Dr Pradeep Mahajan, Regenerative Medicine Researcher, StemRx Bioscience Solutions highlights the importance and other aspects of stem cell technology

Globally, we are seeing a change in the type of age-specific, chronic, debilitating diseases. Thus, the manner in which we diagnose and treat such diseases is also seeing a paradigm shift. From empirical use of drugs to target-specific treatments, we are now advancing towards molecular dysfunction-based therapies.

I have been in the field of clinical medicine and surgery for over 3 decades now and I have always been fascinated by new research. Among the substantial advances in the healthcare field, I believe regenerative medicine and cell-based therapy have been game changers. We saw hematopoietic stem cells being used to treat blood cancers and related diseases for over 3-4 decades. Now we are seeing an expansion in the applications of stem cells in treating various acute, chronic, lifestyle, and even genetic and congenital diseases. The need arose because conventional medicine is gradually losing potency in treating diseases and patients are often left at the mercy of nature to take its course.

With increasing knowledge of stem cells, the trend to utilise the endogenous repair mechanisms of the human body gained popularity. Cells, growth factors and other biological products, when present at the right site; at the right moment, stimulate the natural healing mechanisms of the body and aid in management of health conditions. Cell-based therapy thus marked the beginning of a new era in regenerative medicine.

Stem cells are present in several tissues, namely, embryo, umbilical cord, placenta, as well as adult body tissues. These are the master cells of the body that have roles in development of the body, repairing and regenerating injured tissues (at a cellular level), and maintaining homeostasis even in an healthy individual. Of course, we have all heard of ethical issues regarding the use of embryonic stem cells as well as their tumor-forming issue. Regarding umbilical cord stem cells, the trend of banking this tissue has just begun; therefore, the majority of us would not have the umbilical cord as a source of stem cells. Keeping in mind these aspects, researchers started focusing on adult stem cells that can be derived from different tissues of the human body. The common sources are bone marrow, fat tissue, peripheral blood, and teeth, among others. The chief advantage is that, the source being autologous, the therapy is safe and is not associated with side effects.

Coming to the diseases that can be treated using stem cellswe have just scratched the tip of the iceberg. There are several health conditions that plague mankindarthritis, diabetes, nerve-related conditions, traumatic injuries, etc. Conventionally, one would be prescribed medications (often for prolong periods or even for their lifetime) or be advised surgery. Nonetheless, in several cases, the quality of life of a patient is compromised. The various properties of stem cells help reduce swelling in the body, regulate the immune system, enhance the functioning of other cells, and create a healthy environment for health cells to thrive. Through this, one can target a myriad of pathologies at the molecular level, in a minimally/non-invasive manner.

Patients today are quite aware of the benefits of regenerative medicine and cell based therapy, but there is still a long distance to cover. Countries are promoting research and development in the field of regenerative medicine and cell-based therapy. Research advances pertaining to introducing products with cell and scaffold based technology through tissue engineering are underway. Bioactive scaffolds that are capable of supporting activation and differentiation of host stem cells at the required site are being developed. In the future, it will be possible to use human native sites as micro-niche/micro-environment for potentiation of the human bodys site-specific response. Another breakthrough in the field of cell-based therapy is immunotherapy that aims to utilise certain parts of a persons immune system and stimulate them to fight diseases such as cancer.

The scope of cell-based therapy is endless. All we need is more research, awareness, and implementation to permit reach of the treatment to every stratum of the society. Soon, we will talk about treating diseases with cells and not pills and knives!

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Importance of stem cells-Past, present and future - Express Healthcare

Advance Market Analytics published a new research publication on Stem Cells Market Insights, to 2026 with 232 pages and enriched with self-explained Tables and charts in presentable format. In the Study you will find new evolving Trends, Drivers, Restraints, Opportunities generated by targeting market associated stakeholders. The growth of the Stem Cells Market was mainly driven by the increasing R&D spending across the world.

Some of the key players profiled in the study are:

Smith & Nephew (United Kingdom),Celgene Corporation (United States),BIOTIME, INC. (United States),Cynata (Australia),Applied Cell Technology (Hungary),STEMCELL Technologies Inc. (Canada),BioTime Inc. (United States),Cytori Therapeutics, Inc. (United States),Astellas Pharma Inc. (Japan),U.S. Stem Cell, Inc. (United States),Takara Holdings. (Japan)

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Scope of the Report of Stem Cells

The stem cell is used for treating chronic diseases such as cardiovascular disorders, cancer, diabetes, and others. Growing research and development in stem cell isolation techniques propelling market growth. For instance, a surgeon from Turkey developed a method for obtaining stem cells from the human body without enzymes which are generally used for the isolation of stem cells. Further, growing healthcare infrastructure in the developing economies and government spending on the life science research and development expected to drive the demand for stem cell market over the forecasted period.

Market Trend:

Emphasizing On Development of Regenerative Medicine

Technological Advancement in Stem Cell Harvesting and Isolation Techniques

Market Drivers:

Rising Prevalence of Chronic Diseases such as Cardiovascular Disorders, Cancer, and others

Growing Healthcare Infrastructure in the Developing Economies

Challenges:

Lack of Awareness Regarding Stem Cell Therapy in the Low and Middle Income Group Countries

Opportunities:

Growing Demand for Cellular Therapies

Rising Application of Autologous Therapy

The titled segments and sub-section of the market are illuminated below:by Type (Adult Stem Cells (Neuronal, Hematopoietic, Mesenchymal, Umbilical Cord, Others), Human Embryonic Stem Cells (hESC), Induced Pluripotent Stem Cells, Very Small Embryonic-Like Stem Cells), Application (Regenerative Medicine (Neurology, Orthopedics, Oncology, Hematology, Cardiovascular and Myocardial Infraction, Injuries, Diabetes, Liver Disorder, Incontinence, Others), Drug Discovery and Development), Technology (Cell Acquisition (Bone Marrow Harvest, Umbilical Blood Cord, Apheresis), Cell Production (Therapeutic Cloning, In-vitro Fertilization, Cell Culture, Isolation), Cryopreservation, Expansion and Sub-Culture), Therapy (Autologous, Allogeneic)

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Region Included are: North America, Europe, Asia Pacific, Oceania, South America, Middle East & Africa

Country Level Break-Up: United States, Canada, Mexico, Brazil, Argentina, Colombia, Chile, South Africa, Nigeria, Tunisia, Morocco, Germany, United Kingdom (UK), the Netherlands, Spain, Italy, Belgium, Austria, Turkey, Russia, France, Poland, Israel, United Arab Emirates, Qatar, Saudi Arabia, China, Japan, Taiwan, South Korea, Singapore, India, Australia and New Zealand etc.

Strategic Points Covered in Table of Content of Global Stem Cells Market:

Chapter 1: Introduction, market driving force product Objective of Study and Research Scope the Stem Cells market

Chapter 2: Exclusive Summary the basic information of the Stem Cells Market.

Chapter 3: Displaying the Market Dynamics- Drivers, Trends and Challenges of the Stem Cells

Chapter 4: Presenting the Stem Cells Market Factor Analysis Porters Five Forces, Supply/Value Chain, PESTEL analysis, Market Entropy, Patent/Trademark Analysis.

Chapter 5: Displaying market size by Type, End User and Region 2015-2020

Chapter 6: Evaluating the leading manufacturers of the Stem Cells market which consists of its Competitive Landscape, Peer Group Analysis, BCG Matrix & Company Profile

Chapter 7: To evaluate the market by segments, by countries and by manufacturers with revenue share and sales by key countries (2021-2026).

Chapter 8 & 9: Displaying the Appendix, Methodology and Data Source

Finally, Stem Cells Market is a valuable source of guidance for individuals and companies in decision framework.

Read Detailed Index of full Research Study at @ https://www.advancemarketanalytics.com/reports/72815-global-stem-cells-market-1

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Stem Cells Market to Witness Gigantic Growth by 2026 LSMedia - LSMedia

PALM BEACH, Fla., Dec. 21, 2021 /PRNewswire/ -- FinancialNewsMedia.com News Commentary - Systemic mastocytosis is rare disease which affects very few people and it causes due to C-kit mutation which leads to higher number of mast cell production in the body resulting in accumulation of excessive mast cells in the internal body organs such as spleen, liver, bone marrow and small intestine etc. Recently, the World Health Organization (WHO) updated the prognosis, diagnosis and systemic mastocytosis treatment guidelines for the disease which in turn helped to bring uniformity in the approach by healthcare professionals. The manufacturers in the systemic mastocytosis treatment market are focusing on evaluating possible treatment options to cure the disease by investing heavily in the research & development. Various leading manufacturers are focusing on gaining FDA approval to respective drugs for the systemic mastocytosis treatment to enhance their revenue generation. A reportfrom Future Market Insights said:"Increasing awareness about the systemic mastocytosis treatment as well as symptoms of the disease due to extended effort by non-profit organizations, governmental associations and through other platforms expected to drive the growth of the systemic mastocytosis treatment market Increasing approvals and launches of drugs for the systemic mastocytosis treatment expected to drive the growth of the market Increasing spending on research & development by various pharmaceutical companies to develop novel systemic mastocytosis treatment expected to further fuel the growth of market. Increasing early diagnosis rate subsequently followed by increasing treatment seeking rate further expected to drive the growth of the systemic mastocytosis treatment market." Active companies in the markets today include: Hoth Therapeutics, Inc. (NASDAQ:HOTH), Longeveron Inc. (NASDAQ: LGVN), Bristol Myers Squibb (NYSE: BMY), Takeda Pharmaceutical Company Limited (NYSE: TAK), Amgen (NASDAQ: AMGN).

Future Market Insights continued:"The global systemic mastocytosis treatment market is expected to experience significant growth due to increasing novel treatment options. By drug class, systemic mastocytosis treatment market is expected to be dominated by the mast cell stabilizers due to superior efficacy. By indication, systemic mastocytosis treatment market is expected to be dominated by indolent systemic mastocytosis (ISM) due to higher prevalence. By route o administration, systemic mastocytosis treatment market is expected to be dominated by injectables. By distribution channel, systemic mastocytosis treatment market is expected to be dominated by the retail pharmacies due to higher patient footfall. The global systemic mastocytosis treatment market is expected to be dominated by the North America due to comparatively higher prevalence of the disease. Europe systemic mastocytosis treatment market is expected to be second most lucrative market due to higher treatment seeking rate. Latin America expected to show gradual growth in the systemic mastocytosis treatment market due to steadily increasing diagnosis. Asia-Pacific is emerging systemic mastocytosis treatment market due to increasing diagnosis subsequently followed by treatment. Middle East & Africa is the least lucrative systemic mastocytosis treatment market due to least diagnostic rate and lower awareness about the symptoms."

Hoth Therapeutics, Inc. (NASDAQ:HOTH) BREAKING NEWS: Hoth Therapeutics Announces Submission of Orphan Designation Application for HT-KIT to Treat Mastocytosis Hoth Therapeutics, Inc., a patient-focusedclinical-stage biopharmaceutical company, announced it submitted an Orphan Drug Designation Application to the US Food and Drug Administration (FDA) for HT-KIT for the treatment of mastocyctosis. HT-KIT is an antisense oligonucleotide that targets the proto-oncogene cKIT by inducing mRNA frame shifting, resulting in apoptosis of neoplastic mast cells. The KIT signaling pathway is implicated in multiple diseases, including all types of mastocytosis (such as aggressive systemic mastocytosis (ASM), mast cell leukemia (MCL), and systemic mastocytosis with associated hematological neoplasm (SM-AHN)), acute myeloid leukemia, gastrointestinal stromal tumors, and anaphylaxis.

Drugs intended to treat orphan diseases (rare diseases that affect less than 200,000 people in the US)are eligible to apply for Orphan Drug Designation (ODD), which provides multiple benefits to the sponsor during development and after approval. Hoth intends to pursue these benefits as part of the drug development for HT-KIT for treatment of mastocytosis, pending designation of the ODD application.

Benefits of Orphan Drug Designation - Under the Orphan Drug Act, drug companies can apply for ODD, and if granted, the drug will have a status which gives companies exclusive marketing and development rights along with other benefits to recover the costs of researching and developing the drug. A tax credit of 50% of the qualified clinical drug testing costs awarded upon drug approval is also possible. Regulatory streamlining and provide special assistance to companies that develop drugs for rare patient populations. In addition to exclusive rights and cost benefits, the FDA will provide protocol assistance, potential decreased wait-time for drug approval, discounts on registration fees, and eligibility for market exclusivity after approval.

Key benefits of ODD:

Hoth recently announcedthat its novelanti-cancer therapeuticexhibited highly positive results in humanized mast cell neoplasm models, representative in vitro and in vivo models for aggressive, mast cell-derived cancers such as mast cell leukemia and mast cell sarcoma. CONTINUED Read the Hoth Therapeutics full press release by going to: https://ir.hoththerapeutics.com/news-releases

In other news and developments of note in the markets this week:

Amgen (NASDAQ: AMGN) recently announced that the U.S. Food and Drug Administration (FDA) has approved Amgen and AstraZeneca'sTezspire (tezepelumab-ekko) for the add-on maintenance treatment of adult and pediatric patients aged 12 years and older with severe asthma.

Tezspirewas approved following a Priority Review by the FDA and based on results from the PATHFINDER clinical trial program. The application included results from the pivotal NAVIGATOR Phase 3 trial in whichTezspiredemonstrated superiority across every primary and key secondary endpoint in patients with severe asthma, compared to placebo, when added to standard therapy.

Longeveron Inc. (NASDAQ: LGVN), a clinical stage biotechnology company developing cellular therapies for chronic aging-related and certain life-threatening conditions, recently announced that the U.S. Food and Drug Administration (FDA) has granted Orphan Drug Designation (ODD) for Lomecel-B for the treatment of Hypoplastic Left Heart Syndrome (HLHS), a rare and life-threatening congenital heart defect in infants.

ODD is intended to assist and encourage companies to develop safe and effective therapies for the treatment of rare diseases or conditions. ODD positions Longeveron to be able to potentially leverage a range of financial and regulatory benefits, including government grants for conducting clinical trials, waiver of FDA user fees for the potential submission of a marketing application, and certain tax credits. Receiving ODD may also result in the product receiving seven years market exclusivity upon approval for use in the rare disease or condition for which the product was designated if all of the statutory and regulatory requirements are met.

Bristol Myers Squibb (NYSE: BMY) recently announced thatOrencia(abatacept) was approved by the U.S. Food and Drug Administration (FDA) for the prophylaxis, or prevention, of acute graft versus host disease (aGvHD), in combination with a calcineurin inhibitor (CNI) and methotrexate (MTX), in adults and pediatric patients 2 years of age and older undergoing hematopoietic stem cell transplantation (HSCT) from a matched or 1 allele-mismatched unrelated donor (URD).

"Orenciais the first FDA-approved therapy to prevent acute graft versus host disease following hematopoietic stem cell transplant, a potentially life-threatening complication that can pose a comparatively higher risk to racial and ethnic minority populations in the U.S. due to difficulty finding appropriately matched donors," said Tina Deignan, senior vice president, U.S. Immunology, Bristol Myers Squibb. "With this fourth indication forOrencia,Bristol Myers Squibb draws on its legacy and expertise in both immunology and hematology to deliver an important treatment option for patients in a disease with high unmet need.

Takeda Pharmaceutical Company Limited (NYSE: TAK) announced that the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA) has recommended the approval of intravenous (IV) vedolizumab for the treatment of adult patients with moderately to severely active chronic pouchitis, who have undergone proctocolectomy and ileal pouch-anal anastomosis (IPAA) for ulcerative colitis (UC), and have had an inadequate response with or lost response to antibiotic therapy. The CHMP opinion will now be reviewed by the European Commission. If approved, vedolizumab will become the first treatment indicated for active chronic pouchitis across the European Union.

DISCLAIMER: FN Media Group LLC (FNM), which owns and operates Financialnewsmedia.com and MarketNewsUpdates.com, is a third- party publisher and news dissemination service provider, which disseminates electronic information through multiple online media channels. FNM is NOT affiliated in any manner with any company mentioned herein. FNM and its affiliated companies are a news dissemination solutions provider and are NOT a registered broker/dealer/analyst/adviser, holds no investment licenses and may NOT sell, offer to sell or offer to buy any security. FNM's market updates, news alerts and corporate profiles are NOT a solicitation or recommendation to buy, sell or hold securities. The material in this release is intended to be strictly informational and is NEVER to be construed or interpreted as research material. All readers are strongly urged to perform research and due diligence on their own and consult a licensed financial professional before considering any level of investing in stocks. All material included herein is republished content and details which were previously disseminated by the companies mentioned in this release. FNM is not liable for any investment decisions by its readers or subscribers. Investors are cautioned that they may lose all or a portion of their investment when investing in stocks. For current services performed FNM was compensated twenty five hundred dollars for news coverage of current press release issued by: Hoth Therapeutics, Inc. by a non-affiliated third party.

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Systemic Mastocytosis Treatments Gain Hope Due To Increasing Novel Treatment Options - PRNewswire

Int J Cell Biol. 2016; 2016: 6940283.

Department of Biological Sciences, Indian Institute of Science Education and Research (IISER), Bhopal, Madhya Pradesh 462066, India

Department of Biological Sciences, Indian Institute of Science Education and Research (IISER), Bhopal, Madhya Pradesh 462066, India

Academic Editor: Paul J. Higgins

Received 2016 Mar 13; Accepted 2016 Jun 5.

This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Regenerative medicine, the most recent and emerging branch of medical science, deals with functional restoration of tissues or organs for the patient suffering from severe injuries or chronic disease. The spectacular progress in the field of stem cell research has laid the foundation for cell based therapies of disease which cannot be cured by conventional medicines. The indefinite self-renewal and potential to differentiate into other types of cells represent stem cells as frontiers of regenerative medicine. The transdifferentiating potential of stem cells varies with source and according to that regenerative applications also change. Advancements in gene editing and tissue engineering technology have endorsed the ex vivo remodelling of stem cells grown into 3D organoids and tissue structures for personalized applications. This review outlines the most recent advancement in transplantation and tissue engineering technologies of ESCs, TSPSCs, MSCs, UCSCs, BMSCs, and iPSCs in regenerative medicine. Additionally, this review also discusses stem cells regenerative application in wildlife conservation.

Regenerative medicine, the most recent and emerging branch of medical science, deals with functional restoration of specific tissue and/or organ of the patients suffering with severe injuries or chronic disease conditions, in the state where bodies own regenerative responses do not suffice [1]. In the present scenario donated tissues and organs cannot meet the transplantation demands of aged and diseased populations that have driven the thrust for search for the alternatives. Stem cells are endorsed with indefinite cell division potential, can transdifferentiate into other types of cells, and have emerged as frontline regenerative medicine source in recent time, for reparation of tissues and organs anomalies occurring due to congenital defects, disease, and age associated effects [1]. Stem cells pave foundation for all tissue and organ system of the body and mediates diverse role in disease progression, development, and tissue repair processes in host. On the basis of transdifferentiation potential, stem cells are of four types, that is, (1) unipotent, (2) multipotent, (3) pluripotent, and (4) totipotent [2]. Zygote, the only totipotent stem cell in human body, can give rise to whole organism through the process of transdifferentiation, while cells from inner cells mass (ICM) of embryo are pluripotent in their nature and can differentiate into cells representing three germ layers but do not differentiate into cells of extraembryonic tissue [2]. Stemness and transdifferentiation potential of the embryonic, extraembryonic, fetal, or adult stem cells depend on functional status of pluripotency factors like OCT4, cMYC, KLF44, NANOG, SOX2, and so forth [35]. Ectopic expression or functional restoration of endogenous pluripotency factors epigenetically transforms terminally differentiated cells into ESCs-like cells [3], known as induced pluripotent stem cells (iPSCs) [3, 4]. On the basis of regenerative applications, stem cells can be categorized as embryonic stem cells (ESCs), tissue specific progenitor stem cells (TSPSCs), mesenchymal stem cells (MSCs), umbilical cord stem cells (UCSCs), bone marrow stem cells (BMSCs), and iPSCs (; ). The transplantation of stem cells can be autologous, allogenic, and syngeneic for induction of tissue regeneration and immunolysis of pathogen or malignant cells. For avoiding the consequences of host-versus-graft rejections, tissue typing of human leucocyte antigens (HLA) for tissue and organ transplant as well as use of immune suppressant is recommended [6]. Stem cells express major histocompatibility complex (MHC) receptor in low and secret chemokine that recruitment of endothelial and immune cells is enabling tissue tolerance at graft site [6]. The current stem cell regenerative medicine approaches are founded onto tissue engineering technologies that combine the principles of cell transplantation, material science, and microengineering for development of organoid; those can be used for physiological restoration of damaged tissue and organs. The tissue engineering technology generates nascent tissue on biodegradable 3D-scaffolds [7, 8]. The ideal scaffolds support cell adhesion and ingrowths, mimic mechanics of target tissue, support angiogenesis and neovascularisation for appropriate tissue perfusion, and, being nonimmunogenic to host, do not require systemic immune suppressant [9]. Stem cells number in tissue transplant impacts upon regenerative outcome [10]; in that case prior ex vivo expansion of transplantable stem cells is required [11]. For successful regenerative outcomes, transplanted stem cells must survive, proliferate, and differentiate in site specific manner and integrate into host circulatory system [12]. This review provides framework of most recent (; Figures ) advancement in transplantation and tissue engineering technologies of ESCs, TSPSCs, MSCs, UCSCs, BMSCs, and iPSCs in regenerative medicine. Additionally, this review also discusses stem cells as the tool of regenerative applications in wildlife conservation.

Promises of stem cells in regenerative medicine: the six classes of stem cells, that is, embryonic stem cells (ESCs), tissue specific progenitor stem cells (TSPSCs), mesenchymal stem cells (MSCs), umbilical cord stem cells (UCSCs), bone marrow stem cells (BMSCs), and induced pluripotent stem cells (iPSCs), have many promises in regenerative medicine and disease therapeutics.

ESCs in regenerative medicine: ESCs, sourced from ICM of gastrula, have tremendous promises in regenerative medicine. These cells can differentiate into more than 200 types of cells representing three germ layers. With defined culture conditions, ESCs can be transformed into hepatocytes, retinal ganglion cells, chondrocytes, pancreatic progenitor cells, cone cells, cardiomyocytes, pacemaker cells, eggs, and sperms which can be used in regeneration of tissue and treatment of disease in tissue specific manner.

TSPSCs in regenerative medicine: tissue specific stem and progenitor cells have potential to differentiate into other cells of the tissue. Characteristically inner ear stem cells can be transformed into auditory hair cells, skin progenitors into vascular smooth muscle cells, mesoangioblasts into tibialis anterior muscles, and dental pulp stem cells into serotonin cells. The 3D-culture of TSPSCs in complex biomaterial gives rise to tissue organoids, such as pancreatic organoid from pancreatic progenitor, intestinal tissue organoids from intestinal progenitor cells, and fallopian tube organoids from fallopian tube epithelial cells. Transplantation of TSPSCs regenerates targets tissue such as regeneration of tibialis muscles from mesoangioblasts, cardiac tissue from AdSCs, and corneal tissue from limbal stem cells. Cell growth and transformation factors secreted by TSPSCs can change cells fate to become other types of cell, such that SSCs coculture with skin, prostate, and intestine mesenchyme transforms these cells from MSCs into epithelial cells fate.

MSCs in regenerative medicine: mesenchymal stem cells are CD73+, CD90+, CD105+, CD34, CD45, CD11b, CD14, CD19, and CD79a cells, also known as stromal cells. These bodily MSCs represented here do not account for MSCs of bone marrow and umbilical cord. Upon transplantation and transdifferentiation these bodily MSCs regenerate into cartilage, bones, and muscles tissue. Heart scar formed after heart attack and liver cirrhosis can be treated from MSCs. ECM coating provides the niche environment for MSCs to regenerate into hair follicle, stimulating hair growth.

UCSCs in regenerative medicine: umbilical cord, the readily available source of stem cells, has emerged as futuristic source for personalized stem cell therapy. Transplantation of UCSCs to Krabbe's disease patients regenerates myelin tissue and recovers neuroblastoma patients through restoring tissue homeostasis. The UCSCs organoids are readily available tissue source for treatment of neurodegenerative disease. Peritoneal fibrosis caused by long term dialysis, tendon tissue degeneration, and defective hyaline cartilage can be regenerated by UCSCs. Intravenous injection of UCSCs enables treatment of diabetes, spinal myelitis, systemic lupus erythematosus, Hodgkin's lymphoma, and congenital neuropathies. Cord blood stem cells banking avails long lasting source of stem cells for personalized therapy and regenerative medicine.

BMSCs in regenerative medicine: bone marrow, the soft sponge bone tissue that consisted of stromal, hematopoietic, and mesenchymal and progenitor stem cells, is responsible for blood formation. Even halo-HLA matched BMSCs can cure from disease and regenerate tissue. BMSCs can regenerate craniofacial tissue, brain tissue, diaphragm tissue, and liver tissue and restore erectile function and transdifferentiation monocytes. These multipotent stem cells can cure host from cancer and infection of HIV and HCV.

iPSCs in regenerative medicine: using the edge of iPSCs technology, skin fibroblasts and other adult tissues derived, terminally differentiated cells can be transformed into ESCs-like cells. It is possible that adult cells can be transformed into cells of distinct lineages bypassing the phase of pluripotency. The tissue specific defined culture can transform skin cells to become trophoblast, heart valve cells, photoreceptor cells, immune cells, melanocytes, and so forth. ECM complexation with iPSCs enables generation of tissue organoids for lung, kidney, brain, and other organs of the body. Similar to ESCs, iPSCs also can be transformed into cells representing three germ layers such as pacemaker cells and serotonin cells.

Stem cells in wildlife conservation: tissue biopsies obtained from dead and live wild animals can be either cryopreserved or transdifferentiated to other types of cells, through culture in defined culture medium or in vivo maturation. Stem cells and adult tissue derived iPSCs have great potential of regenerative medicine and disease therapeutics. Gonadal tissue procured from dead wild animals can be matured, ex vivo and in vivo for generation of sperm and egg, which can be used for assistive reproductive technology oriented captive breeding of wild animals or even for resurrection of wildlife.

Application of stem cells in regenerative medicine: stem cells (ESCs, TSPSCs, MSCs, UCSCs, BMSCs, and iPSCs) have diverse applications in tissue regeneration and disease therapeutics.

For the first time in 1998, Thomson isolated human ESCs (hESCs) [13]. ESCs are pluripotent in their nature and can give rise to more than 200 types of cells and promises for the treatment of any kinds of disease [13]. The pluripotency fate of ESCs is governed by functional dynamics of transcription factors OCT4, SOX2, NANOG, and so forth, which are termed as pluripotency factors. The two alleles of the OCT4 are held apart in pluripotency state in ESCs; phase through homologues pairing during embryogenesis and transdifferentiation processes [14] has been considered as critical regulatory switch for lineage commitment of ESCs. The diverse lineage commitment potential represents ESCs as ideal model for regenerative therapeutics of disease and tissue anomalies. This section of review on ESCs discusses transplantation and transdifferentiation of ESCs into retinal ganglion, hepatocytes, cardiomyocytes, pancreatic progenitors, chondrocytes, cones, egg sperm, and pacemaker cells (; ). Infection, cancer treatment, and accidents can cause spinal cord injuries (SCIs). The transplantation of hESCs to paraplegic or quadriplegic SCI patients improves body control, balance, sensation, and limbal movements [15], where transplanted stem cells do homing to injury sites. By birth, humans have fixed numbers of cone cells; degeneration of retinal pigment epithelium (RPE) of macula in central retina causes age-related macular degeneration (ARMD). The genomic incorporation of COCO gene (expressed during embryogenesis) in the developing embryo leads lineage commitment of ESCs into cone cells, through suppression of TGF, BMP, and Wnt signalling pathways. Transplantation of these cone cells to eye recovers individual from ARMD phenomenon, where transplanted cone cells migrate and form sheet-like structure in host retina [16]. However, establishment of missing neuronal connection of retinal ganglion cells (RGCs), cones, and PRE is the most challenging aspect of ARMD therapeutics. Recently, Donald Z Jacks group at John Hopkins University School of Medicine has generated RGCs from CRISPER-Cas9-m-Cherry reporter ESCs [17]. During ESCs transdifferentiation process, CRIPER-Cas9 directs the knock-in of m-Cherry reporter into 3UTR of BRN3B gene, which is specifically expressed in RGCs and can be used for purification of generated RGCs from other cells [17]. Furthermore, incorporation of forskolin in transdifferentiation regime boosts generation of RGCs. Coaxing of these RGCs into biomaterial scaffolds directs axonal differentiation of RGCs. Further modification in RGCs generation regime and composition of biomaterial scaffolds might enable restoration of vision for ARMD and glaucoma patients [17]. Globally, especially in India, cardiovascular problems are a more common cause of human death, where biomedical therapeutics require immediate restoration of heart functions for the very survival of the patient. Regeneration of cardiac tissue can be achieved by transplantation of cardiomyocytes, ESCs-derived cardiovascular progenitors, and bone marrow derived mononuclear cells (BMDMNCs); however healing by cardiomyocytes and progenitor cells is superior to BMDMNCs but mature cardiomyocytes have higher tissue healing potential, suppress heart arrhythmias, couple electromagnetically into hearts functions, and provide mechanical and electrical repair without any associated tumorigenic effects [18, 19]. Like CM differentiation, ESCs derived liver stem cells can be transformed into Cytp450-hepatocytes, mediating chemical modification and catabolism of toxic xenobiotic drugs [20]. Even today, availability and variability of functional hepatocytes are a major a challenge for testing drug toxicity [20]. Stimulation of ESCs and ex vivo VitK12 and lithocholic acid (a by-product of intestinal flora regulating drug metabolism during infancy) activates pregnane X receptor (PXR), CYP3A4, and CYP2C9, which leads to differentiation of ESCs into hepatocytes; those are functionally similar to primary hepatocytes, for their ability to produce albumin and apolipoprotein B100 [20]. These hepatocytes are excellent source for the endpoint screening of drugs for accurate prediction of clinical outcomes [20]. Generation of hepatic cells from ESCs can be achieved in multiple ways, as serum-free differentiation [21], chemical approaches [20, 22], and genetic transformation [23, 24]. These ESCs-derived hepatocytes are long lasting source for treatment of liver injuries and high throughput screening of drugs [20, 23, 24]. Transplantation of the inert biomaterial encapsulated hESCs-derived pancreatic progenitors (CD24+, CD49+, and CD133+) differentiates into -cells, minimizing high fat diet induced glycemic and obesity effects in mice [25] (). Addition of antidiabetic drugs into transdifferentiation regime can boost ESCs conservation into -cells [25], which theoretically can cure T2DM permanently [25]. ESCs can be differentiated directly into insulin secreting -cells (marked with GLUT2, INS1, GCK, and PDX1) which can be achieved through PDX1 mediated epigenetic reprogramming [26]. Globally, osteoarthritis affects millions of people and occurs when cartilage at joints wears away, causing stiffness of the joints. The available therapeutics for arthritis relieve symptoms but do not initiate reverse generation of cartilage. For young individuals and athletes replacement of joints is not feasible like old populations; in that case transplantation of stem cells represents an alternative for healing cartilage injuries [27]. Chondrocytes, the cartilage forming cells derived from hESC, embedded in fibrin gel effectively heal defective cartilage within 12 weeks, when transplanted to focal cartilage defects of knee joints in mice without any negative effect [27]. Transplanted chondrocytes form cell aggregates, positive for SOX9 and collagen II, and defined chondrocytes are active for more than 12wks at transplantation site, advocating clinical suitability of chondrocytes for treatment of cartilage lesions [27]. The integrity of ESCs to integrate and differentiate into electrophysiologically active cells provides a means for natural regulation of heart rhythm as biological pacemaker. Coaxing of ESCs into inert biomaterial as well as propagation in defined culture conditions leads to transdifferentiation of ESCs to become sinoatrial node (SAN) pacemaker cells (PCs) [28]. Genomic incorporation TBox3 into ESCs ex vivo leads to generation of PCs-like cells; those express activated leukocyte cells adhesion molecules (ALCAM) and exhibit similarity to PCs for gene expression and immune functions [28]. Transplantation of PCs can restore pacemaker functions of the ailing heart [28]. In summary, ESCs can be transdifferentiated into any kinds of cells representing three germ layers of the body, being most promising source of regenerative medicine for tissue regeneration and disease therapy (). Ethical concerns limit the applications of ESCs, where set guidelines need to be followed; in that case TSPSCs, MSCs, UCSCs, BMSCs, and iPSCs can be explored as alternatives.

TSPSCs maintain tissue homeostasis through continuous cell division, but, unlike ESCs, TSPSCs retain stem cells plasticity and differentiation in tissue specific manner, giving rise to few types of cells (). The number of TSPSCs population to total cells population is too low; in that case their harvesting as well as in vitro manipulation is really a tricky task [29], to explore them for therapeutic scale. Human body has foundation from various types of TSPSCs; discussing the therapeutic application for all types is not feasible. This section of review discusses therapeutic application of pancreatic progenitor cells (PPCs), dental pulp stem cells (DPSCs), inner ear stem cells (IESCs), intestinal progenitor cells (IPCs), limbal progenitor stem cells (LPSCs), epithelial progenitor stem cells (EPSCs), mesoangioblasts (MABs), spermatogonial stem cells (SSCs), the skin derived precursors (SKPs), and adipose derived stem cells (AdSCs) (; ). During embryogenesis PPCs give rise to insulin-producing -cells. The differentiation of PPCs to become -cells is negatively regulated by insulin [30]. PPCs require active FGF and Notch signalling; growing more rapidly in community than in single cell populations advocates the functional importance of niche effect in self-renewal and transdifferentiation processes. In 3D-scaffold culture system, mice embryo derived PPCs grow into hollow organoid spheres; those finally differentiate into insulin-producing -cell clusters [29]. The DSPSCs, responsible for maintenance of teeth health status, can be sourced from apical papilla, deciduous teeth, dental follicle, and periodontal ligaments, have emerged as regenerative medicine candidate, and might be explored for treatment of various kinds of disease including restoration neurogenic functions in teeth [31, 32]. Expansion of DSPSCs in chemically defined neuronal culture medium transforms them into a mixed population of cholinergic, GABAergic, and glutaminergic neurons; those are known to respond towards acetylcholine, GABA, and glutamine stimulations in vivo. These transformed neuronal cells express nestin, glial fibrillary acidic protein (GFAP), III-tubulin, and voltage gated L-type Ca2+ channels [32]. However, absence of Na+ and K+ channels does not support spontaneous action potential generation, necessary for response generation against environmental stimulus. All together, these primordial neuronal stem cells have possible therapeutic potential for treatment of neurodental problems [32]. Sometimes, brain tumor chemotherapy can cause neurodegeneration mediated cognitive impairment, a condition known as chemobrain [33]. The intrahippocampal transplantation of human derived neuronal stem cells to cyclophosphamide behavioural decremented mice restores cognitive functions in a month time. Here the transplanted stem cells differentiate into neuronal and astroglial lineage, reduce neuroinflammation, and restore microglial functions [33]. Furthermore, transplantation of stem cells, followed by chemotherapy, directs pyramidal and granule-cell neurons of the gyrus and CA1 subfields of hippocampus which leads to reduction in spine and dendritic cell density in the brain. These findings suggest that transplantation of stem cells to cranium restores cognitive functions of the chemobrain [33]. The hair cells of the auditory system produced during development are not postmitotic; loss of hair cells cannot be replaced by inner ear stem cells, due to active state of the Notch signalling [34]. Stimulation of inner ear progenitors with -secretase inhibitor ({"type":"entrez-nucleotide","attrs":{"text":"LY411575","term_id":"1257853995","term_text":"LY411575"}}LY411575) abrogates Notch signalling through activation of transcription factor atonal homologue 1 (Atoh1) and directs transdifferentiation of progenitors into cochlear hair cells [34]. Transplantation of in vitro generated hair cells restores acoustic functions in mice, which can be the potential regenerative medicine candidates for the treatment of deafness [34]. Generation of the hair cells also can be achieved through overexpression of -catenin and Atoh1 in Lrg5+ cells in vivo [35]. Similar to ear progenitors, intestine of the digestive tract also has its own tissue specific progenitor stem cells, mediating regeneration of the intestinal tissue [34, 36]. Dysregulation of the common stem cells signalling pathways, Notch/BMP/TGF-/Wnt, in the intestinal tissue leads to disease. Information on these signalling pathways [37] is critically important in designing therapeutics. Coaxing of the intestinal tissue specific progenitors with immune cells (macrophages), connective tissue cells (myofibroblasts), and probiotic bacteria into 3D-scaffolds of inert biomaterial, crafting biological environment, is suitable for differentiation of progenitors to occupy the crypt-villi structures into these scaffolds [36]. Omental implementation of these crypt-villi structures to dogs enhances intestinal mucosa through regeneration of goblet cells containing intestinal tissue [36]. These intestinal scaffolds are close approach for generation of implantable intestinal tissue, divested by infection, trauma, cancer, necrotizing enterocolitis (NEC), and so forth [36]. In vitro culture conditions cause differentiation of intestinal stem cells to become other types of cells, whereas incorporation of valproic acid and CHIR-99021 in culture conditions avoids differentiation of intestinal stem cells, enabling generation of indefinite pool of stem cells to be used for regenerative applications [38]. The limbal stem cells of the basal limbal epithelium, marked with ABCB5, are essential for regeneration and maintenance of corneal tissue [39]. Functional status of ABCB5 is critical for survival and functional integrity of limbal stem cells, protecting them from apoptotic cell death [39]. Limbal stem cells deficiency leads to replacement of corneal epithelium with visually dead conjunctival tissue, which can be contributed by burns, inflammation, and genetic factors [40]. Transplanted human cornea stem cells to mice regrown into fully functional human cornea, possibly supported by blood eye barrier phenomena, can be used for treatment of eye diseases, where regeneration of corneal tissue is critically required for vision restoration [39]. Muscle degenerative disease like duchenne muscular dystrophy (DMD) can cause extensive thrashing of muscle tissue, where tissue engineering technology can be deployed for functional restoration of tissue through regeneration [41]. Encapsulation of mouse or human derived MABs (engineered to express placental derived growth factor (PDGF)) into polyethylene glycol (PEG) fibrinogen hydrogel and their transplantation beneath the skin at ablated tibialis anterior form artificial muscles, which are functionally similar to those of normal tibialis anterior muscles [41]. The PDGF attracts various cell types of vasculogenic and neurogenic potential to the site of transplantation, supporting transdifferentiation of mesoangioblasts to become muscle fibrils [41]. The therapeutic application of MABs in skeletal muscle regeneration and other therapeutic outcomes has been reviewed by others [42]. One of the most important tissue specific stem cells, the male germline stem cells or spermatogonial stem cells (SSCs), produces spermatogenic lineage through mesenchymal and epithets cells [43] which itself creates niche effect on other cells. In vivo transplantation of SSCs with prostate, skin, and uterine mesenchyme leads to differentiation of these cells to become epithelia of the tissue of origin [43]. These newly formed tissues exhibit all physical and physiological characteristics of prostate and skin and the physical characteristics of prostate, skin, and uterus, express tissue specific markers, and suggest that factors secreted from SSCs lead to lineage conservation which defines the importance of niche effect in regenerative medicine [43]. According to an estimate, more than 100 million people are suffering from the condition of diabetic retinopathy, a progressive dropout of vascularisation in retina that leads to loss of vision [44]. The intravitreal injection of adipose derived stem cells (AdSCs) to the eye restores microvascular capillary bed in mice. The AdSCs from healthy donor produce higher amounts of vasoprotective factors compared to glycemic mice, enabling superior vascularisation [44]. However use of AdSCs for disease therapeutics needs further standardization for cell counts in dose of transplant and monitoring of therapeutic outcomes at population scale [44]. Apart from AdSCs, other kinds of stem cells also have therapeutic potential in regenerative medicine for treatment of eye defects, which has been reviewed by others [45]. Fallopian tubes, connecting ovaries to uterus, are the sites where fertilization of the egg takes place. Infection in fallopian tubes can lead to inflammation, tissue scarring, and closure of the fallopian tube which often leads to infertility and ectopic pregnancies. Fallopian is also the site where onset of ovarian cancer takes place. The studies on origin and etiology of ovarian cancer are restricted due to lack of technical advancement for culture of epithelial cells. The in vitro 3D organoid culture of clinically obtained fallopian tube epithelial cells retains their tissue specificity, keeps cells alive, which differentiate into typical ciliated and secretory cells of fallopian tube, and advocates that ectopic examination of fallopian tube in organoid culture settings might be the ideal approach for screening of cancer [46]. The sustained growth and differentiation of fallopian TSPSCs into fallopian tube organoid depend both on the active state of the Wnt and on paracrine Notch signalling [46]. Similar to fallopian tube stem cells, subcutaneous visceral tissue specific cardiac adipose (CA) derived stem cells (AdSCs) have the potential of differentiation into cardiovascular tissue [47]. Systemic infusion of CA-AdSCs into ischemic myocardium of mice regenerates heart tissue and improves cardiac function through differentiation to endothelial cells, vascular smooth cells, and cardiomyocytes and vascular smooth cells. The differentiation and heart regeneration potential of CA-AdSCs are higher than AdSCs [48], representing CA-AdSCs as potent regenerative medicine candidates for myocardial ischemic therapy [47]. The skin derived precursors (SKPs), the progenitors of dermal papilla/hair/hair sheath, give rise to multiple tissues of mesodermal and/or ectodermal origin such as neurons, Schwann cells, adipocytes, chondrocytes, and vascular smooth muscle cells (VSMCs). VSMCs mediate wound healing and angiogenesis process can be derived from human foreskin progenitor SKPs, suggesting that SKPs derived VSMCs are potential regenerative medicine candidates for wound healing and vasculature injuries treatments [49]. In summary, TSPSCs are potentiated with tissue regeneration, where advancement in organoid culture (; ) technologies defines the importance of niche effect in tissue regeneration and therapeutic outcomes of ex vivo expanded stem cells.

MSCs, the multilineage stem cells, differentiate only to tissue of mesodermal origin, which includes tendons, bone, cartilage, ligaments, muscles, and neurons [50]. MSCs are the cells which express combination of markers: CD73+, CD90+, CD105+, CD11b, CD14, CD19, CD34, CD45, CD79a, and HLA-DR, reviewed elsewhere [50]. The application of MSCs in regenerative medicine can be generalized from ongoing clinical trials, phasing through different state of completions, reviewed elsewhere [90]. This section of review outlines the most recent representative applications of MSCs (; ). The anatomical and physiological characteristics of both donor and receiver have equal impact on therapeutic outcomes. The bone marrow derived MSCs (BMDMSCs) from baboon are morphologically and phenotypically similar to those of bladder stem cells and can be used in regeneration of bladder tissue. The BMDMSCs (CD105+, CD73+, CD34, and CD45), expressing GFP reporter, coaxed with small intestinal submucosa (SIS) scaffolds, augment healing of degenerated bladder tissue within 10wks of the transplantation [51]. The combinatorial CD characterized MACs are functionally active at transplantation site, which suggests that CD characterization of donor MSCs yields superior regenerative outcomes [51]. MSCs also have potential to regenerate liver tissue and treat liver cirrhosis, reviewed elsewhere [91]. The regenerative medicinal application of MSCs utilizes cells in two formats as direct transplantation or first transdifferentiation and then transplantation; ex vivo transdifferentiation of MSCs deploys retroviral delivery system that can cause oncogenic effect on cells. Nonviral, NanoScript technology, comprising utility of transcription factors (TFs) functionalized gold nanoparticles, can target specific regulatory site in the genome effectively and direct differentiation of MSCs into another cell fate, depending on regime of TFs. For example, myogenic regulatory factor containing NanoScript-MRF differentiates the adipose tissue derived MSCs into muscle cells [92]. The multipotency characteristics represent MSCs as promising candidate for obtaining stable tissue constructs through coaxed 3D organoid culture; however heterogeneous distribution of MSCs slows down cell proliferation, rendering therapeutic applications of MSCs. Adopting two-step culture system for MSCs can yield homogeneous distribution of MSCs in biomaterial scaffolds. For example, fetal-MSCs coaxed in biomaterial when cultured first in rotating bioreactor followed with static culture lead to homogeneous distribution of MSCs in ECM components [7]. Occurrence of dental carries, periodontal disease, and tooth injury can impact individual's health, where bioengineering of teeth can be the alternative option. Coaxing of epithelial-MSCs with dental stem cells into synthetic polymer gives rise to mature teeth unit, which consisted of mature teeth and oral tissue, offering multiple regenerative therapeutics, reviewed elsewhere [52]. Like the tooth decay, both human and animals are prone to orthopedic injuries, affecting bones, joint, tendon, muscles, cartilage, and so forth. Although natural healing potential of bone is sufficient to heal the common injuries, severe trauma and tumor-recession can abrogate germinal potential of bone-forming stem cells. In vitro chondrogenic, osteogenic, and adipogenic potential of MSCs advocates therapeutic applications of MSCs in orthopedic injuries [53]. Seeding of MSCs, coaxed into biomaterial scaffolds, at defective bone tissue, regenerates defective bone tissues, within fourwks of transplantation; by the end of 32wks newly formed tissues integrate into old bone [54]. Osteoblasts, the bone-forming cells, have lesser actin cytoskeleton compared to adipocytes and MSCs. Treatment of MSCs with cytochalasin-D causes rapid transportation of G-actin, leading to osteogenic transformation of MSCs. Furthermore, injection of cytochalasin-D to mice tibia also promotes bone formation within a wk time frame [55]. The bone formation processes in mice, dog, and human are fundamentally similar, so outcomes of research on mice and dogs can be directional for regenerative application to human. Injection of MSCs to femur head of Legg-Calve-Perthes suffering dog heals the bone very fast and reduces the injury associated pain [55]. Degeneration of skeletal muscle and muscle cramps are very common to sledge dogs, animals, and individuals involved in adventurous athletics activities. Direct injection of adipose tissue derived MSCs to tear-site of semitendinosus muscle in dogs heals injuries much faster than traditional therapies [56]. Damage effect treatment for heart muscle regeneration is much more complex than regeneration of skeletal muscles, which needs high grade fine-tuned coordination of neurons with muscles. Coaxing of MSCs into alginate gel increases cell retention time that leads to releasing of tissue repairing factors in controlled manner. Transplantation of alginate encapsulated cells to mice heart reduces scar size and increases vascularisation, which leads to restoration of heart functions. Furthermore, transplanted MSCs face host inhospitable inflammatory immune responses and other mechanical forces at transplantation site, where encapsulation of cells keeps them away from all sorts of mechanical forces and enables sensing of host tissue microenvironment, and respond accordingly [57]. Ageing, disease, and medicine consumption can cause hair loss, known as alopecia. Although alopecia has no life threatening effects, emotional catchments can lead to psychological disturbance. The available treatments for alopecia include hair transplantation and use of drugs, where drugs are expensive to afford and generation of new hair follicle is challenging. Dermal papillary cells (DPCs), the specialized MSCs localized in hair follicle, are responsible for morphogenesis of hair follicle and hair cycling. The layer-by-layer coating of DPCs, called GAG coating, consists of coating of geletin as outer layer, middle layer of fibroblast growth factor 2 (FGF2) loaded alginate, and innermost layer of geletin. GAG coating creates tissue microenvironment for DPCs that can sustain immunological and mechanical obstacles, supporting generation of hair follicle. Transplantation of GAG-coated DPCs leads to abundant hair growth and maturation of hair follicle, where GAG coating serves as ECM, enhancing intrinsic therapeutic potential of DPCs [58]. During infection, the inflammatory cytokines secreted from host immune cells attract MSCs to the site of inflammation, which modulates inflammatory responses, representing MSCs as key candidate of regenerative medicine for infectious disease therapeutics. Coculture of macrophages (M) and adipose derived MSCs from Leishmania major (LM) susceptible and resistant mice demonstrates that AD-MSCs educate M against LM infection, differentially inducing M1 and M2 phenotype that represents AD-MSC as therapeutic agent for leishmanial therapy [93]. In summary, the multilineage differentiation potential of MSCs, as well as adoption of next-generation organoid culture system, avails MSCs as ideal regenerative medicine candidate.

Umbilical cord, generally thrown at the time of child birth, is the best known source for stem cells, procured in noninvasive manner, having lesser ethical constraints than ESCs. Umbilical cord is rich source of hematopoietic stem cells (HSCs) and MSCs, which possess enormous regeneration potential [94] (; ). The HSCs of cord blood are responsible for constant renewal of all types of blood cells and protective immune cells. The proliferation of HSCs is regulated by Musashi-2 protein mediated attenuation of Aryl hydrocarbon receptor (AHR) signalling in stem cells [95]. UCSCs can be cryopreserved at stem cells banks (; ), in operation by both private and public sector organization. Public stem cells banks operate on donation formats and perform rigorous screening for HLA typing and donated UCSCs remain available to anyone in need, whereas private stem cell banks operation is more personalized, availing cells according to donor consent. Stem cell banking is not so common, even in developed countries. Survey studies find that educated women are more eager to donate UCSCs, but willingness for donation decreases with subsequent deliveries, due to associated cost and safety concerns for preservation [96]. FDA has approved five HSCs for treatment of blood and other immunological complications [97]. The amniotic fluid, drawn during pregnancy for standard diagnostic purposes, is generally discarded without considering its vasculogenic potential. UCSCs are the best alternatives for those patients who lack donors with fully matched HLA typing for peripheral blood and PBMCs and bone marrow [98]. One major issue with UCSCs is number of cells in transplant, fewer cells in transplant require more time for engraftment to mature, and there are also risks of infection and mortality; in that case ex vivo propagation of UCSCs can meet the demand of desired outcomes. There are diverse protocols, available for ex vivo expansion of UCSCs, reviewed elsewhere [99]. Amniotic fluid stem cells (AFSCs), coaxed to fibrin (required for blood clotting, ECM interactions, wound healing, and angiogenesis) hydrogel and PEG supplemented with vascular endothelial growth factor (VEGF), give rise to vascularised tissue, when grafted to mice, suggesting that organoid cultures of UCSCs have promise for generation of biocompatible tissue patches, for treating infants born with congenital heart defects [59]. Retroviral integration of OCT4, KLF4, cMYC, and SOX2 transforms AFSCs into pluripotency stem cells known as AFiPSCs which can be directed to differentiate into extraembryonic trophoblast by BMP2 and BMP4 stimulation, which can be used for regeneration of placental tissues [60]. Wharton's jelly (WJ), the gelatinous substance inside umbilical cord, is rich in mucopolysaccharides, fibroblast, macrophages, and stem cells. The stem cells from UCB and WJ can be transdifferentiated into -cells. Homogeneous nature of WJ-SCs enables better differentiation into -cells; transplantation of these cells to streptozotocin induced diabetic mice efficiently brings glucose level to normal [7]. Easy access and expansion potential and plasticity to differentiate into multiple cell lineages represent WJ as an ideal candidate for regenerative medicine but cells viability changes with passages with maximum viable population at 5th-6th passages. So it is suggested to perform controlled expansion of WJ-MSCS for desired regenerative outcomes [9]. Study suggests that CD34+ expression leads to the best regenerative outcomes, with less chance of host-versus-graft rejection. In vitro expansion of UCSCs, in presence of StemRegenin-1 (SR-1), conditionally expands CD34+ cells [61]. In type I diabetic mellitus (T1DM), T-cell mediated autoimmune destruction of pancreatic -cells occurs, which has been considered as tough to treat. Transplantation of WJ-SCs to recent onset-T1DM patients restores pancreatic function, suggesting that WJ-MSCs are effective in regeneration of pancreatic tissue anomalies [62]. WJ-MSCs also have therapeutic importance for treatment of T2DM. A non-placebo controlled phase I/II clinical trial demonstrates that intravenous and intrapancreatic endovascular injection of WJ-MSCs to T2DM patients controls fasting glucose and glycated haemoglobin through improvement of -cells functions, evidenced by enhanced c-peptides and reduced inflammatory cytokines (IL-1 and IL-6) and T-cells counts [63]. Like diabetes, systematic lupus erythematosus (SLE) also can be treated with WJ-MSCs transplantation. During progression of SLE host immune system targets its own tissue leading to degeneration of renal, cardiovascular, neuronal, and musculoskeletal tissues. A non-placebo controlled follow-up study on 40 SLE patients demonstrates that intravenous infusion of WJ-MSC improves renal functions and decreases systematic lupus erythematosus disease activity index (SLEDAI) and British Isles Lupus Assessment Group (BILAG), and repeated infusion of WJ-MSCs protects the patient from relapse of the disease [64]. Sometimes, host inflammatory immune responses can be detrimental for HSCs transplantation and blood transfusion procedures. Infusion of WJ-MSC to patients, who had allogenic HSCs transplantation, reduces haemorrhage inflammation (HI) of bladder, suggesting that WJ-MSCs are potential stem cells adjuvant in HSCs transplantation and blood transfusion based therapies [100]. Apart from WJ, umbilical cord perivascular space and cord vein are also rich source for obtaining MSCs. The perivascular MSCs of umbilical cord are more primitive than WJ-MSCs and other MSCs from cord suggest that perivascular MSCs might be used as alternatives for WJ-MSCs for regenerative therapeutics outcome [101]. Based on origin, MSCs exhibit differential in vitro and in vivo properties and advocate functional characterization of MSCs, prior to regenerative applications. Emerging evidence suggests that UCSCs can heal brain injuries, caused by neurodegenerative diseases like Alzheimer's, Krabbe's disease, and so forth. Krabbe's disease, the infantile lysosomal storage disease, occurs due to deficiency of myelin synthesizing enzyme (MSE), affecting brain development and cognitive functions. Progression of neurodegeneration finally leads to death of babies aged two. Investigation shows that healing of peripheral nervous system (PNS) and central nervous system (CNS) tissues with Krabbe's disease can be achieved by allogenic UCSCs. UCSCs transplantation to asymptomatic infants with subsequent monitoring for 46 years reveals that UCSCs recover babies from MSE deficiency, improving myelination and cognitive functions, compared to those of symptomatic babies. The survival rate of transplanted UCSCs in asymptomatic and symptomatic infants was 100% and 43%, respectively, suggesting that early diagnosis and timely treatment are critical for UCSCs acceptance for desired therapeutic outcomes. UCSCs are more primitive than BMSCs, so perfect HLA typing is not critically required, representing UCSCs as an excellent source for treatment of all the diseases involving lysosomal defects, like Krabbe's disease, hurler syndrome, adrenoleukodystrophy (ALD), metachromatic leukodystrophy (MLD), Tay-Sachs disease (TSD), and Sandhoff disease [65]. Brain injuries often lead to cavities formation, which can be treated from neuronal parenchyma, generated ex vivo from UCSCs. Coaxing of UCSCs into human originated biodegradable matrix scaffold and in vitro expansion of cells in defined culture conditions lead to formation of neuronal organoids, within threewks' time frame. These organoids structurally resemble brain tissue and consisted of neuroblasts (GFAP+, Nestin+, and Ki67+) and immature stem cells (OCT4+ and SOX2+). The neuroblasts of these organoids further can be differentiated into mature neurons (MAP2+ and TUJ1+) [66]. Administration of high dose of drugs in divesting neuroblastoma therapeutics requires immediate restoration of hematopoiesis. Although BMSCs had been promising in restoration of hematopoiesis UCSCs are sparely used in clinical settings. A case study demonstrates that neuroblastoma patients who received autologous UCSCs survive without any associated side effects [12]. During radiation therapy of neoplasm, spinal cord myelitis can occur, although occurrence of myelitis is a rare event and usually such neurodegenerative complication of spinal cord occurs 624 years after exposure to radiations. Transplantation of allogenic UC-MSCs in laryngeal patients undergoing radiation therapy restores myelination [102]. For treatment of neurodegenerative disease like Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), traumatic brain injuries (TBI), Parkinson's, SCI, stroke, and so forth, distribution of transplanted UCSCs is critical for therapeutic outcomes. In mice and rat, injection of UCSCs and subsequent MRI scanning show that transplanted UCSCs migrate to CNS and multiple peripheral organs [67]. For immunomodulation of tumor cells disease recovery, transplantation of allogenic DCs is required. The CD11c+DCs, derived from UCB, are morphologically and phenotypically similar to those of peripheral blood derived CTLs-DCs, suggesting that UCB-DCs can be used for personalized medicine of cancer patient, in need for DCs transplantation [103]. Coculture of UCSCs with radiation exposed human lung fibroblast stops their transdifferentiation, which suggests that factors secreted from UCSCs may restore niche identity of fibroblast, if they are transplanted to lung after radiation therapy [104]. Tearing of shoulder cuff tendon can cause severe pain and functional disability, whereas ultrasound guided transplantation of UCB-MSCs in rabbit regenerates subscapularis tendon in fourwks' time frame, suggesting that UCB-MSCs are effective enough to treat tendons injuries when injected to focal points of tear-site [68]. Furthermore, transplantation of UCB-MSCs to chondral cartilage injuries site in pig knee along with HA hydrogel composite regenerates hyaline cartilage [69], suggesting that UCB-MSCs are effective regenerative medicine candidate for treating cartilage and ligament injuries. Physiologically circulatory systems of brain, placenta, and lungs are similar. Infusion of UCB-MSCs to preeclampsia (PE) induced hypertension mice reduces the endotoxic effect, suggesting that UC-MSCs are potential source for treatment of endotoxin induced hypertension during pregnancy, drug abuse, and other kinds of inflammatory shocks [105]. Transplantation of UCSCs to severe congenital neutropenia (SCN) patients restores neutrophils count from donor cells without any side effect, representing UCSCs as potential alternative for SCN therapy, when HLA matched bone marrow donors are not accessible [106]. In clinical settings, the success of myocardial infarction (MI) treatment depends on ageing, systemic inflammation in host, and processing of cells for infusion. Infusion of human hyaluronan hydrogel coaxed UCSCs in pigs induces angiogenesis, decreases scar area, improves cardiac function at preclinical level, and suggests that the same strategy might be effective for human [107]. In stem cells therapeutics, UCSCs transplantation can be either autologous or allogenic. Sometimes, the autologous UCSCs transplants cannot combat over tumor relapse, observed in Hodgkin's lymphoma (HL), which might require second dose transplantation of allogenic stem cells, but efficacy and tolerance of stem cells transplant need to be addressed, where tumor replace occurs. A case study demonstrates that second dose allogenic transplants of UCSCs effective for HL patients, who had heavy dose in prior transplant, increase the long term survival chances by 30% [10]. Patients undergoing long term peritoneal renal dialysis are prone to peritoneal fibrosis and can change peritoneal structure and failure of ultrafiltration processes. The intraperitoneal (IP) injection of WJ-MSCs prevents methylglyoxal induced programmed cell death and peritoneal wall thickening and fibrosis, suggesting that WJ-MSCs are effective in therapeutics of encapsulating peritoneal fibrosis [70]. In summary, UCB-HSCs, WJ-MSCs, perivascular MSCs, and UCB-MSCs have tissue regeneration potential.

Bone marrow found in soft spongy bones is responsible for formation of all peripheral blood and comprises hematopoietic stem cells (producing blood cells) and stromal cells (producing fat, cartilage, and bones) [108] (; ). Visually bone marrow has two types, red marrow (myeloid tissue; producing RBC, platelets, and most of WBC) and yellow marrow (producing fat cells and some WBC) [108]. Imbalance in marrow composition can culminate to the diseased condition. Since 1980, bone marrow transplantation is widely accepted for cancer therapeutics [109]. In order to avoid graft rejection, HLA typing of donors is a must, but completely matched donors are limited to family members, which hampers allogenic transplantation applications. Since matching of all HLA antigens is not critically required, in that case defining the critical antigens for haploidentical allogenic donor for patients, who cannot find fully matched donor, might relieve from donor constraints. Two-step administration of lymphoid and myeloid BMSCs from haploidentical donor to the patients of aplastic anaemia and haematological malignancies reconstructs host immune system and the outcomes are almost similar to fully matched transplants, which recommends that profiling of critically important HLA is sufficient for successful outcomes of BMSCs transplantation. Haploidentical HLA matching protocol is the major process for minorities and others who do not have access to matched donor [71]. Furthermore, antigen profiling is not the sole concern for BMSCs based therapeutics. For example, restriction of HIV1 (human immune deficiency virus) infection is not feasible through BMSCs transplantation because HIV1 infection is mediated through CD4+ receptors, chemokine CXC motif receptor 4 (CXCR4), and chemokine receptor 5 (CCR5) for infecting and propagating into T helper (Th), monocytes, macrophages, and dendritic cells (DCs). Genetic variation in CCR2 and CCR5 receptors is also a contributory factor; mediating protection against infection has been reviewed elsewhere [110]. Engineering of hematopoietic stem and progenitor cells (HSPCs) derived CD4+ cells to express HIV1 antagonistic RNA, specifically designed for targeting HIV1 genome, can restrict HIV1 infection, through immune elimination of latently infected CD4+ cells. A single dose infusion of genetically modified (GM), HIV1 resistant HSPCs can be the alternative of HIV1 retroviral therapy. In the present scenario stem cells source, patient selection, transplantation-conditioning regimen, and postinfusion follow-up studies are the major factors, which can limit application of HIV1 resistant GM-HSPCs (CD4+) cells application in AIDS therapy [72, 73]. Platelets, essential for blood clotting, are formed from megakaryocytes inside the bone marrow [74]. Due to infection, trauma, and cancer, there are chances of bone marrow failure. To an extent, spongy bone marrow microenvironment responsible for lineage commitment can be reconstructed ex vivo [75]. The ex vivo constructed 3D-scaffolds consisted of microtubule and silk sponge, flooded with chemically defined organ culture medium, which mimics bone marrow environment. The coculture of megakaryocytes and embryonic stem cells (ESCs) in this microenvironment leads to generation of functional platelets from megakaryocytes [75]. The ex vivo 3D-scaffolds of bone microenvironment can stride the path for generation of platelets in therapeutic quantities for regenerative medication of burns [75] and blood clotting associated defects. Accidents, traumatic injuries, and brain stroke can deplete neuronal stem cells (NSCs), responsible for generation of neurons, astrocytes, and oligodendrocytes. Brain does not repopulate NSCs and heal traumatic injuries itself and transplantation of BMSCs also can heal neurodegeneration alone. Lipoic acid (LA), a known pharmacological antioxidant compound used in treatment of diabetic and multiple sclerosis neuropathy when combined with BMSCs, induces neovascularisation at focal cerebral injuries, within 8wks of transplantation. Vascularisation further attracts microglia and induces their colonization into scaffold, which leads to differentiation of BMSCs to become brain tissue, within 16wks of transplantation. In this approach, healing of tissue directly depends on number of BMSCs in transplantation dose [76]. Dental caries and periodontal disease are common craniofacial disease, often requiring jaw bone reconstruction after removal of the teeth. Traditional therapy focuses on functional and structural restoration of oral tissue, bone, and teeth rather than biological restoration, but BMSCs based therapies promise for regeneration of craniofacial bone defects, enabling replacement of missing teeth in restored bones with dental implants. Bone marrow derived CD14+ and CD90+ stem and progenitor cells, termed as tissue repair cells (TRC), accelerate alveolar bone regeneration and reconstruction of jaw bone when transplanted in damaged craniofacial tissue, earlier to oral implants. Hence, TRC therapy reduces the need of secondary bone grafts, best suited for severe defects in oral bone, skin, and gum, resulting from trauma, disease, or birth defects [77]. Overall, HSCs have great value in regenerative medicine, where stem cells transplantation strategies explore importance of niche in tissue regeneration. Prior to transplantation of BMSCs, clearance of original niche from target tissue is necessary for generation of organoid and organs without host-versus-graft rejection events. Some genetic defects can lead to disorganization of niche, leading to developmental errors. Complementation with human blastocyst derived primary cells can restore niche function of pancreas in pigs and rats, which defines the concept for generation of clinical grade human pancreas in mice and pigs [111]. Similar to other organs, diaphragm also has its own niche. Congenital defects in diaphragm can affect diaphragm functions. In the present scenario functional restoration of congenital diaphragm defects by surgical repair has risk of reoccurrence of defects or incomplete restoration [8]. Decellularization of donor derived diaphragm offers a way for reconstruction of new and functionally compatible diaphragm through niche modulation. Tissue engineering technology based decellularization of diaphragm and simultaneous perfusion of bone marrow mesenchymal stem cells (BM-MSCs) facilitates regeneration of functional scaffolds of diaphragm tissues [8]. In vivo replacement of hemidiaphragm in rats with reseeded scaffolds possesses similar myography and spirometry as it has in vivo in donor rats. These scaffolds retaining natural architecture are devoid of immune cells, retaining intact extracellular matrix that supports adhesion, proliferation, and differentiation of seeded cells [8]. These findings suggest that cadaver obtained diaphragm, seeded with BM-MSCs, can be used for curing patients in need for restoration of diaphragm functions (; ). However, BMSCs are heterogeneous population, which might result in differential outcomes in clinical settings; however clonal expansion of BMSCs yields homogenous cells population for therapeutic application [8]. One study also finds that intracavernous delivery of single clone BMSCs can restore erectile function in diabetic mice [112] and the same strategy might be explored for adult human individuals. The infection of hepatitis C virus (HCV) can cause liver cirrhosis and degeneration of hepatic tissue. The intraparenchymal transplantation of bone marrow mononuclear cells (BMMNCs) into liver tissue decreases aspartate aminotransferase (AST), alanine transaminase (ALT), bilirubin, CD34, and -SMA, suggesting that transplanted BMSCs restore hepatic functions through regeneration of hepatic tissues [113]. In order to meet the growing demand for stem cells transplantation therapy, donor encouragement is always required [8]. The stem cells donation procedure is very simple; with consent donor gets an injection of granulocyte-colony stimulating factor (G-CSF) that increases BMSCs population. Bone marrow collection is done from hip bone using syringe in 4-5hrs, requiring local anaesthesia and within a wk time frame donor gets recovered donation associated weakness.

The field of iPSCs technology and research is new to all other stem cells research, emerging in 2006 when, for the first time, Takahashi and Yamanaka generated ESCs-like cells through genetic incorporation of four factors, Sox2, Oct3/4, Klf4, and c-Myc, into skin fibroblast [3]. Due to extensive nuclear reprogramming, generated iPSCs are indistinguishable from ESCs, for their transcriptome profiling, epigenetic markings, and functional competence [3], but use of retrovirus in transdifferentiation approach has questioned iPSCs technology. Technological advancement has enabled generation of iPSCs from various kinds of adult cells phasing through ESCs or direct transdifferentiation. This section of review outlines most recent advancement in iPSC technology and regenerative applications (; ). Using the new edge of iPSCs technology, terminally differentiated skin cells directly can be transformed into kidney organoids [114], which are functionally and structurally similar to those of kidney tissue in vivo. Up to certain extent kidneys heal themselves; however natural regeneration potential cannot meet healing for severe injuries. During kidneys healing process, a progenitor stem cell needs to become 20 types of cells, required for waste excretion, pH regulation, and restoration of water and electrolytic ions. The procedure for generation of kidney organoids ex vivo, containing functional nephrons, has been identified for human. These ex vivo kidney organoids are similar to fetal first-trimester kidneys for their structure and physiology. Such kidney organoids can serve as model for nephrotoxicity screening of drugs, disease modelling, and organ transplantation. However generation of fully functional kidneys is a far seen event with today's scientific technologies [114]. Loss of neurons in age-related macular degeneration (ARMD) is the common cause of blindness. At preclinical level, transplantation of iPSCs derived neuronal progenitor cells (NPCs) in rat limits progression of disease through generation of 5-6 layers of photoreceptor nuclei, restoring visual acuity [78]. The various approaches of iPSCs mediated retinal regeneration including ARMD have been reviewed elsewhere [79]. Placenta, the cordial connection between mother and developing fetus, gets degenerated in certain pathophysiological conditions. Nuclear programming of OCT4 knock-out (KO) and wild type (WT) mice fibroblast through transient expression of GATA3, EOMES, TFAP2C, and +/ cMYC generates transgene independent trophoblast stem-like cells (iTSCs), which are highly similar to blastocyst derived TSCs for DNA methylation, H3K7ac, nucleosome deposition of H2A.X, and other epigenetic markings. Chimeric differentiation of iTSCs specifically gives rise to haemorrhagic lineages and placental tissue, bypassing pluripotency phase, opening an avenue for generation of fully functional placenta for human [115]. Neurodegenerative disease like Alzheimer's and obstinate epilepsies can degenerate cerebrum, controlling excitatory and inhibitory signals of the brain. The inhibitory tones in cerebral cortex and hippocampus are accounted by -amino butyric acid secreting (GABAergic) interneurons (INs). Loss of these neurons often leads to progressive neurodegeneration. Genomic integration of Ascl1, Dlx5, Foxg1, and Lhx6 to mice and human fibroblast transforms these adult cells into GABAergic-INs (iGABA-INs). These cells have molecular signature of telencephalic INs, release GABA, and show inhibition to host granule neuronal activity [81]. Transplantation of these INs in developing embryo cures from genetic and acquired seizures, where transplanted cells disperse and mature into functional neuronal circuits as local INs [82]. Dorsomorphin and SB-431542 mediated inhibition of TGF- and BMP signalling direct transformation of human iPSCs into cortical spheroids. These cortical spheroids consisted of both peripheral and cortical neurons, surrounded by astrocytes, displaying transcription profiling and electrophysiology similarity with developing fetal brain and mature neurons, respectively [83]. The underlying complex biology and lack of clear etiology and genetic reprogramming and difficulty in recapitulation of brain development have barred understanding of pathophysiology of autism spectrum disorder (ASD) and schizophrenia. 3D organoid cultures of ASD patient derived iPSC generate miniature brain organoid, resembling fetal brain few months after gestation. The idiopathic conditions of these organoids are similar with brain of ASD patients; both possess higher inhibitory GABAergic neurons with imbalanced neuronal connection. Furthermore these organoids express forkhead Box G1 (FOXG1) much higher than normal brain tissue, which explains that FOXG1 might be the leading cause of ASD [84]. Degeneration of other organs and tissues also has been reported, like degeneration of lungs which might occur due to tuberculosis infection, fibrosis, and cancer. The underlying etiology for lung degeneration can be explained through organoid culture. Coaxing of iPSC into inert biomaterial and defined culture leads to formation of lung organoids that consisted of epithelial and mesenchymal cells, which can survive in culture for months. These organoids are miniature lung, resemble tissues of large airways and alveoli, and can be used for lung developmental studies and screening of antituberculosis and anticancer drugs [87]. The conventional multistep reprogramming for iPSCs consumes months of time, while CRISPER-Cas9 system based episomal reprogramming system that combines two steps together enables generation of ESCs-like cells in less than twowks, reducing the chances of culture associated genetic abrasions and unwanted epigenetic [80]. This approach can yield single step ESCs-like cells in more personalized way from adults with retinal degradation and infants with severe immunodeficiency, involving correction for genetic mutation of OCT4 and DNMT3B [80]. The iPSCs expressing anti-CCR5-RNA, which can be differentiated into HIV1 resistant macrophages, have applications in AIDS therapeutics [88]. The diversified immunotherapeutic application of iPSCs has been reviewed elsewhere [89]. The -1 antitrypsin deficiency (A1AD) encoded by serpin peptidase inhibitor clade A member 1 (SERPINA1) protein synthesized in liver protects lungs from neutrophils elastase, the enzyme causing disruption of lungs connective tissue. A1AD deficiency is common cause of both lung and liver disease like chronic obstructive pulmonary disease (COPD) and liver cirrhosis. Patient specific iPSCs from lung and liver cells might explain pathophysiology of A1AD deficiency. COPD patient derived iPSCs show sensitivity to toxic drugs which explains that actual patient might be sensitive in similar fashion. It is known that A1AD deficiency is caused by single base pair mutation and correction of this mutation fixes the A1AD deficiency in hepatic-iPSCs [85]. The high order brain functions, like emotions, anxiety, sleep, depression, appetite, breathing heartbeats, and so forth, are regulated by serotonin neurons. Generation of serotonin neurons occurs prior to birth, which are postmitotic in their nature. Any sort of developmental defect and degeneration of serotonin neurons might lead to neuronal disorders like bipolar disorder, depression, and schizophrenia-like psychiatric conditions. Manipulation of Wnt signalling in human iPSCs in defined culture conditions leads to an in vitro differentiation of iPSCs to serotonin-like neurons. These iPSCs-neurons primarily localize to rhombomere 2-3 segment of rostral raphe nucleus, exhibit electrophysiological properties similar to serotonin neurons, express hydroxylase 2, the developmental marker, and release serotonin in dose and time dependent manner. Transplantation of these neurons might cure from schizophrenia, bipolar disorder, and other neuropathological conditions [116]. The iPSCs technology mediated somatic cell reprogramming of ventricular monocytes results in generation of cells, similar in morphology and functionality with PCs. SA note transplantation of PCs to large animals improves rhythmic heart functions. Pacemaker needs very reliable and robust performance so understanding of transformation process and site of transplantation are the critical aspect for therapeutic validation of iPSCs derived PCs [28]. Diabetes is a major health concern in modern world, and generation of -cells from adult tissue is challenging. Direct reprogramming of skin cells into pancreatic cells, bypassing pluripotency phase, can yield clinical grade -cells. This reprogramming strategy involves transformation of skin cells into definitive endodermal progenitors (cDE) and foregut like progenitor cells (cPF) intermediates and subsequent in vitro expansion of these intermediates to become pancreatic -cells (cPB). The first step is chemically complex and can be understood as nonepisomal reprogramming on day one with pluripotency factors (OCT4, SOX2, KLF4, and hair pin RNA against p53), then supplementation with GFs and chemical supplements on day seven (EGF, bFGF, CHIR, NECA, NaB, Par, and RG), and two weeks later (Activin-A, CHIR, NECA, NaB, and RG) yielding DE and cPF [86]. Transplantation of cPB yields into glucose stimulated secretion of insulin in diabetic mice defines that such cells can be explored for treatment of T1DM and T2DM in more personalized manner [86]. iPSCs represent underrated opportunities for drug industries and clinical research laboratories for development of therapeutics, but safety concerns might limit transplantation applications (; ) [117]. Transplantation of human iPSCs into mice gastrula leads to colonization and differentiation of cells into three germ layers, evidenced with clinical developmental fat measurements. The acceptance of human iPSCs by mice gastrula suggests that correct timing and appropriate reprogramming regime might delimit human mice species barrier. Using this fact of species barrier, generation of human organs in closely associated primates might be possible, which can be used for treatment of genetic factors governed disease at embryo level itself [118]. In summary, iPSCs are safe and effective for treatment of regenerative medicine.

The unstable growth of human population threatens the existence of wildlife, through overexploitation of natural habitats and illegal killing of wild animals, leading many species to face the fate of being endangered and go for extinction. For wildlife conservation, the concept of creation of frozen zoo involves preservation of gene pool and germ plasm from threatened and endangered species (). The frozen zoo tissue samples collection from dead or live animal can be DNA, sperms, eggs, embryos, gonads, skin, or any other tissue of the body [119]. Preserved tissue can be reprogrammed or transdifferentiated to become other types of tissues and cells, which opens an avenue for conservation of endangered species and resurrection of life (). The gonadal tissue from young individuals harbouring immature tissue can be matured in vivo and ex vivo for generation of functional gametes. Transplantation of SSCs to testis of male from the same different species can give rise to spermatozoa of donor cells [120], which might be used for IVF based captive breeding of wild animals. The most dangerous fact in wildlife conservation is low genetic diversity, too few reproductively capable animals which cannot maintain adequate genetic diversity in wild or captivity. Using the edge of iPSC technology, pluripotent stem cells can be generated from skin cells. For endangered drill, Mandrillus leucophaeus, and nearly extinct white rhinoceros, Ceratotherium simum cottoni, iPSC has been generated in 2011 [121]. The endangered animal drill (Mandrillus leucophaeus) is genetically very close to human and often suffers from diabetes, while rhinos are genetically far removed from other primates. The progress in iPSCs, from the human point of view, might be transformed for animal research for recapturing reproductive potential and health in wild animals. However, stem cells based interventions in wild animals are much more complex than classical conservation planning and biomedical research has to face. Conversion of iPSC into egg or sperm can open the door for generation of IVF based embryo; those might be transplanted in womb of live counterparts for propagation of population. Recently, iPSCs have been generated for snow leopard (Panthera uncia), native to mountain ranges of central Asia, which belongs to cat family; this breakthrough has raised the possibilities for cryopreservation of genetic material for future cloning and other assisted reproductive technology (ART) applications, for the conservation of cat species and biodiversity. Generation of leopard iPSCs has been achieved through retroviral-system based genomic integration of OCT4, SOX2, KLF4, cMYC, and NANOG. These iPSCs from snow leopard also open an avenue for further transformation of iPSCs into gametes [122]. The in vivo maturation of grafted tissue depends both on age and on hormonal status of donor tissue. These facts are equally applicable to accepting host. Ectopic xenografts of cryopreserved testis tissue from Indian spotted deer (Moschiola indica) to nude mice yielded generation of spermatocytes [123], suggesting that one-day procurement of functional sperm from premature tissue might become a general technique in wildlife conservation. In summary, tissue biopsies from dead or live animals can be used for generation of iPSCs and functional gametes; those can be used in assisted reproductive technology (ART) for wildlife conservation.

The spectacular progress in the field of stem cells research represents great scope of stem cells regenerative therapeutics. It can be estimated that by 2020 or so we will be able to produce wide array of tissue, organoid, and organs from adult stem cells. Inductions of pluripotency phenotypes in terminally differentiated adult cells have better therapeutic future than ESCs, due to least ethical constraints with adult cells. In the coming future, there might be new pharmaceutical compounds; those can activate tissue specific stem cells, promote stem cells to migrate to the side of tissue injury, and promote their differentiation to tissue specific cells. Except few countries, the ongoing financial and ethical hindrance on ESCs application in regenerative medicine have more chance for funding agencies to distribute funding for the least risky projects on UCSCs, BMSCs, and TSPSCs from biopsies. The existing stem cells therapeutics advancements are more experimental and high in cost; due to that application on broad scale is not feasible in current scenario. In the near future, the advancements of medical science presume using stem cells to treat cancer, muscles damage, autoimmune disease, and spinal cord injuries among a number of impairments and diseases. It is expected that stem cells therapies will bring considerable benefits to the patients suffering from wide range of injuries and disease. There is high optimism for use of BMSCs, TSPSCs, and iPSCs for treatment of various diseases to overcome the contradictions associated with ESCs. For advancement of translational application of stem cells, there is a need of clinical trials, which needs funding rejoinder from both public and private organizations. The critical evaluation of regulatory guidelines at each phase of clinical trial is a must to comprehend the success and efficacy in time frame.

Dr. Anuradha Reddy from Centre for Cellular and Molecular Biology Hyderabad and Mrs. Sarita Kumari from Department of Yoga Science, BU, Bhopal, India, are acknowledged for their critical suggestions and comments on paper.

There are no competing interests associated with this paper.

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Stem Cells Applications in Regenerative Medicine and ...

Stem cell therapy, sometimes called regenerative medicine, is one of the most exciting areas of the life sciences sector right now.

Since the pandemic, the sector has emerged into the publics spotlight with new developments in mRNA-based vaccines and therapies.

Nasdaq is the obvious breeding ground for world-class stem cell companies with the likes of Moderna and BioNTech, and lesser known names like Anavex and Enochian.

In Australia, Mesoblast (ASX:MSB) has long been the local poster child for the regenerative medicine industry.

Mesoblast has developed a platform of innovative cellular medicines, but the company has struggled since the FDA rejected its drug in October last year.

Now, other ASX companies like Cynata Therapeutics (ASX:CYP)are making rapid progress to take over the mantle from MSB in this hot field.

Cynata is developing a mesenchymal stem cells (or MSC) technology, which it says has huge therapeutic potential for numerous unmet medical needs.

This includes asthma, heart attack, sepsis, and acute respiratory distress syndrome (ARDS), which all add up to a market opportunity worth $46bn, says the company.

According to CEO Dr Ross Macdonald, who spoke to Stockhead today, MSC is the hottest segment of stem cell therapy at the moment, and has gained a lot of attention recently.

There is a huge interest, and theres been more than 1000 clinical trials conducted around the world using MSC, Dr Macdonald told Stockhead.

He explains that the humans immune system controls many of the bodys functions responsible for repairing tissue after injury or disease, and defending against invading germs like viruses or bacteria.

And just like an orchestral conductor, MSC seems to be playing a central role in that coordination within our immune system.

We now have a firm understanding of how those cells coordinate the bodys responses, and can use that knowledge to enhance those processes that they control, Dr Macdonald explained.

In short, MSC therapies work by expressing a variety of chemokines and cytokines that aid in repair of degraded tissue, restoration of normal tissue metabolism and, most importantly, counteracting inflammation.

And because MSCs play that co-ordination role within the immune system, they can be used to treat different diseases.

However theres one big problem with cell-based therapies, and its not to do with the safety and efficacy.

Its how to manufacture these products on a mass scale, that is the greatest challenge right now, says Dr Macdonald.

Unlike aspirin where it can be synthesised in a chemical lab and produced in bulk, manufacturing a living drug like a cell is a whole lot more complicated.

But that big challenge is the exact area of strength and competitive advantage that Cynata has, Dr Macdonald told Stockhead.

He says Cynata has a technology platform which allows it to manufacture essentially limitless quantities of MSCs, consistently and economically.

Dr Macdonald explains there are two approaches to using cell therapy, the autologous and the allogeneic approach.

The autologous approach is where the patient themselves serves as their own donor.

This is obviously bespoke and inefficient, because the drug can only be manufactured for that one patient, and is obviously not an industrialised process, he said.

But by taking an allogeneic approach, Cynata has the ability to start with a one time donation of cells from one single donor.

Well never have to go back to that human donor ever again, so our process of producing cells has become a very much more typical industrialised process.

The company has a patent for this, with two clinical trials underway and two more under preparation.

A Phase 3 clinical trial for osteoarthritis which is funded by a NHMRC grant has progressed the furthest, while a Phase 2 trial in COVID-19 is ongoing.

Meanwhile a Phase 1 study in GvHD, which was published in prestigious journal Nature Medicine, is probably the closest to commercialisation according to Dr Macdonald.

GvHD is a challenging disease which occurs in patients who have had a bone marrow transplant as part of their chemotherapy treatment for cancer.

Chemo is still very much a sledgehammer therapy where you use very toxic drugs that do kill the cancer cells, but they also kill the surrounding healthy cells that grow hair and bone marrow.

Unfortunately for many patients, the bone marrow transplant reacts against their body and starts to attack all of the tissues in the body, and its ultimately fatal.

Its a horrible death, destroying the lungs, liver, intestines and the skin, Macdonald explains.

Cynatas MSC therapy has been shown to reset that reaction, so the patient can recover from the GvHD, and also recover from their underlying cancer.

With all these clinical trials concurrently under way, Macdonald believes there is a clear significant upside potential for Cynata, particularly given its small market cap of $70m compared to other similar plays like Mesoblast ($1 billion market cap).

Osteopore (ASX:OSX) focuses in bones and specialises in the production of 3D printed bioresorbable implants that are used in surgical procedures to assist with the natural stages of bone healing.

The 3D bio-printer makes a scaffold that mimics bone, with a patented micro-architecture which traps the patients own stem cells.

Orthocell (ASX:OCC) develops collagen medical devices and cellular therapies for the repair and regeneration of human tendons, bone, nerve and cartilage defects.

Its flagship product, the CelGro, is a naturally derived collagen medical device for tissue repair.

Aroa Biosurgery (ASX:ARX) develops FDA-approved medical devices for wounds and tissue repair using its extracellular matrix (ECM) technology, mainly in the United States.

Recent study shows 100% success rates from the use of its Myriad product when patients underwent surgical reconstruction of exposed vital structures such as bone and tendon.

Regeneus (ASX:RGS) Progenza is a cellular therapy targeting pain and inflammation which uses Secretome to improve not only the resident tissue, but the MSCs themselves.

It fills a gap in the current treatment market for osteoarthritis, by providing disease modification and pain relief to address patient symptoms.

Anteris Technologies (ASX:AVR) claims that its Adapt Technology is the first and only bio-scaffold technology that completely re-engineers xenograft tissue into a pure collagen scaffold.

A recent study indicated that Adapt-treated tissue has superior anti-calcification attributes compared with tissues used in competitor valves.

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Mesoblast has long been the one poster child for stem cell therapy. Now Cynata and other ASX stocks have e ... - Stockhead

KENILWORTH, N.J.--(BUSINESS WIRE)--Merck (NYSE: MRK), known as MSD outside the United States and Canada, today announced that the U.S. Food and Drug Administration (FDA) has approved KEYTRUDA, Mercks anti-PD-1 therapy, for the adjuvant treatment of patients with renal cell carcinoma (RCC) at intermediate-high or high risk of recurrence following nephrectomy, or following nephrectomy and resection of metastatic lesions. The approval is based on data from the pivotal Phase 3 KEYNOTE-564 trial, in which KEYTRUDA demonstrated a statistically significant improvement in disease-free survival (DFS), reducing the risk of disease recurrence or death by 32% (HR=0.68 [95% CI, 0.53-0.87]; p=0.0010) compared to placebo. Median DFS has not been reached for either group.

Despite decades of research, limited adjuvant treatment options have been available for earlier-stage renal cell carcinoma patients who are often at risk for recurrence. In KEYNOTE-564, pembrolizumab reduced the risk of disease recurrence or death by 32%, providing a promising new treatment option for certain patients at intermediate-high or high risk of recurrence, said Dr. Toni K. Choueiri, director, Lank Center for Genitourinary Oncology, Dana-Farber Cancer Institute, and professor of medicine, Harvard Medical School. With this FDA approval, pembrolizumab may address a critical unmet treatment need and has the potential to become a new standard of care in the adjuvant setting for appropriately selected patients.

Immune-mediated adverse reactions, which may be severe or fatal, can occur in any organ system or tissue and can affect more than one body system simultaneously. Immune-mediated adverse reactions can occur at any time during or after treatment with KEYTRUDA, including pneumonitis, colitis, hepatitis, endocrinopathies, nephritis, dermatologic reactions, solid organ transplant rejection, and complications of allogeneic hematopoietic stem cell transplantation. Important immune-mediated adverse reactions listed here may not include all possible severe and fatal immune-mediated adverse reactions. Early identification and management of immune-mediated adverse reactions are essential to ensure safe use of KEYTRUDA. Based on the severity of the adverse reaction, KEYTRUDA should be withheld or permanently discontinued and corticosteroids administered if appropriate. KEYTRUDA can also cause severe or life-threatening infusion-related reactions. Based on its mechanism of action, KEYTRUDA can cause fetal harm when administered to a pregnant woman. For more information, see Selected Important Safety Information below.

KEYTRUDA is foundational for the treatment of patients with certain advanced cancers, and this approval marks the fourth indication for KEYTRUDA in earlier stages of cancer, said Dr. Scot Ebbinghaus, vice president, clinical research, Merck Research Laboratories. KEYTRUDA is now the first immunotherapy approved for the adjuvant treatment of certain patients with renal cell carcinoma. This milestone is a testament to our commitment to help more people living with cancer.

In RCC, Merck has a broad clinical development program exploring KEYTRUDA, as monotherapy or in combination, as well as other investigational products across multiple settings and stages of RCC, including adjuvant and advanced or metastatic disease.

Data Supporting the Approval

KEYTRUDA demonstrated a statistically significant improvement in DFS in patients with RCC at intermediate-high or high risk of recurrence following nephrectomy, or following nephrectomy and resection of metastatic lesions compared with placebo (HR=0.68 [95% CI, 0.53-0.87]; p=0.0010). The trial will continue to assess overall survival (OS) as a secondary outcome measure.

In KEYNOTE-564, the median duration of exposure to KEYTRUDA was 11.1 months (range, 1 day to 14.3 months). Serious adverse reactions occurred in 20% of these patients receiving KEYTRUDA. Serious adverse reactions (1%) were acute kidney injury, adrenal insufficiency, pneumonia, colitis and diabetic ketoacidosis (1% each). Fatal adverse reactions occurred in 0.2% of those treated with KEYTRUDA, including one case of pneumonia. Adverse reactions leading to discontinuation occurred in 21% of patients receiving KEYTRUDA; the most common (1%) were increased alanine aminotransferase (1.6%), colitis and adrenal insufficiency (1% each). The most common adverse reactions (all grades 20%) in the KEYTRUDA arm were musculoskeletal pain (41%), fatigue (40%), rash (30%), diarrhea (27%), pruritus (23%) and hypothyroidism (21%).

About KEYNOTE-564

KEYNOTE-564 (ClinicalTrials.gov, NCT03142334) is a multicenter, randomized, double-blind, placebo-controlled Phase 3 trial evaluating KEYTRUDA as adjuvant therapy for RCC in 994 patients with intermediate-high or high risk of recurrence of RCC or M1 no evidence of disease (NED). Patients must have undergone a partial nephroprotective or radical complete nephrectomy (and complete resection of solid, isolated, soft tissue metastatic lesion[s] in M1 NED participants) with negative surgical margins for at least four weeks prior to the time of screening. Patients were excluded from the trial if they had received prior systemic therapy for advanced RCC. Patients with active autoimmune disease or a medical condition that required immunosuppression were also ineligible. The major efficacy outcome measure was investigator-assessed DFS, defined as time to recurrence, metastasis or death. An additional outcome measure was OS. Patients were randomized (1:1) to receive KEYTRUDA 200 mg administered intravenously every three weeks or placebo for up to one year until disease recurrence or unacceptable toxicity.

About Renal Cell Carcinoma (RCC)

Renal cell carcinoma is by far the most common type of kidney cancer; about nine out of 10 kidney cancer diagnoses are RCCs. Renal cell carcinoma is about twice as common in men than in women. Most cases of RCC are discovered incidentally during imaging tests for other abdominal diseases. Worldwide, it is estimated there were more than 431,000 new cases of kidney cancer diagnosed and more than 179,000 deaths from the disease in 2020. In the U.S., it is estimated there will be more than 76,000 new cases of kidney cancer diagnosed and almost 14,000 deaths from the disease in 2021.

About Mercks Early-Stage Cancer Clinical Program

Finding cancer at an earlier stage may give patients a greater chance of long-term survival. Many cancers are considered most treatable and potentially curable in their earliest stage of disease. Building on the strong understanding of the role of KEYTRUDA in later-stage cancers, Merck is studying KEYTRUDA in earlier disease states, with approximately 20 ongoing registrational studies across multiple types of cancer.

About KEYTRUDA (pembrolizumab) Injection, 100 mg

KEYTRUDA is an anti-programmed death receptor-1 (PD-1) therapy that works by increasing the ability of the bodys immune system to help detect and fight tumor cells. KEYTRUDA is a humanized monoclonal antibody that blocks the interaction between PD-1 and its ligands, PD-L1 and PD-L2, thereby activating T lymphocytes which may affect both tumor cells and healthy cells.

Merck has the industrys largest immuno-oncology clinical research program. There are currently more than 1,600 trials studying KEYTRUDA across a wide variety of cancers and treatment settings. The KEYTRUDA clinical program seeks to understand the role of KEYTRUDA across cancers and the factors that may predict a patient's likelihood of benefitting from treatment with KEYTRUDA, including exploring several different biomarkers.

Selected KEYTRUDA (pembrolizumab) Indications in the U.S.

Melanoma

KEYTRUDA is indicated for the treatment of patients with unresectable or metastatic melanoma.

KEYTRUDA is indicated for the adjuvant treatment of patients with melanoma with involvement of lymph node(s) following complete resection.

Non-Small Cell Lung Cancer

KEYTRUDA, in combination with pemetrexed and platinum chemotherapy, is indicated for the first-line treatment of patients with metastatic nonsquamous non-small cell lung cancer (NSCLC), with no EGFR or ALK genomic tumor aberrations.

KEYTRUDA, in combination with carboplatin and either paclitaxel or paclitaxel protein-bound, is indicated for the first-line treatment of patients with metastatic squamous NSCLC.

KEYTRUDA, as a single agent, is indicated for the first-line treatment of patients with NSCLC expressing PD-L1 [tumor proportion score (TPS) 1%] as determined by an FDA-approved test, with no EGFR or ALK genomic tumor aberrations, and is:

KEYTRUDA, as a single agent, is indicated for the treatment of patients with metastatic NSCLC whose tumors express PD-L1 (TPS 1%) as determined by an FDA-approved test, with disease progression on or after platinum-containing chemotherapy. Patients with EGFR or ALK genomic tumor aberrations should have disease progression on FDA-approved therapy for these aberrations prior to receiving KEYTRUDA.

Head and Neck Squamous Cell Cancer

KEYTRUDA, in combination with platinum and fluorouracil (FU), is indicated for the first-line treatment of patients with metastatic or with unresectable, recurrent head and neck squamous cell carcinoma (HNSCC).

KEYTRUDA, as a single agent, is indicated for the first-line treatment of patients with metastatic or with unresectable, recurrent HNSCC whose tumors express PD-L1 [combined positive score (CPS 1)] as determined by an FDA-approved test.

KEYTRUDA, as a single agent, is indicated for the treatment of patients with recurrent or metastatic HNSCC with disease progression on or after platinum-containing chemotherapy.

Classical Hodgkin Lymphoma

KEYTRUDA is indicated for the treatment of adult patients with relapsed or refractory classical Hodgkin lymphoma (cHL).

KEYTRUDA is indicated for the treatment of pediatric patients with refractory cHL, or cHL that has relapsed after 2 or more lines of therapy.

Primary Mediastinal Large B-Cell Lymphoma

KEYTRUDA is indicated for the treatment of adult and pediatric patients with refractory primary mediastinal large B-cell lymphoma (PMBCL), or who have relapsed after 2 or more prior lines of therapy. KEYTRUDA is not recommended for treatment of patients with PMBCL who require urgent cytoreductive therapy.

Urothelial Carcinoma

KEYTRUDA is indicated for the treatment of patients with locally advanced or metastatic urothelial carcinoma (mUC):

Non-muscle Invasive Bladder Cancer

KEYTRUDA is indicated for the treatment of patients with Bacillus Calmette-Guerin-unresponsive, high-risk, non-muscle invasive bladder cancer (NMIBC) with carcinoma in situ with or without papillary tumors who are ineligible for or have elected not to undergo cystectomy.

Microsatellite Instability-High or Mismatch Repair Deficient Cancer

KEYTRUDA is indicated for the treatment of adult and pediatric patients with unresectable or metastatic microsatellite instability-high (MSI-H) or mismatch repair deficient (dMMR) solid tumors that have progressed following prior treatment and who have no satisfactory alternative treatment options.

This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials. The safety and effectiveness of KEYTRUDA in pediatric patients with MSI-H central nervous system cancers have not been established.

Microsatellite Instability-High or Mismatch Repair Deficient Colorectal Cancer

KEYTRUDA is indicated for the treatment of patients with unresectable or metastatic MSI-H or dMMR colorectal cancer (CRC).

Gastric Cancer

KEYTRUDA, in combination with trastuzumab, fluoropyrimidine- and platinum-containing chemotherapy, is indicated for the first-line treatment of patients with locally advanced unresectable or metastatic HER2-positive gastric or gastroesophageal junction (GEJ) adenocarcinoma.

This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Esophageal Cancer

KEYTRUDA is indicated for the treatment of patients with locally advanced or metastatic esophageal or GEJ (tumors with epicenter 1 to 5 centimeters above the GEJ) carcinoma that is not amenable to surgical resection or definitive chemoradiation either:

Cervical Cancer

KEYTRUDA, in combination with chemotherapy, with or without bevacizumab, is indicated for the treatment of patients with persistent, recurrent, or metastatic cervical cancer whose tumors express PD-L1 (CPS 1) as determined by an FDA-approved test.

KEYTRUDA, as a single agent, is indicated for the treatment of patients with recurrent or metastatic cervical cancer with disease progression on or after chemotherapy whose tumors express PD-L1 (CPS 1) as determined by an FDA-approved test.

Hepatocellular Carcinoma

KEYTRUDA is indicated for the treatment of patients with hepatocellular carcinoma (HCC) who have been previously treated with sorafenib. This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Merkel Cell Carcinoma

KEYTRUDA is indicated for the treatment of adult and pediatric patients with recurrent locally advanced or metastatic Merkel cell carcinoma (MCC). This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Renal Cell Carcinoma

KEYTRUDA, in combination with axitinib, is indicated for the first-line treatment of adult patients with advanced renal cell carcinoma (RCC).

KEYTRUDA is indicated for the adjuvant treatment of patients with RCC at intermediate-high or high risk of recurrence following nephrectomy, or following nephrectomy and resection of metastatic lesions.

Tumor Mutational Burden-High Cancer

KEYTRUDA is indicated for the treatment of adult and pediatric patients with unresectable or metastatic tumor mutational burden-high (TMB-H) [10 mutations/megabase] solid tumors, as determined by an FDA-approved test, that have progressed following prior treatment and who have no satisfactory alternative treatment options. This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials. The safety and effectiveness of KEYTRUDA in pediatric patients with TMB-H central nervous system cancers have not been established.

Cutaneous Squamous Cell Carcinoma

KEYTRUDA is indicated for the treatment of patients with recurrent or metastatic cutaneous squamous cell carcinoma (cSCC) or locally advanced cSCC that is not curable by surgery or radiation.

Triple-Negative Breast Cancer

KEYTRUDA is indicated for the treatment of patients with high-risk early-stage triple-negative breast cancer (TNBC) in combination with chemotherapy as neoadjuvant treatment, and then continued as a single agent as adjuvant treatment after surgery.

KEYTRUDA, in combination with chemotherapy, is indicated for the treatment of patients with locally recurrent unresectable or metastatic TNBC whose tumors express PD-L1 (CPS 10) as determined by an FDA-approved test.

Selected Important Safety Information for KEYTRUDA

Severe and Fatal Immune-Mediated Adverse Reactions

KEYTRUDA is a monoclonal antibody that belongs to a class of drugs that bind to either the PD-1 or the PD-L1, blocking the PD-1/PD-L1 pathway, thereby removing inhibition of the immune response, potentially breaking peripheral tolerance and inducing immune-mediated adverse reactions. Immune-mediated adverse reactions, which may be severe or fatal, can occur in any organ system or tissue, can affect more than one body system simultaneously, and can occur at any time after starting treatment or after discontinuation of treatment. Important immune-mediated adverse reactions listed here may not include all possible severe and fatal immune-mediated adverse reactions.

Monitor patients closely for symptoms and signs that may be clinical manifestations of underlying immune-mediated adverse reactions. Early identification and management are essential to ensure safe use of antiPD-1/PD-L1 treatments. Evaluate liver enzymes, creatinine, and thyroid function at baseline and periodically during treatment. For patients with TNBC treated with KEYTRUDA in the neoadjuvant setting, monitor blood cortisol at baseline, prior to surgery, and as clinically indicated. In cases of suspected immune-mediated adverse reactions, initiate appropriate workup to exclude alternative etiologies, including infection. Institute medical management promptly, including specialty consultation as appropriate.

Withhold or permanently discontinue KEYTRUDA depending on severity of the immune-mediated adverse reaction. In general, if KEYTRUDA requires interruption or discontinuation, administer systemic corticosteroid therapy (1 to 2 mg/kg/day prednisone or equivalent) until improvement to Grade 1 or less. Upon improvement to Grade 1 or less, initiate corticosteroid taper and continue to taper over at least 1 month. Consider administration of other systemic immunosuppressants in patients whose adverse reactions are not controlled with corticosteroid therapy.

Immune-Mediated Pneumonitis

KEYTRUDA can cause immune-mediated pneumonitis. The incidence is higher in patients who have received prior thoracic radiation. Immune-mediated pneumonitis occurred in 3.4% (94/2799) of patients receiving KEYTRUDA, including fatal (0.1%), Grade 4 (0.3%), Grade 3 (0.9%), and Grade 2 (1.3%) reactions. Systemic corticosteroids were required in 67% (63/94) of patients. Pneumonitis led to permanent discontinuation of KEYTRUDA in 1.3% (36) and withholding in 0.9% (26) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 23% had recurrence. Pneumonitis resolved in 59% of the 94 patients.

Pneumonitis occurred in 8% (31/389) of adult patients with cHL receiving KEYTRUDA as a single agent, including Grades 3-4 in 2.3% of patients. Patients received high-dose corticosteroids for a median duration of 10 days (range: 2 days to 53 months). Pneumonitis rates were similar in patients with and without prior thoracic radiation. Pneumonitis led to discontinuation of KEYTRUDA in 5.4% (21) of patients. Of the patients who developed pneumonitis, 42% interrupted KEYTRUDA, 68% discontinued KEYTRUDA, and 77% had resolution.

Immune-Mediated Colitis

KEYTRUDA can cause immune-mediated colitis, which may present with diarrhea. Cytomegalovirus infection/reactivation has been reported in patients with corticosteroid-refractory immune-mediated colitis. In cases of corticosteroid-refractory colitis, consider repeating infectious workup to exclude alternative etiologies. Immune-mediated colitis occurred in 1.7% (48/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (1.1%), and Grade 2 (0.4%) reactions. Systemic corticosteroids were required in 69% (33/48); additional immunosuppressant therapy was required in 4.2% of patients. Colitis led to permanent discontinuation of KEYTRUDA in 0.5% (15) and withholding in 0.5% (13) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 23% had recurrence. Colitis resolved in 85% of the 48 patients.

Hepatotoxicity and Immune-Mediated Hepatitis

KEYTRUDA as a Single Agent

KEYTRUDA can cause immune-mediated hepatitis. Immune-mediated hepatitis occurred in 0.7% (19/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.4%), and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 68% (13/19) of patients; additional immunosuppressant therapy was required in 11% of patients. Hepatitis led to permanent discontinuation of KEYTRUDA in 0.2% (6) and withholding in 0.3% (9) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, none had recurrence. Hepatitis resolved in 79% of the 19 patients.

KEYTRUDA with Axitinib

KEYTRUDA in combination with axitinib can cause hepatic toxicity. Monitor liver enzymes before initiation of and periodically throughout treatment. Consider monitoring more frequently as compared to when the drugs are administered as single agents. For elevated liver enzymes, interrupt KEYTRUDA and axitinib, and consider administering corticosteroids as needed. With the combination of KEYTRUDA and axitinib, Grades 3 and 4 increased alanine aminotransferase (ALT) (20%) and increased aspartate aminotransferase (AST) (13%) were seen at a higher frequency compared to KEYTRUDA alone. Fifty-nine percent of the patients with increased ALT received systemic corticosteroids. In patients with ALT 3 times upper limit of normal (ULN) (Grades 2-4, n=116), ALT resolved to Grades 0-1 in 94%. Among the 92 patients who were rechallenged with either KEYTRUDA (n=3) or axitinib (n=34) administered as a single agent or with both (n=55), recurrence of ALT 3 times ULN was observed in 1 patient receiving KEYTRUDA, 16 patients receiving axitinib, and 24 patients receiving both. All patients with a recurrence of ALT 3 ULN subsequently recovered from the event.

Immune-Mediated Endocrinopathies

Adrenal Insufficiency

KEYTRUDA can cause primary or secondary adrenal insufficiency. For Grade 2 or higher, initiate symptomatic treatment, including hormone replacement as clinically indicated. Withhold KEYTRUDA depending on severity. Adrenal insufficiency occurred in 0.8% (22/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.3%), and Grade 2 (0.3%) reactions. Systemic corticosteroids were required in 77% (17/22) of patients; of these, the majority remained on systemic corticosteroids. Adrenal insufficiency led to permanent discontinuation of KEYTRUDA in <0.1% (1) and withholding in 0.3% (8) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Hypophysitis

KEYTRUDA can cause immune-mediated hypophysitis. Hypophysitis can present with acute symptoms associated with mass effect such as headache, photophobia, or visual field defects. Hypophysitis can cause hypopituitarism. Initiate hormone replacement as indicated. Withhold or permanently discontinue KEYTRUDA depending on severity. Hypophysitis occurred in 0.6% (17/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.3%), and Grade 2 (0.2%) reactions. Systemic corticosteroids were required in 94% (16/17) of patients; of these, the majority remained on systemic corticosteroids. Hypophysitis led to permanent discontinuation of KEYTRUDA in 0.1% (4) and withholding in 0.3% (7) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Thyroid Disorders

KEYTRUDA can cause immune-mediated thyroid disorders. Thyroiditis can present with or without endocrinopathy. Hypothyroidism can follow hyperthyroidism. Initiate hormone replacement for hypothyroidism or institute medical management of hyperthyroidism as clinically indicated. Withhold or permanently discontinue KEYTRUDA depending on severity. Thyroiditis occurred in 0.6% (16/2799) of patients receiving KEYTRUDA, including Grade 2 (0.3%). None discontinued, but KEYTRUDA was withheld in <0.1% (1) of patients.

Hyperthyroidism occurred in 3.4% (96/2799) of patients receiving KEYTRUDA, including Grade 3 (0.1%) and Grade 2 (0.8%). It led to permanent discontinuation of KEYTRUDA in <0.1% (2) and withholding in 0.3% (7) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement. Hypothyroidism occurred in 8% (237/2799) of patients receiving KEYTRUDA, including Grade 3 (0.1%) and Grade 2 (6.2%). It led to permanent discontinuation of KEYTRUDA in <0.1% (1) and withholding in 0.5% (14) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement. The majority of patients with hypothyroidism required long-term thyroid hormone replacement. The incidence of new or worsening hypothyroidism was higher in 1185 patients with HNSCC, occurring in 16% of patients receiving KEYTRUDA as a single agent or in combination with platinum and FU, including Grade 3 (0.3%) hypothyroidism. The incidence of new or worsening hypothyroidism was higher in 389 adult patients with cHL (17%) receiving KEYTRUDA as a single agent, including Grade 1 (6.2%) and Grade 2 (10.8%) hypothyroidism.

Type 1 Diabetes Mellitus (DM), Which Can Present With Diabetic Ketoacidosis

Monitor patients for hyperglycemia or other signs and symptoms of diabetes. Initiate treatment with insulin as clinically indicated. Withhold KEYTRUDA depending on severity. Type 1 DM occurred in 0.2% (6/2799) of patients receiving KEYTRUDA. It led to permanent discontinuation in <0.1% (1) and withholding of KEYTRUDA in <0.1% (1) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Immune-Mediated Nephritis With Renal Dysfunction

KEYTRUDA can cause immune-mediated nephritis. Immune-mediated nephritis occurred in 0.3% (9/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.1%), and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 89% (8/9) of patients. Nephritis led to permanent discontinuation of KEYTRUDA in 0.1% (3) and withholding in 0.1% (3) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, none had recurrence. Nephritis resolved in 56% of the 9 patients.

Immune-Mediated Dermatologic Adverse Reactions

KEYTRUDA can cause immune-mediated rash or dermatitis. Exfoliative dermatitis, including Stevens-Johnson syndrome, drug rash with eosinophilia and systemic symptoms, and toxic epidermal necrolysis, has occurred with antiPD-1/PD-L1 treatments. Topical emollients and/or topical corticosteroids may be adequate to treat mild to moderate nonexfoliative rashes. Withhold or permanently discontinue KEYTRUDA depending on severity. Immune-mediated dermatologic adverse reactions occurred in 1.4% (38/2799) of patients receiving KEYTRUDA, including Grade 3 (1%) and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 40% (15/38) of patients. These reactions led to permanent discontinuation in 0.1% (2) and withholding of KEYTRUDA in 0.6% (16) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 6% had recurrence. The reactions resolved in 79% of the 38 patients.

Other Immune-Mediated Adverse Reactions

The following clinically significant immune-mediated adverse reactions occurred at an incidence of <1% (unless otherwise noted) in patients who received KEYTRUDA or were reported with the use of other antiPD-1/PD-L1 treatments. Severe or fatal cases have been reported for some of these adverse reactions. Cardiac/Vascular: Myocarditis, pericarditis, vasculitis; Nervous System: Meningitis, encephalitis, myelitis and demyelination, myasthenic syndrome/myasthenia gravis (including exacerbation), Guillain-Barr syndrome, nerve paresis, autoimmune neuropathy; Ocular: Uveitis, iritis and other ocular inflammatory toxicities can occur. Some cases can be associated with retinal detachment. Various grades of visual impairment, including blindness, can occur. If uveitis occurs in combination with other immune-mediated adverse reactions, consider a Vogt-Koyanagi-Harada-like syndrome, as this may require treatment with systemic steroids to reduce the risk of permanent vision loss; Gastrointestinal: Pancreatitis, to include increases in serum amylase and lipase levels, gastritis, duodenitis; Musculoskeletal and Connective Tissue: Myositis/polymyositis, rhabdomyolysis (and associated sequelae, including renal failure), arthritis (1.5%), polymyalgia rheumatica; Endocrine: Hypoparathyroidism; Hematologic/Immune: Hemolytic anemia, aplastic anemia, hemophagocytic lymphohistiocytosis, systemic inflammatory response syndrome, histiocytic necrotizing lymphadenitis (Kikuchi lymphadenitis), sarcoidosis, immune thrombocytopenic purpura, solid organ transplant rejection.

Infusion-Related Reactions

KEYTRUDA can cause severe or life-threatening infusion-related reactions, including hypersensitivity and anaphylaxis, which have been reported in 0.2% of 2799 patients receiving KEYTRUDA. Monitor for signs and symptoms of infusion-related reactions. Interrupt or slow the rate of infusion for Grade 1 or Grade 2 reactions. For Grade 3 or Grade 4 reactions, stop infusion and permanently discontinue KEYTRUDA.

Complications of Allogeneic Hematopoietic Stem Cell Transplantation (HSCT)

Read more:
FDA Approves Merck's KEYTRUDA (pembrolizumab) as Adjuvant Therapy for Certain Patients With Renal Cell Carcinoma (RCC) Following Surgery - Business...

Passage Of California Stem Cell Proposition Boosts Research

For the last decade and more, Stem Cell research and regenerative medicine have been the rave of the healthcare industry, a delicate area that has seen steady advancements over the last few years.

The promise of regenerative medicine is simple but profound that one day medical experts will be able to diagnose a problem, remove some of our body cells called stem cells and use them to grow a cure for our ailment. Using our body cells will create a highly personalized therapy attuned to our genes and systems.

The terminologies often used in this field of medicine can get a bit fuzzy for the uninitiated, so in this article, I have relied heavily on the insights of Christian Drapeau, a neurophysiologist and stem cell expert.

Drapeau was one of the first voices who discovered and began to speak about stem cells being the bodys repair system in the early 2000s. Since then, he has gone on to discover the first stem cell mobilizer, and his studies and research delivered the proof of concept that the AFA (Aphanizomenon flos-aquae) extract was capable of enhancing repair from muscle injury.

Christian Drapeau is also the founder of Kalyagen, astem cell research-based company, and the manufacturers of Stemregen. This stem cell mobilizer combines some of the most effective stem cell mobilizers Drapeau has discovered to create an effective treatment for varying diseases.

How exactly do stem cell-based treatments work? And how is it delivering on its promise of boosting our abilities to regenerate or self-heal?

Drapeau explains the concept for us;

Stem cells are mother cells or blank cells produced by the bone marrow. As they are released from the bone marrow stem cells can travel to any organ and tissue of the body, where they can transform into cells of that tissue.Stem cells constitute the repair system of the body.

The discovery of this function has led scientists on a long journey to discover how to use stem cells to cure diseases, which are essentially caused by cellular loss. Diseases like Diabetes and age-related degenerative diseases are all associated with the loss of a type of cell or cellular function.

However, what Drapeaus research has unearthed over the last few decades is that there are naturally occurring substances that show a demonstrated ability to induce the release of stem cells from the bone marrow. These stem cells then enter the bloodstream, from where they can travel to sites of cell deficiency or injury in the body to aid healing and regeneration. This process is referred to as Endogenous Stem Cell Mobilization (ESCM).

Stemregen is our most potent creation so far, explains Drapeau, and it has shown excellent results with the treatment of problems in the endocrine system, muscles, kidneys, respiratory systems, and even with issues of erectile dysfunction.

Despite the stunning advancements that have been made so far, a concern that both Drapeau and I share is how this innovation can be merged with another exciting innovation; AI.

Is it even a possibility? Drapeau, an AI enthusiast, explains that AI has already been a life-saver in stem cell research and has even more potential.

On closer observation, there are a few areas in which AI has greatly benefited stem cell research and regenerative medicine.

One obstacle that scientists have consistently faced with delivering the full promise of regenerative medicine is the complexity of the available data.Cells are so different from each other that scientists can struggle with predicting what the cells will do in any given therapeutic scenario. Scientists are faced with millions of ways that medical therapy could go wrong.

Most AI experts believe that in almost any field, AI can provide a solution whenever there is a problem with data analysis and predictive analysis.

Carl Simon, a biologist at the National Institute of Standards and Technology (NIST) and Nicholas Schaub recentlytested this hypothesiswhen they applied Deep Neural Networks (DNN), an AI program to the data they had collected in their experiments on eye cells. Their research revolved around causes and solutions for age-related eye degeneration. The results were stunning; the AI made only one incorrect prediction about cell changes out of 36 predictions it was asked to make.

Their program learned how to predict cell function in different scenarios and settings from annotated images of cells. It soon could rapidly analyze images of the lab-grown eye tissues to classify the tissues as good or bad. This discovery has raised optimism in the stem cell research space.

Drapeau explains why this is so exciting;

When we talk about stem cells in general, we say stem cells as if they were all one thing, but there are many different types of stem cells.For example, hair follicle and dental pulp stem cells contain neuronal markers and can easily transform into neurons to repair the brain. Furthermore, the tissue undergoing repair must signal to attract stem cells and must secrete compounds to stimulate stem cell function. A complex analysis of the tissue that needs repair and the conditions of that tissue using AI, in any specific individual, will help select the right type of stem cells and the best cells in that stem cell population, along with the accompanying treatment to optimize stem cell-based tissue repair.

Christian Drapeau

Ina study published in Februaryof this year inStem Cells, researchers from Tokyo Medical and Dental University (TMDU) reported that their AI system, called DeepACT, had successfully identified healthy, productive skin stem cells with the same accuracy that a human could. This discovery further strengthens Drapeaus argument on the potentials of AI in this field.

This experiment owes its success to AIs machine learning capabilities, but it is expected that Deep Learning can be beneficially introduced into regenerative medicine.There are many futuristic projections for these possibilities, but many of them are not as far-fetched as they may first seem.

Researchers believe that AI can help fast-track the translation of regenerative medicine into clinical practice; the technology can be used to predict cell behavior in different environments. Therefore, hypothetically, it can be used to simulate the human environment. This means that researchers can gain in-depth information more rapidly.

Perhaps the most daring expectation is the possibility of using AI to pioneer the 3D printing of organs. In a world where organ shortage is a harsh reality, this would certainly come in handy. AI algorithms can be utilized to identify the best materials for artificial organs, understand the anatomic challenges during treatment, and design the organ.

Can stem cells actually be used along with other biological materials to grow functional 3D-printed organs? If this is possible, then pacemakers will soon give way to 3D-printed hearts. A 3D-printedheart valvehas already become a reality in India, making this even more of an imminent possibility.

While all of these possibilities excite Drapeau, he is confident that AIs capabilities with data analysis and prediction, which is already largely in use, would go down as its most beneficial contribution to stem cell research;

It was already shown that stem cells laid on the connective tissue of the heart, the soft skeleton of the heart, can lead the entire formation of a new heart. Stem cells have this enormous regenerative potential. AI can take this to another level by helping establish the conditions in which this type of regeneration can be orchestrated inside the body.But we have to be grateful for what we already have, over the last 20 years, I have studied endogenous stem cell mobilization and today the fact that we have such amazing results with Stemregen is testament that regenerative medicine is already a success.

As AI continues to scale over industry boundaries, we can only sit back and hope it delivers on its full potential promise. Who knows? Perhaps AI really can change the world.

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How The Overlap Between Artificial Intelligence And Stem Cell Research Is Producing Exciting Results - Forbes

Sponsored Article

Although stem cells are known to work wonders, there is still a lot of misunderstanding about what they are, what they do, and how they work.

The good news is that StemCells21 can clear everything up for you. SC21 produces all of its cellular medications in-house, and all of its treatments are performed at its cutting-edge medical centre in Bangkok. Its a one-stop shop that adheres to high-quality standards.

This company will be on display at the Thailand International Boat Show, which will be hosted at Royal Phuket Marina from January 6 to 9 next year. Staff from StemCells21 will be on hand to walk you through the producers, pricing, and techniques.

StemCells21s laboratory is a full-scale culture & analysis laboratory specialising in the production & treatment of Mesenchymal Stem Cells (StemCells21), and Natural Killer Cells (ImmuneCells21). It has also launched a new generation of regenerative medicine called Pluripotent Stem Cells (iPSC21), which hold great potential for impacting chronic diseases in the quest for anti-ageing.

The lab has seven scientists & stem cell researchers, a couple of who have worked with Professor Shinya Yamanaka, who was awarded the Nobel Prize in Physiology or Medicine in 2012 for the discovery that mature cells can be reprogrammed to become pluripotent (iPS cells).

Photo Via: Stemcells 21

Before StemCells21 was created, Managing Director Paul Collier and co-founder Sergei Dmitrievs experienced the power of stem cells either first hand or through the treatment of someone close to them. They knew that stem cells could deliver positive health results, and also knew stem cell treatmentsand the clinics that administered themhad room for improvement.

After deep laboratory investigation, they came to see that most clinics utilised relatively low-quality stem cells and incomplete treatments. While these clinics could deliver a certain level of positive results, they were only scratching the surface of the promise that stem cell treatments could deliver.

Furthermore, the clinics themselves frequently provided a less-than-ideal patient experience. Clinics were generally hectic, unprofessional, and unwelcoming. Patients were often administered a single treatment and sent on their way, unsure if they had experienced an efficacious treatment or if they had travelled and paid for nothing.

StemCells21 was created to offer superior results and give you a welcoming experience. It was set up to provide the global community with access to treatments that few people are aware of, and to offer health benefits that are superior to what most people ever imagined were possible.

The SC21 complex in Bangkok houses the StemCells21, ImmuneCells21, and IPS21 laboratories, as well as the premium 5* IntelliHealth+ (IH+) Clinic.

IntelliHealth+ is a state-of-the-art medical centre licensed by the Thai medical authorities. The luxurious design, efficient workflow layouts, and modern treatments make it the ideal choice for customers seeking a premium level of healthcare in 5* settings.

The centre treats patients from all over the world and has staff who speak fluent English, Arabic, Chinese, Russian, Thai and Spanish.

Furthermore, SC21s come from all corners of the globe for these cutting edge treatments. Many VIPs travel to the clinic including presidents, prime ministers, sports stars, football managers, bank owners and heads of major corporations, many of whom return every six to twelve months and have been doing so for years.

Recently, SC21 treated a ten-year-old British boy who had Ewing sarcoma develop in his arm, which then spread to other areas. He had tried every treatment option in the UK. His trip and treatment were sponsored by UK football teams and the public. Since he started treatment hes put on weight, hes vibrant, and his demeanour has totally changed. Various tests and scans have shown he is responding very well to the immunotherapy course and will perform another round in a few months time.

SC21 focuses on three main areas: anti-ageing and longevity; orthopaedic and muscular-skeletal issues (knee, hip, back & shoulder); and chronic diseases (diabetes, liver cirrhosis, lung, respiratory, hearing & vision disorders). Aside from that, the clinic can also help with chronic fatigue and burn-out syndrome.

Outpatient services for anti-ageing, immunotherapy and regenerative medicine are available at the centre. The anti-ageing clinic has a cutting-edge approach to skin rejuvenation, dermatology, detoxification, and wellbeing. A youthful appearance, more energy, improved mental capacity and mobility, reduced aches and pains, and a stronger immune system are among the benefits.

Photo Via: Stemcells 21

The high level of traditional medicine and the unique protocols designed by the IH+ teams give patients real therapeutic benefits and longevity.

According to Paul Collier, a client typically receives two sessions of stem cell injections during a treatment intravenous for systemic and local to the target and is required to stay in Bangkok for two days following their procedure to monitor any complications that may arise. Then theyre given a two-month take-home kit that comprises self-administered injections (similar to insulin) that target specific growth factors in organs or tissues that need to be repaired. These can also be taken orally, but they are less effective.

He goes on to say that stem cells are the foundation of the human body. They split over and over to produce humans from an embryo at the start of our lives. They restore cells in your blood, bone, skin, and organs throughout your life to keep you alive and functioning. Stem cells have two distinct properties that distinguish them from other types of cells in our bodies.

First, they can self-renew (mitosis), which is a stage of the cell cycle in which replicated chromosomes are divided into two new nuclei. As a result, identical duplicated cells are produced.

Secondly, they have the ability to differentiate into specialized cells such as cartilage, heart cells, liver cells, and neurons. No other cell in the body has the natural ability to generate new cell types.

Mesenchymal Stem Cells (MSCs) are at the core of StemCells21s regenerative programs. They are multipotent stem cells derived from various adult and fetal tissues. A large number of studies have shown the beneficial effects of MSC-based therapies to treat different pathologies, including neurological disorders, cardiac ischemia, diabetes, and bone and cartilage diseases.

StemCells21 also has arthritis treatment, which reduces inflammation & joint pain, increases cartilage growth, improves mobility & joint stability and lessens dependence on medication. The clinics degenerative spine treatments help discs regenerate and stabilize the spine.

On top of that, it provides lung & liver disease treatment as well as treatments for autism, cerebral palsy, diabetes, motor neuron disease, multiple sclerosis and immune disorders.

Theres even eye treatment, which reduces blurred vision & field of vision defects, improves night vision & enhances colour texture.

Photo Via: Stemcells 21

SC21 can even help with certain types of cancer by taking a clients blood and growing their natural killer cells (immunotherapy) over a 21-day period. Through various stimuli, their cytotoxicity is increased which kills cancer and virally-affected cells.

Paul says stem cell therapy should be looked at before undergoing any kind of invasive surgery. The type of medicine should certainly be an intervention before surgery. If you are looking at knee replacement, why not consider an injection of a biologic that would only take a couple of days and has the potential to remodel the cartilage, because once you perform surgery there is no going back.

SC21 also produces a wide range of stem-cell extract-based cosmetics and nutritional supplements, which are available at their medical centres and online under the brand SC21 Biotech.

The Thailand International Boat Show will feature Paul Collier and his team. Theyll be able to answer any of your questions about the cost, procedure, and treatment. On top of that, they will also assist you in educating yourself and managing your expectations so that you do not expect more than stem cell therapy can provide. If you want to get treatment, they will also provide you with a complete report on all treatments. SC21 is fully compliant with international regulations and guidelines.

http://www.stemcells21.com http://www.intellihealthplus.com

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SC21- 21st century cellular medicines specialists - The Thaiger

NEW YORK, Nov. 21, 2021 (GLOBE NEWSWIRE) -- Mesoblast Limited (Nasdaq:MESO; ASX:MSB), global leader in allogeneic cellular medicines for inflammatory diseases, today announced that it has successfully refinanced its existing senior debt facility with a new US$90 million five year facility provided by funds managed by Oaktree Capital Management, L.P. (“Oaktree”).

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Mesoblast and Oaktree Capital Enter Into Refinancing and Expansion of Senior Debt Facility

UW Health treats first patient in U.S. with investigational cell therapy for heart disease

Appleton resident Donald Krause became the first patient in the country last week to undergo an investigational cell therapy for a debilitating heart condition called chronic myocardial ischemia (CMI). Krause was treated by Amish Raval, MD, an interventional cardiologist at UW Health, supported by Peiman Hematti, MD, a bone marrow transplantation hematologist at the UW School of Medicine and Public Health.

October 29, 2021SMPH News

Center members Dr. Anita Bhattacharyy and Dr. Su-Chun Zhang, in collaboration with Waisman and the University of Washington-Seattle and Seattle Childrens Hospital, have been awarded an $11 million Transformative Research grant from the National Institutes of Health to create a new approach using stem cells that may reveal how brain development in individuals with Down syndrome differs from typically developing individuals, identify features that will help understand their intellectual disability, and find potential targets for therapy. They will also address questions that remain unanswered about brain development overall.

October 7th, 2021UW News

The U.S. Food and Drug Administration on Tuesday approved StrataGraft, a topical treatment for severe burns made from skin tissue, providing a boost for Madison-based firm Stratatech. Stratatech was founded in 2000 by SCRMC member Lynn Allen-Hoffman, the first female University of Wisconsin-Madison faculty member to start a biotech company.

June 16, 2021The Cap Times

The Food and Drug Administration-approved trial will use a form of transplant that replaces a patients bone marrow with alpha-beta T-cell depleted peripheral blood stem cells from closely matched unrelated donors or family members.

May 27, 2021

Over the past two decades, stem cell research at UW-Madison has grown from involving a handful of scientists to nearly 100 from more than 30 schools, colleges and departments.

May 25, 2021Quarterly Magazine, Vol. 23, No. 1

Nine University of WisconsinMadison postdoctoral researchers have been recognized with the inaugural Postdoc Excellence Awards for their teaching, service and mentoring. Daniel Z. Radecki (Comparative Biosciences) received one of these awards.

The defining feature of Dans work with the (UWMadison Postdoctoral Association) and others is his commitment to bettering the lives of all postdocs. He envisions how each event and initiative can best impact the individual, through the lenses of diversity and inclusion, immigration status, postdocs personal lives (e.g. childcare considerations), department/discipline, and more.

Congratulations, Daniel!

April 29, 2021

Researchers at UWMadison have made new photoreceptors from human pluripotent stem cells. However, it remains challenging to precisely deliver those photoreceptors within the diseased or damaged eye so that they can form appropriate connections, says David Gamm, director of the McPherson Eye Research Institute and professor of ophthalmology and visual sciences at the UW School of Medicine and Public Health.

While it was a breakthrough to be able to make the spare parts these photoreceptors its still necessary to get them to the right spot so they can effectively reconstruct the retina, he says. So, we started thinking, How can we deliver these cells in a more intelligent way? Thats when we reached out to our world-class engineers at UWMadison.

Research from the University of WisconsinMadison finds that a new therapeutic approach for heart failure could help restore cardiac function by regenerating heart muscle. In a study recently published in the journal Circulation, the UW team describes its success in improving, in a mouse model, the function of heart muscle by temporarily blocking a key metabolic enzyme after a heart attack. This simple intervention, the researchers say, could ultimately help people regain cardiac function. Our goal was to gain new understanding of how the heart can heal itself following injury at the molecular and cellular level and see if there was a way to restore cardiac function to an earlier state, says UWMadisons Ahmed Mahmoud, professor of cell and regenerative biology in the School of Medicine and Public Health.

Learn more about the research here.April 15, 2021

Grafting neurons grown from monkeys own cells into their brains relieved the debilitating movement and depression symptoms associated with Parkinsons disease, researchers at the University of WisconsinMadison reported today. In a study published in the journal Nature Medicine the UW team describes its success with neurons made from cells from the monkeys own bodies after reprogramming to induced pluripotent stem cells. UWMadison neuroscientist Su-Chun Zhang, whose Waisman Center lab grew the brain cells, said this approach avoided complications with the primates immune systems and takes an important step toward a treatment for millions of human Parkinsons patients. Learn more about their work here.March 1, 2021

The project, led by David Gamm, MD, PhD, director of the McPherson Eye Research Institute and professor of ophthalmology and visual sciences at the UW School of Medicine and Public Health, will develop a transplantable patch to restore vision to members of the armed forces who have been injured by blasts or lasers.December 11, 2020

This week, the NIH Office of Research Infrastructure Programs highlights Dr. Marina Emborg, her WNPRC lab team and their UWMadison colleagues advances in detecting heart disease in Parkinsons and evaluating new therapies that specifically target nerve disease within the human heart.December 2020

Its been 25 years since University of WisconsinMadison scientist James Thomson became the first in the world to successfully isolate and culture primate embryonic stem cells. He accomplished this breakthrough first with nonhuman primates at the Wisconsin National Primate Research Center in 1995, using rhesus monkey cells, then in 1996 with marmoset cells. Thomson then published his world-changing breakthrough on human embryonic stem cell derivation in Science on Nov. 6, 1998.November 6, 2020

EEMs and exosomes each have attractive characteristics as therapeutics, Dr. Hematti, UW-Madisons Department of Medicine, noted. As a cell therapy, EEMs will not proliferate or differentiate to undesirable cell types, which remains a concern for many stem cell therapies. Moreover, EEMs could be generated from a patients own monocytes using off-the-shelf exosomes, resulting in a faster and more facile process compared to autologous MSCs. Alternatively, exosome therapy could be a cell free, shelf-stable therapeutic to deliver biologically active components. Altogether, we believe our studies results support the use of EEMs and/or exosomes to improve ligament healing by modulating inflammation and tissue remodeling, Dr. Vanderby concluded.November 3, 2020

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Our Need For Things Lab-Grown

What was once something of the movies objects forming themselves in thin air is real now. Various things can be grown in a laboratory setting, some even on a large scale for commercial distribution. This technology could be a big part of the solution to establish sustainable societies. At the moment, we harvest organs from the deceased, rear animals for meat and dairy, destroy forests by cutting down trees for wood, mine the earth for diamonds, and the list goes on. All these things can already be lab-made or are on the brink of reality.

Once these staples of society can be mass-made affordably, they could supply the world while minimally impacting the natural environment. Acres of land wouldnt need to be used for food and building materials, meaning deforestation can cease, for starters. Looking at lab-grown meats alone: they require 99% less land than traditionally farmed meats, generate up to 96% fewer emissions, use up to 96% less water, and no animals need to be slaughtered in the process.

Naturally, there will be short-term disruptions, particularly job-related. For example, eco-friendly agriculture will mean fewer farms and agriculture jobs. But new employment opportunities will emerge in the scientific and technical fields related to lab-grown foods.

Whats the difference between 3D printing (additive manufacturing) and lab-grown, you may be wondering? 3D printing uses material as ink anything from plastic to cellular material whereas lab-grown materials start off as a bit of material that multiplies on its own, replicating natural processes. Thus, lab-grown material has the same cellular structure as the naturally occurring material and mimics the natural formation process but within a much shorter period.

In the future, we are bound to see various lab-grown breakthroughs coming from the medical field. Eventually, there should be alternative sources for organs and blood cultured from stem cells. In addition, there will likely be lab-produced medicines (lotions, ointments, balms, nutraceuticals, energy drinks, etc.), breast milk, and more.

Scientists are well on the way to functioning full-sized organs, with several innovations in fully functional mini-organs, or organoids, making headlines in recent years. For now, these organoids are tools for testing new drugs and studying human diseases. But soon enough, these research teams will take the technology to the next level and develop organs that can be used for implantation when someone needs an organ replacement. So far, the brain, liver, lungs, thymus, heart, blood, and blood vessels are among the growing list of lab-grown medical accomplishments.

A team of scientists from the University of Pittsburgh managed to grow miniature human livers using induced pluripotent stem cells (IPSCs) made from human skin cells. Meaning, in the far future, someone needing a liver transplant could have the organ grown from their own skin cells! This method may even reduce the chances of a patients immune system rejecting the new tissue because it would recognize the cells as self. Whats more, their lab-grown livers matured in under a month compared to two years in a natural environment.

The scientists tested their fully-functional mini-livers by transplanting them into rats. In this proof-of-concept study, the lab-made organs survived for four days inside their animal hosts, secreting bile acids and urea like a healthy liver would.

A research team led by the University Hospital Dsseldorf induced pluripotent stem cells (iPSCs) to grow into pea-sized brain organoids with rudimentary eye structures that sense light and send signals to the rest of the brain. They used skin cells taken from adult donors, reverted them back into stem cells, and placed them into a culture mimicking a developing brains environment, which encourages them to form specific brain cells. Their mini-brains grew optic cups, vision structures of the eye found where the optic nerve and retina meet. The cups even grew symmetrically, as eyes would, and were functional!

Jay Gopalakrishnan, a senior author of the study, said:

Our work highlights the remarkable ability of brain organoids to generate primitive sensory structures that are light sensitive and harbor cell types similar to those found in the body. These organoids can help to study brain-eye interactions during embryo development, model congenital retinal disorders, and generate patient-specific retinal cell types for personalized drug testing and transplantation therapies.

This achievement is the first time an in vitro system shows nerve fibers of retinal ganglion cells reaching out to connect with their brain target an essential aspect of the mammalian brain.

Scientists from Michigan State University developed functional miniature human heart models grown from stem cells complete with all primary heart cell types and with functioning chambers and vascular tissue. The models could help researchers better understand how hearts develop and provide an ethical platform for treating disease and testing drugs or new treatments.

The teams lab-grown mini hearts follow the fetal development of a human heart, offering a new view into that process. The organoids start beating by day six, and they grow into spheres approximately 1 mm (0.4 in) wide, with all significant cardiac cell types and multiple internal chambers by day 15.

Aside from research purposes, full-sized lab-grown hearts could solve the shortage problem of hearts the world faces today. More than 25 million people suffer heart failure each year. In the United States, approximately 2,500 of the 4,000 people in line for heart transplants receive them. That means almost 50% of the people needing a new heart to keep them alive wont get it.

Unlimited supplies of blood for transfusions are possible with lab-growing technology. Blood has been challenging to grow in the lab. However, real breakthroughs in creating artificial blood have sprung up!

A couple of years ago, Japanese researchers developed universal artificial blood that worked for all blood types. It even has a shelf life of one year stored at room temperature, therefore eliminating the problem of identifying blood type and storage simultaneously.

Like that wasnt impressive enough, last year, a team of scientists from the South China University of Technology, the University of New Mexico, and Sandia National Laboratories created artificial red blood cells (RBCs) with more potential capabilities than real ones! The synthetic RBCs mimic the properties of natural ones such as oxygen transport, flexibility, and long circulation times with the addition of a few new tricks up their sleeves, such as toxin detection, magnetic targeting, and therapeutic drug delivery. In addition, blood contains platelets and red blood cells, so these new cells could be used to make superior artificial blood.

Researchers from the University of British Columbia successfully coaxed stem cells to grow into human blood vessels. The thing that is so remarkable about this study is that the system of blood vessels grown in the lab is virtually identical to the ones currently transporting blood throughout the body. They are using this now to generate new leads in diabetes treatment. They put the lab-grown blood vessels in a petri dish designed to mimic a diabetic environment.

The global demand for meat and dairy is expected to rise by almost 90% over the next 30 years, regardless of the need to cut back on meat consumption. The risk of environmental damage and the rising food demand itself is a problem many have recently addressed. Thats why companies worldwide are on the verge of scaling up all sorts of lab processes to produce various food items, including steaks, chicken, cheese, milk, ice cream, fruits, and more.

Thinktank RethinkX even published research suggesting that proteins from precision fermentation (lab-grown protein using microbes) will be about ten times cheaper than animal protein by 2035, resulting in a collapse of the livestock industry. It says the new food economy will subsequently:

replace an extravagantly inefficient system that requires enormous quantities of inputs and produces considerable amounts of waste with one that is precise, targeted, and tractable. [Using tiny land areas, with a massively reduced requirement for water and nutrients, it] presents the most significant opportunity for environmental restoration in human historyFarm-free food offers hope where hope is missing. We will soon be able to feed the world without devouring it.

The worlds pace of meat consumption is placing a significant strain on the environment. Many studies show that eating less meat is just as crucial to slowing down global warming as using solar panels and zero-emissions vehicles. Unfortunately, animal farming generates an obscene amount of greenhouse gas emissions. Yet again, scientists come to the rescue, working diligently to fix this situation.

Over a decade ago, researchers created something akin to ground beef, but the complex structure of steak didnt happen until recently, with Aleph Farms debuting its thick-cut rib-eye steak in 2018. Furthermore, that first burger cost around US$345,000, but now the price has dropped dramatically to the point that lab-grown chicken is to be commercially produced and hit grocery store shelves as of this year.

SuperMeat, Eat Just, and Aleph Farms are todays most prominent startups working on getting lab-grown meats to people looking to lower their carbon and environmental footprints. In addition, their products are made from actual animal cells, so theyre real meat, but no animals had to be hurt or killed.

Speaking of Aleph Farms, the company also grew meat in space to show that it can even be done in a zero-gravity environment with limited resources.

Aside from Aleph Farms figuring out how to make steak like an authentic steak, a group of Harvard John A. Paulson School of Engineering and Applied Sciences (SEAS) researchers also devised a solution to the texture challenge. First, they made edible gelatin scaffolds that have the texture and consistency of real meat. Then, they grew rabbit and cow muscle cells on this scaffolding. The research demonstrates how realistic meat products are possible!

Parker and his Disease Biophysics Group developed a technique to produce the scaffolding. Its a fiber-production system inspired by cotton candy known as immersion Rotary Jet-Spinning (iRJS). It enabled the team to spin long nanofibers of a specific shape and size using centrifugal force. So, they spun food-safe gelatin fibers, creating the base upon which cells could grow.

Natural muscle tissue is composed of an extracellular matrix, which is the glue that holds the tissue together. As a result, it contributes to the texture of the meat. The spun gelatin fibers mimicked this extracellular matrix and provided the texture to make the lab-grown meat realistic. When the team seeded the fibers with animal (rabbit and cow) muscle cells, they anchored to the gelatin scaffolding and grew in long, thin structures, similar to real meat.

Meanwhile, Boston College developed a new, even greener technology that uses the skeleton of spinach leaves to support bovine animal protein growth. However, animal products arent eliminated from the process entirely. For example, lab-grown steak and chicken are created by painlessly harvesting muscle cells from a living cow, subsequently fed and nurtured to multiply and develop muscle tissue. But for this to have the same texture as real meat, the cells need structural support to flourish and are therefore placed in a scaffold.

Singapore is leading the way, becoming the first country in the world to approve the sale of Eat Justs cultured chicken. The company will start by selling nuggets at a restaurant. Meanwhile, SuperMeat has been handing out lab-grown chicken burgers in Israel for free. Theyre aiming to gain public acceptance of the idea.

The cultured chicken starts as a tiny number of harvested cells. Those cells are put into a bioreactor and fed the same nutrients the living animal would consume to grow. The cells multiply and turn into an edible portion of cultured chicken meat. The meats composition is identical to that of real chicken and offers the same nutritional value. And its cleaner because its antibiotic-free!

Labs are manufacturing dairy products by utilizing the fermentation process of living microbes to produce dairy proteins like whey and casein. These proteins are then used to make dairy products like butter, cheese, and ice cream. Two leading companies in this category are Imagindairy and Perfect Day, which already have several products on supermarket shelves in the United States.

Researchers havent figured out how to make fruits and vegetables yet, but a team is perfecting a cell cultivation process that generates plant biomass. The stuff tastes like the natural-grown product from which the cells were obtained and even exceeded its nutritional properties. Although, the texture of the biomass is different. For example, an apple isnt a solid apple akin to one grown from a tree. Instead, its like applesauce.

Lab-produced materials Including wood, diamonds, leather, glass, clothing, crystals, gels, cardboard, and plastics for making objects are either under development or already available. Many materials need to be taken from nature mined from the earth or cut down from forests. If they can be made in a lab instead, then people could leave nature alone!

A recent project led by a Ph.D. student at MIT paves the way for lab-grown wood one of the worlds most vital resources used to make paper, build houses, heat buildings, and so much more. The process begins with live plant cells cultivated in a growth medium coaxed using plant hormones to become wood-like structures. Next, a gel matrix is used to guide the shape of the cellular growth, and controlling the levels of plant hormones regulates the structural characteristics. Therefore, the technology could grow anything from tables and chairs to doors to boats and so on.

The environmental and socio-economic impact of traditionally mined diamonds has been exposed in recent years, and as awareness grows, the rising popularity of lab-grown diamonds does too. Mined diamonds are linked to bloody conflicts, and their excavation produces carbon emissions, requires substantial water use, and causes severe land disturbances.

Research has found that 1,000 tons of earth have to be shifted, 3,890 liters or more of water is used, and 108kg of carbon is emitted per one-carat stone produced. In addition, the traditional diamond mining industry causes irreversible damage to the environment, hence why, a decade ago, researchers started experimenting with how to grow them in the lab. Its been a feat a long time in the making, but we finally have lab-grown diamonds available for eco-conscious consumers to buy.

Diamonds are made of pure carbon. It takes extreme heat and pressure for carbon to crystalize. In nature, this happens hundreds of miles beneath the Earths surface. The ones being mined were shot out by a volcano millions of years ago. So how have scientists managed to hack such an intense and time-consuming process?

They began by investigating the mechanisms behind the diamond formation, zooming in at the atomic level. This led to the invention of a novel technology that utilizes the process of HPHT (high pressure, high temperature) to mimic the natural atmospheric conditions of diamond formation. Labs can use it to replicate the process and turn pure carbon into diamonds in 2-6 weeks.

Lab-grown gems are eco-friendly rocks, especially when theyre made entirely from the sky, like SkyDiamonds. Even the electricity used to grow its stones is from renewables, so theyll indeed be the worlds first zero-impact diamonds.

But how are the diamonds created out of thin air? They are made of carbon from the sky and rainwater. The sky mining facility is in Stroud. Energy is sourced from wind and sunlight. The CO2 is sourced directly from the air. Hydrogen is produced by splitting rainwater molecules in an electrolysis machine using renewable energy. The captured carbon and hydrogen are then used to make methane, used to grow the diamonds. The final product is a diamond anatomically identical to those mined from the ground. It is even accredited, fully certified, and graded by the International Gemological Institute.

Another company, Climeworks, is also making diamonds using carbon sucked from the sky. However, SkyDiamonds takes it a step forward by using rainwater and sunshine in the process.

The last lab-grown object were going to discuss is not something in the works, but an idea a fantastic and outlandish one thats jumping far into the future but was thought up in 2010 by Mercedes Benz. The luxury car companys ambitious BIOME idea shows just how wild imagination can get with lab-grown technology. It envisions a day when it can grow an entire supercar from scratch.

Mercedes-Benz explained when launching the concept:

The interior of the BIOME grows from the DNA in the Mercedes star on the front of the vehicle, while the exterior grows from the star on the rear. The Mercedes star is genetically engineered in each case to accommodate specific customer requirements, and the vehicle grows when the genetic code is combined with the seed capsule. The wheels are grown from four separate seeds.

This list of lab-grown possibilities is just the tip of the iceberg! Other materials in the pipeline include leather, chocolate, and silk. This intelligent technology can make anything a scientist can dream up! The only limit is the imagination and dedication of brilliant people.

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Lab-Growing Everything Might Be The Only Way To Attain A Sustainable World - Intelligent Living

Phovia is highly effective for treating superficial and deep skin infections.

This article is sponsored by Vetoquinol.

Superficial bacterial folliculitis, also called superficial pyoderma, is a commonly diagnosed dermatological condition in dogs.1,2 These infections are secondary to primary conditions affecting normal skin barrier function (eg, allergic skin disease, trauma, burns), keratinization (eg, nutritional deficiency, liver disease), and immune regulation (eg, neoplasia, autoimmunity, endocrinopathy).2 Cats less commonly develop superficial pyoderma perhaps because of decreased adhesion of staphylococci to feline corneocytes, but the primary issues causing infection are similar to those seen in dogs.3-8

The primary pathogen associated with superficial pyoderma in dogs and cats is a normal resident of the skin, Staphylococcus pseudintermedius, but other flora may be involved.2,8-12 As the normal homeostasis of this organism is disrupted from a primary disease, these gram-positive cocci invade deeper regions of the epidermis and hair follicle epithelium, increase in number, and enhance inflammation.

Classical clinical lesions of superficial pyoderma include papules and pustules that may eventually progress to alopecia, epidermal collarettes, scales, and crusts. Often the skin is erythematous and pruritic. Chronic cases may demonstrate lichenification, hyperpigmentation, and scarring alopecia from long-standing inflammation and infection.2 Cats may develop even more unique cutaneous reaction patterns and skin lesionsespecially when allergic skin disease is presentincluding miliary dermatitis, eosinophilic plaques, rodent ulcers, and eosinophilic granulomas.5

Identifying and addressing the primary disease is paramount in achieving complete, permanent resolution of the superficial pyoderma. Therefore, treatment is multifactorial and aimed at addressing the primary disease, reducing skin inflammation, and treating the infection directly. Current guidelines for the treatment of superficial pyoderma in dogs recommend the use of topical antimicrobials as sole therapy whenever possible; however, overuse of systemic antibiotics remains common.2,13-16

Topical therapy has many benefits including direct antimicrobial effects without use of an antibiotic, reduction in antibiotic-resistant bacterial populations, restoration of the normal skin barrier, enhancement of skin hydration, physical removal of keratinous debris, and removal of offending allergens from the haircoat.2,14 However, topical therapy is met with challenges that impede clinical application. Adherence is the biggest concern when recommending topical therapy to pet owners. Frequent bathing or application of medicated solutions to the skin can be difficult when busy owner lifestyles combine with a nonadherent patient. Skin inflammation can be painful and animals may be resistant to topical therapy. Cats are fastidious groomers and may lick away a medicated topical therapy before it can achieve appropriate contact time. Additionally, some topical agents can cause oral erosions and ulcerations or even gastrointestinal disturbance when groomed off. For these reasons, systemic antibiotics continue to be a common prescribing practice for superficial pyoderma.

All antibiotic use, despite duration or frequency, contributes to the development of antibiotic-resistant bacterial populations on the animal and in the environment.17-19 From that very first dose, bacteria are constantly evolving to implement inherent and acquired resistance mechanisms necessary for survival. One well-recognized mechanism is oxacillin resistance through the mecA gene, which produces a penicillin-binding protein receptor with poor affinity for -lactam antibiotics.2,14,15,20-23 Even more concerning than these oxacillin-resistant strains are those that develop multidrug resistance, which is defined as resistance to 3 or more antibiotic drug classes. This may happen over time with repeated antibiotic exposure or after a single dose of certain antibiotics such as fluorinated quinolones.2,20,23-25 The continued emergence of antibiotic-resistant bacteria inhibits the successful treatment of bacterial infections in pets and humans. As veterinarians consider how their antibiotic use contributes to this growing pandemic, they must look for alternative, safe, effective, affordable, and convenient antibacterial treatment modalities.

Phovia as a solution

Investigation into the photobiological effects of light therapy has been ongoing for the past 50 years. Photobiomodulation (PBM) therapy is a type of light treatment that uses visible or near infrared light to promote therapeutic benefits including induction of tissue healing and regeneration and inhibition of biological responses that induce pain or inflammation. The treatment distance, wavelength, fluence, pulse parameters, spot size, and irradiation time influence the effects of light energy on tissue. Visible light with wavelengths ranging from 400 to 700 nm can stimulate positive photobiomodulatory effects that promote wound healing, reduce inflammation and pain, modulate stem cell populations, and reduce bacterial contamination of wounds.26,27

Once visible light enters the skin, it is absorbed by the cells and initiates chemical changes dependent on the wavelength (or color) of light and the chromophore within the skin.27 Within each cell, membrane-bound organelles called mitochondria contain chromophores that absorb the light energy and begin making energy (adenosine triphosphate; ATP) via activation of cytochrome c oxidase. Outcomes of the mitochondrial respiratory pathway activation include stimulation of secondary messenger pathways, production of transcription factors and growth factors, and increased ATP production. However, excessive light energy exposure will overstimulate mitochondrial respiration and cause expenditure of all ATP reserves, which creates oxidative stress resulting in damaging elevations of nitric oxide, production of harmful free radicals, and activation of cytotoxic mitochondrial-signaling pathways leading to apoptosis.27,28 This is why creating PBM therapy protocols is important for targeting the beneficial effects while avoiding unintended harm.

Specific benefits of light energy within the visible light spectrum can be broken down into each color of light. Blue light (400-500 nm) has a lower penetration depth and primarily interacts with keratinocytes, reduces bacterial adhesion and growth, and increases intracellular calcium and osteoblast differentiation.29-31 Green light (495-570 nm) affects the superficial tissue and alters melanogenesis, reduces hyperpigmentation of the skin, and reduces tissue swelling.29,30 Red light (600-750 nm) penetrates deeper into the dermis and subcutis where it acts on cellular mitochondria to reduce inflammation and promote collagen synthesis through fibroblast proliferation and production of transforming growth factor-, fibroblast growth factor, platelet derived growth factor, and others.26-28,32,33 Red light has proliferative effects on mesenchymal stem cells and induces proliferation of epithelial colony forming units important for tissue repair and regeneration.34,35

Phovia, sold by Vetoquinol, is a form of fluorescent PBM therapy utilizing a blue light emitting diode (LED lamp, 400-460 nm) and topical photoconverter gel that emits low-energy fluorescent light (510-600 nm) when illuminated by the LED lamp.36,37 This interaction results in the formation of multiple wavelengths of visible light, each with a unique depth of penetration and effect on the tissue as described above. Application is fast and simple. The affected skin may be clipped free of hair and cellular debris removed with gentle cleaning. The skin is allowed to dry before application of the photoconverter gel. Just prior to application, 1 ampule of fluorescence chromophore gel is added to 1 container of photoconverter carrier gel and mixed thoroughly. The mixture is applied in a 2-mm layer to the affected skin, and the LED lamp is held 5 cm above the lesion and used to illuminate the area for 2 minutes. The gel is wiped away using saline-soaked gauze. The application can be repeated immediately after 5 to 10 minutes of rest or a second application can occur a few days later. Twice-weekly applications are continued until the wound is healed. Appropriate eyewear is required to protect the operator from the intensely bright light. Application is pain free and stress free for the patient, so sedation is not typically required.

Benefits of Phovia

Phovia shows great promise as a safe, effective therapy for treatment of numerous inflammatory dermatoses in dogs including superficial pyoderma,38 deep pyoderma,39 perianal fistula,40 interdigital dermatitis,41 calcinosis cutis,42 acute traumatic wounds,43 chronic wounds,37 surgical wounds,44 and otitis externa.45 Phovia as a sole therapy speeds time to healing by 36% in canine superficial pyoderma as compared with dogs receiving oral antibiotics alone.38 In one study, dogs with superficial pyoderma were treated with Phovia alone or with an oral antibiotic alone. Dogs treated twice weekly with Phovia demonstrated complete clinical healing in about 2.3 weeks (P < .05)whereas dogs receiving oral antibiotic healed in about 3.75 weeks.38 Additionally, Phovia speeds time to healing by nearly 50% in deep pyoderma when used with an oral antibiotic (5.7 weeks of treatment) compared with dogs receiving only oral antibiotic (11.7 weeks of treatment).39 The ability of this fluorescent PBM therapy to eliminate or significantly reduce duration of exposure to antibiotics will decrease the spread of antibiotic-resistant bacterial strains within pets and humans.

Phovias high safety profile makes it a beneficial tool to implement in everyday practice. Training the veterinary team to communicate therapy benefits with clients as well as to perform treatments is fast and easy. Training the veterinary technicians to perform treatments will give the veterinarian time to examine other patients. A single back-to-back application takes about 15 minutes, so pet owners can be in and out of the clinic quickly; however, the 2 weekly treatments can be separated by a few days if the veterinarian prefers to evaluate the patient more frequently. Additionally, when used as a sole therapy, clients are not required to administer oral or topical medications at home. This greatly improves treatment adherence and success. Instruct clients to use once-daily smartphone photos to document improvement at home. This can be useful when deciding how many treatments to perform. Most cases of superficial pyoderma will resolve completely by the third treatment.38 It is a good idea to communicate to clients that 3 to 4 weekly treatments may be required.

Conclusion

Phovia is a versatile, innovative therapeutic approach to numerous types of dermatitis.36 It is easy to implement in general practice, and is safe, pain free, and affordable. Phovia is highly effective for superficial and deep skin infections and eliminates the need for clients to administer numerous at-home treatments. This greatly improves the pet-owner bond and treatment outcomes by promoting adherence. Phovia accelerates time to wound healing, which decreases duration of antibiotic exposure and may reduce risk of antibiotic resistance development in these cases.2,13,36-39 Phovias efficacy against antibiotic-susceptible and antibiotic-resistant bacteria shows promise as an alternative therapeutic approach that promotes the principles of antimicrobial stewardship.36 If you are interested in purchasing this medical device for your practice, contact your Vetoquinol service representative.

Amelia G. White, DVM, MS, DACVD is an associate clinical professor of dermatology at Auburn University College of Veterinary Medicine.

REFERENCES

Original post:
Managing superficial pyoderma with light therapy - DVM 360

image:Dr. Ulrich G. Steidl view more

Credit: Albert Einstein College of Medicine

October 27, 2021(BRONX, NY)Ulrich G. Steidl, M.D., Ph.D., co-director of the Blood Cancer Institute and associate director of basic science at the Albert Einstein Cancer Center (AECC), has received a prestigious Outstanding Investigator Award from the National Cancer Institute (NCI). This award is accompanied by a seven-year, $7 million grant to study the molecular and cellular mechanisms that lead to two related blood diseases, myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Dr. Steidl is one of only 17 recipients of this award in 2021, which is given to accomplished leaders in cancer research who provide significant contributions in their field. The ultimate goal of this research is to develop new treatments and cures for these usually fatal disorders.

Clinical outcomes in MDS and AML have not significantly improved over the past half-century, and cure rates remain below 15% for most patients, said Dr. Steidl, who is also professor of cell biology and of medicine and the Diane and Arthur B. Belfer Faculty Scholar in Cancer Research at Einstein. There is an urgent need to improve our understanding of how these diseases develop and to devise more effective therapies.

MDS and AML Explained

MDS occurs when blood-forming (hematopoietic) stem cells in the bone marrow acquire genetic and non-genetic irregularities, leading to the production of abnormal, dysfunctional blood cells, which out-compete healthy cells. Common symptoms include anemia, infections, and bleeding.

The incidence of MDS in the United States is unclear, with estimates ranging from 10,000 to 40,000 new cases annually; about one-third of MDS patients will go on to develop AML. Treatment for MDS is generally limited to preventing or reducing complications, particularly severe anemia. The only cure is a bone-marrow transplanta therapy not easily tolerated and therefore often reserved for the youngest, most resilient patients. However, most people diagnosed with MDS are elderly.

AML, like MDS, begins with abnormal bone marrow stem cells. But in AML, those cells, after becoming cancerous, proliferate rapidly and quickly spread to the blood and other hematopoietic organs, such as the bone marrow and spleen, and sometimes to other tissues, causing many of the same symptoms seen in MDS, plus others. AML is often fatal within just a few months and afflicts about 21,000 Americans each year. It is usually treated with chemotherapy. Bone-marrow transplantation can cure AML in some patients.

From Stem Cells to Cancer

Recent studies led by Dr. Steidl and his research team have shown that both MDS and AML arise from pre-leukemic stems cells (pre-LSCs), a subpopulation of blood-forming stem cells that have genetic and non-genetic aberrations. Certain varieties (clones) of these pre-LSCs go on to develop into leukemic stem cells (LSCs)cancer cells that are capable of self-renewal. These LSCs lead to sustained leukemia growth and are particularly resistant to drugs. We now know that the considerable diversity of pre-LSC clones affects the development, progression, and treatment resistance of both MDS and AML, said Dr. Steidl, one of the nations leading authorities on both diseases.

What causes some pre-LSCs but not others to become leukemic is not clear, but transcription factors are thought to play a key role. Transcription factors are proteins that turn specific genes on or off, determining a cells function by regulating the activity of genes. In the case of stem cells, transcription factors guide their differentiation into mature cells. Our recent work has shown that the actions of key transcription factors are dysregulated in pre-LSCs and LSCs, meaning that the transcription factors and the molecular programs they govern behave abnormally, he added.

Thanks to his new NCI grant, Dr. Steidl hopes to:

To accomplish these goals, Dr. Steidls research team will employ novel tools for analyzing stem cell clones in patients, as well as newly developed mouse models of pre-LSC progression to MDS and AML.

Developing New Cancer Therapies

The knowledge we gain from this research should enable us to develop drugs that target pre-LSCs and their aberrant transcription factors, said Dr. Steidl. Such an approach holds the promise of achieving lasting remissions and, ultimately, even cures. Hopefully, our understanding of the early events in the progression of MDS and AML may even allow us in the future to prevent these diseases by interrupting the transformation of pre-LSCs to LSCs before overt leukemia can occur.

The grant (R35CA253127) is titled Molecular and Cellular Regulation of Pre-Leukemic Stem Cells and their Therapeutic Targeting.

***

About Albert Einstein College of Medicine

Albert Einstein College of Medicine is one of the nations premier centers for research, medical education and clinical investigation. During the 2020-21 academic year, Einstein is home to 721 M.D. students, 178 Ph.D. students, 109 students in the combined M.D./Ph.D. program, and 265 postdoctoral research fellows. The College of Medicine has more than 1,900 full-time faculty members located on the main campus and at its clinical affiliates. In 2020, Einstein received more than $197 million in awards from the National Institutes of Health (NIH). This includes the funding of major research centers at Einstein in aging, intellectual development disorders, diabetes, cancer, clinical and translational research, liver disease, and AIDS. Other areas where the College of Medicine is concentrating its efforts include developmental brain research, neuroscience, cardiac disease, and initiatives to reduce and eliminate ethnic and racial health disparities. Its partnership with Montefiore, the University Hospital and academic medical center for Einstein, advances clinical and translational research to accelerate the pace at which new discoveries become the treatments and therapies that benefit patients. Einstein runs one of the largest residency and fellowship training programs in the medical and dental professions in the United States through Montefiore and an affiliation network involving hospitals and medical centers in the Bronx, Brooklyn and on Long Island. For more information, please visit einsteinmed.org, read our blog, followus on Twitter, like us on Facebook, and view us on YouTube.

Disclaimer: AAAS and EurekAlert! are not responsible for the accuracy of news releases posted to EurekAlert! by contributing institutions or for the use of any information through the EurekAlert system.

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Albert Einstein Cancer Center researcher receives NCI Outstanding Investigator Award to study two deadly blood diseases - EurekAlert

Curr Cardiol Rev. 2013 Feb; 9(1): 6372.

1Department of Medicine, University of Minnesota Medical School, Minneapolis, Minnesota, USA

2Stem Cell Institute, University of Minnesota Medical School, Minneapolis, Minnesota, USA

1Department of Medicine, University of Minnesota Medical School, Minneapolis, Minnesota, USA

1Department of Medicine, University of Minnesota Medical School, Minneapolis, Minnesota, USA

2Stem Cell Institute, University of Minnesota Medical School, Minneapolis, Minnesota, USA

3Department of Biomedical Engineering, University of Minnesota, Minneapolis, Minnesota, USA

1Department of Medicine, University of Minnesota Medical School, Minneapolis, Minnesota, USA

2Stem Cell Institute, University of Minnesota Medical School, Minneapolis, Minnesota, USA

3Department of Biomedical Engineering, University of Minnesota, Minneapolis, Minnesota, USA

Received 2012 Jun 11; Revised 2012 Jul 31; Accepted 2012 Aug 27.

Induced pluripotent stem (iPS) cells, are a type of pluripotent stem cell derived from adult somatic cells. They have been reprogrammed through inducing genes and factors to be pluripotent. iPS cells are similar to embryonic stem (ES) cells in many aspects. This review summarizes the recent progresses in iPS cell reprogramming and iPS cell based therapy, and describe patient specific iPS cells as a disease model at length in the light of the literature. This review also analyzes and discusses the problems and considerations of iPS cell therapy in the clinical perspective for the treatment of disease.

Keywords: Cellular therapy, disease model, embryonic stem cells, induced pluripotent stem cells, reprogramm.

Induced pluripotent stem (iPS) cells, are a type of pluripotent stem cell derived from adult somatic cells that have been genetically reprogrammed to an embryonic stem (ES) cell-like state through the forced expression of genes and factors important for maintaining the defining properties of ES cells.

Mouse iPS cells from mouse fibroblasts were first reported in 2006 by the Yamanaka lab at Kyoto University [1]. Human iPS cells were first independently produced by Yamanakas and Thomsons groups from human fibroblasts in late 2007 [2, 3]. iPS cells are similar to ES cells in many aspects, including the expression of ES cell markers, chromatin methylation patterns, embryoid body formation, teratoma formation, viable chimera formation, pluripotency and the ability to contribute to many different tissues in vitro.

The breakthrough discovery of iPS cells allow researchers to obtain pluripotent stem cells without the controversial use of embryos, providing a novel and powerful method to "de-differentiate" cells whose developmental fates had been traditionally assumed to be determined. Furthermore, tissues derived from iPS cells will be a nearly identical match to the cell donor, which is an important factor in research of disease modeling and drug screening. It is expected that iPS cells will help researchers learn how to reprogram cells to repair damaged tissues in the human body.

The purpose of this paper is to summarize the recent progresses in iPS cell development and iPS cell-based therapy, and describe patient specific iPS cells as a disease model, analyze the problems and considerations of iPS therapy in the clinical treatment of disease.

The methods of reprogramming somatic cells into iPS cells are summarized in Table . It was first demonstrated that genomic integration and high expression of four factors, Oct4/Sox2/Klf4/c-Myc or Oct4/Sox2/Nanog/LIN28 by virus, can reprogram fibroblast cells into iPS cells [1-3]. Later, it was shown that iPS cells can be generated from fibroblasts by viral integration of Oct4/Sox2/Klf4 without c-Myc [4]. Although these iPS cells showed reduced tumorigenicity in chimeras and progeny mice, the reprogramming process is much slower, and efficiency is substantially reduced. These studies suggest that the ectopic expression of these three transcription factors (Oct4/Klf4/Sox2) is required for reprogramming of somatic cells in iPS cells.

Various growth factors and chemical compounds have recently been found to improve the induction efficiency of iPS cells. Shi et al., [5] demonstrated that small molecules, able to compensate for Sox2, could successfully reprogram mouse embryonic fibroblasts (MEF) into iPS cells. They combined Oct4/Klf4 transduction with BIX-01294 and BayK8644s and derived MEF into iPS cells. Huangfu et al., [6, 7] reported that 5-azacytidine, DNA methyltransferase inhibitor, and valproic acid, a histone deacetylase inhibitor, improved reprogramming of MEF by more than 100 folds. Valproic acid enables efficient reprogramming of primary human fibroblasts with only Oct4 and Sox2.

Kim et al. showed that mouse neural stem cells, expressing high endogenous levels of Sox2, can be reprogrammed into iPS cells by transduction Oct4 together with either Klf4 or c-Myc [19]. This suggests that endogenous expression of transcription factors, that maintaining stemness, have a role in the reprogramming process of pluripotency. More recently, Tsai et al., [20] demonstrated that mouse iPS cells could be generated from the skin hair follicle papilla (DP) cell with Oct4 alone since the skin hair follicle papilla cells expressed endogenously three of the four reprogramming factors: Sox2, c-Myc, and Klf4. They showed that reprogramming could be achieved after 3 weeks with efficiency similar to other cell types reprogrammed with four factors, comparable to ES cells.

Retroviruses are being extensively used to reprogram somatic cells into iPS cells. They are effective for integrating exogenous genes into the genome of somatic cells to produce both mouse and human iPS cells. However, retroviral vectors may have significant risks that could limit their use in patients. Permanent genetic alterations, due to multiple retroviral insertions, may cause retrovirus-mediated gene therapy as seen in treatment of severe combined immunodeficiency [25]. Second, although retroviral vectors are silenced during reprogramming [26], this silencing may not be permanent, and reactivation of transgenes may occur upon the differentiation of iPS cells. Third, expression of exogenous reprogramming factors could occur. This may trigger the expression of oncogenes that stimulate cancer growth and alter the properties of the cells. Fourth, the c-Myc over-expression may cause tumor development after transplantation of iPS derived cells. Okita et al. [10] reported that the chimeras and progeny derived from iPS cells frequently showed tumor formation. They found that the retroviral expression of c-Myc was reactivated in these tumors. Therefore, it would be desirable to produce iPS cells with minimal, or free of, genomic integration. Several new strategies have been recently developed to address this issue (Table ).

Stadtfeld et al. [16] used an adenoviral vector to transduce mouse fibroblasts and hepatocytes, and generated mouse iPS cells at an efficiency of about 0.0005%. Fusaki et al. [22] used Sendai virus to efficiently generate iPS cells from human skin fibroblasts without genome integration. Okita et al. [27] repeatedly transfected MEF with two plasmids, one carrying the complementary DNAs (cDNAs) of Oct3/4, Sox2, and Klf4 and the other carrying the c-Myc cDNA. This generated iPS cells without evidence of plasmid integration. Using a polycistronic plasmid co-expressing Oct4, Sox2, Klf4, and c-Myc, Gonzalez et al., [28] reprogrammed MEF into iPS cells without genomic integration. Yu et al. [29] demonstrated that oriP/EBNA1 (EpsteinBarr nuclear antigen-1)-based episomal vectors could be used to generate human iPS cells free of exogenous gene integration. The reprogramming efficiency was about 36 colonies/1 million somatic cells. Narsinh et al., [21] derived human iPS cells via transfection of human adipocyte stromal cells with a nonviral minicircle DNA by repeated transfection. This produced hiPS cells colonies from an adipose tissue sample in about 4 weeks.

When iPS cells generated from either plasmid transfection or episomes were carefully analyzed to identify random vector integration, it was possible to have vector fragments integrated somewhere. Thus, reprogramming strategies entirely free of DNA-based vectors are being sought. In April 2009, it was shown that iPS cells could be generated using recombinant cell-penetrating reprogramming proteins [30]. Zhou et al. [30] purified Oct4, Sox2, Klf4 and c-Myc proteins, and incorporated poly-arginine peptide tags. It allows the penetration of the recombinant reprogramming proteins through the plasma membrane of MEF. Three iPS cell clones were successfully generated from 5x 104 MEFs after four rounds of protein supplementation and subsequent culture of 2328 days in the presence of valproic acid.

A similar approach has also been demonstrated to be able to generate human iPS cells from neonatal fibroblasts [31]. Kim et al. over-expressed reprogramming factor proteins in HEK293 cells. Whole cell proteins of the transduced HEK293 were extracted and used to culture fibroblast six times within the first week. After eight weeks, five cell lines had been established at a yield of 0.001%, which is one-tenth of viral reprogramming efficiency. Strikingly, Warren et al., [24] demonstrated that human iPS cells can be derived using synthetic mRNA expressing Oct3/4, Klf4, Sox2 and c-Myc. This method efficiently reprogrammed fibroblast into iPS cells without genome integration.

Strenuous efforts are being made to improve the reprogramming efficiency and to establish iPS cells with either substantially fewer or no genetic alterations. Besides reprogramming vectors and factors, the reprogramming efficiency is also affected by the origin of iPS cells.

A number of somatic cells have been successfully reprogrammed into iPS cells (Table ). Besides mouse and human somatic cells, iPS cells from other species have been successfully generated (Table ).

The origin of iPS cells has an impact on choice of reprogramming factors, reprogramming and differentiation efficiencies. The endogenous expression of transcription factors may facilitate the reprogramming procedure [19]. Mouse neural stem cells express higher endogenous levels of Sox2 and c-Myc than ES cells. Thus, two transcription factors, exogenous Oct4 together with either Klf4 or c-Myc, are sufficient to generate iPS cells from neural stem cells [19]. Ahmed et al. [14] demonstrated that mouse skeletal myoblasts endogenously expressed Sox2, Klf4, and c-Myc and can be easily reprogrammed to iPS cells.

It is possible that iPS cells may demonstrate memory of parental source and therefore have low differentiation efficiency into other tissue cells. Kim et al. [32] showed that iPS cells reprogrammed from peripheral blood cells could efficiently differentiate into the hematopoietic lineage cells. It was found, however, that these cells showed very low differentiation efficiency into neural cells. Similarly, Bar-Nur et al. found that human cell-derived iPS cells have the epigenetic memory and may differentiate more readily into insulin producing cells [33]. iPS cells from different origins show similar gene expression patterns in the undifferentiated state. Therefore, the memory could be epigenetic and are not directly related to the pluripotent status.

The cell source of iPS cells can also affect the safety of the established iPS cells. Miura et al. [54] compared the safety of neural differentiation of mouse iPS cells derived from various tissues including MEFs, tail-tip fibroblasts, hepatocyte and stomach. Tumorigenicity was examined. iPS cells that reprogrammed from tail-tip fibroblasts showed many undifferentiated pluripotent cells after three weeks of in vitro differentiation into the neural sphere. These cells developed teratoma after transplantation into an immune-deficient mouse brain. The possible mechanism of this phenomenon may be attributable to epigenetic memory and/or genomic stability. Pre-evaluated, non-tumorigenic and safe mouse iPS cells have been reported by Tsuji et al. [55]. Safe iPS cells were transplanted into non-obese diabetic/severe combined immunodeficiency mouse brain, and found to produce electrophysiologically functional neurons, astrocytes, and oligodendrocytes in vitro.

The cell source of iPS cells is important for patients as well. It is important to carefully evaluate clinically available sources. Human iPS cells have been successfully generated from adipocyte derived stem cells [35], amniocytes [36], peripheral blood [38], cord blood [39], dental pulp cells [40], oral mucosa [41], and skin fibroblasts (Table ). The properties and safety of these iPS cells should be carefully examined before they can be used for treatment.

Shimada et al. [17] demonstrated that combination of chemical inhibitors including A83-01, CHIR99021, PD0325901, sodium butyrate, and Y-27632 under conditions of physiological hypoxia human iPS cells can be rapidly generated from adipocyte stem cells via retroviral transduction of Oct4, Sox2, Klf4, and L-Myc. Miyoshi et al., [42] generated human iPS cells from cells isolated from oral mucosa via the retroviral gene transfer of Oct4, Sox2, c-Myc, and Klf4. Reprogrammed cells showed ES-like morphology and expressed undifferentiated markers. Yan et al., [40] demonstrated that dental tissue-derived mesenchymal-like stem cells can easily be reprogrammed into iPS cells at relatively higher rates as compared to human fibroblasts. Human peripheral blood cells have also been successfully reprogrammed into iPS cells [38]. Anchan et al. [36] described a system that can efficiently derive iPS cells from human amniocytes, while maintaining the pluripotency of these iPS cells on mitotically inactivated feeder layers prepared from the same amniocytes. Both cellular components of this system are autologous to a single donor. Takenaka et al. [39] derived human iPS cells from cord blood. They demonstrated that repression of p53 expression increased the reprogramming efficiency by 100-fold.

All of the human iPS cells described here are indistinguishable from human ES cells with respect to morphology, expression of cell surface antigens and pluripotency-associated transcription factors, DNA methylation status at pluripotent cell-specific genes and the capacity to differentiate in vitro and in teratomas. The ability to reprogram cells from human somatic cells or blood will allow investigating the mechanisms of the specific human diseases.

The iPS cell technology provides an opportunity to generate cells with characteristics of ES cells, including pluripotency and potentially unlimited self-renewal. Studies have reported a directed differentiation of iPS cells into a variety of functional cell types in vitro, and cell therapy effects of implanted iPS cells have been demonstrated in several animal models of disease.

A few studies have demonstrated the regenerative potential of iPS cells for three cardiac cells: cardiomyocytes, endothelial cells, and smooth muscle cells in vitro and in vivo. Mauritz [56] and Zhang [57] independently demonstrated the ability of mouse and human iPS cells to differentiate into functional cardiomyocytes in vitro through embryonic body formation. Rufaihah [58], et al. derived endothelial cells from human iPS cells, and showed that transplantation of these endothelial cells resulted in increased capillary density in a mouse model of peripheral arterial disease. Nelson et al. [59] demonstrated for the first time the efficacy of iPS cells to treat acute myocardial infarction. They showed that iPS cells derived from MEF could restore post-ischemic contractile performance, ventricular wall thickness, and electrical stability while achieving in situ regeneration of cardiac, smooth muscle, and endothelial tissue. Ahmed et al. [14] demonstrated that beating cardiomyocyte-like cells can be differentiated from iPS cells in vitro. The beating cells expressed early and late cardiac-specific markers. In vivo studies showed extensive survival of iPS and iPS-derived cardiomyocytes in mouse hearts after transplantation in a mouse experimental model of acute myocardial infarction. The iPs derived cardiomyocyte transplantation attenuated infarct size and improved cardiac function without tumorgenesis, while tumors were observed in the direct iPS cell transplantation animals.

Strategies to enhance the purity of iPS derived cardiomyocytes and to exclude the presence of undifferentiated iPS are required. Implantation of pre-differentiation or guided differentiation of iPS would be a safer and more effective approach for transplantation. Selection of cardiomyocytes from iPS cells, based on signal-regulatory protein alpha (SIRPA) or combined with vascular cell adhesion protein-1 (VCAM-1), has been reported. Dubois et al. [60] first demonstrated that SIRPA was a marker specifically expressed on cardiomyocytes derived from human ES cells and human iPS cells. Cell sorting with an antibody against SIRPA could enrich cardiac precursors and cardiomyocytes up to 98% troponin T+ cells from human ESC or iPS cell differentiation cultures. Elliott et al. [61] adopted a cardiac-specific reporter gene system (NKX2-5eGFP/w) and identified that VCAM-1 and SIRPA were cell-surface markers of cardiac lineage during differentiation of human ES cells.

Regeneration of functional cells from human stem cells represents the most promising approach for treatment of type 1 diabetes mellitus (T1DM). This may also benefit the patients with type 2 diabetes mellitus (T2DM) who need exogenous insulin. At present, technology for reprogramming human somatic cell into iPS cells brings a remarkable breakthrough in the generation of insulin-producing cells.

Human ES cells can be directed to become fully developed cells and it is expected that iPS cells could also be similarly differentiated. Stem cell based approaches could also be used for modulation of the immune system in T1DM, or to address the problems of obesity and insulin resistance in T2DM.

Tateishi et al., [62] demonstrated that insulin-producing islet-like clusters (ILCs) can be generated from the human iPS cells under feeder-free conditions. The iPS cell derived ILCs not only contain C-peptide positive and glucagon-positive cells but also release C-peptide upon glucose stimulation. Similarly, Zhang et al., [63] reported a highly efficient approach to induce human ES and iPS cells to differentiate into mature insulin-producing cells in a chemical-defined culture system. These cells produce insulin/C-peptide in response to glucose stimuli in a manner comparable to that of adult human islets. Most of these cells co-expressed mature cell-specific markers such as NKX6-1 and PDX1, indicating a similar gene expression pattern to adult islet beta cells in vivo.

Alipo et al. [64] used mouse skin derived iPS cells for differentiation into -like cells that were similar to the endogenous insulin-secreting cells in mice. These -like cells were able to secrete insulin in response to glucose and to correct a hyperglycemic phenotype in mouse models of both T1DM and T2DM after iPS cell transplant. A long-term correction of hyperglycemia could be achieved as determined by hemoglobin A1c levels. These results are encouraging and suggest that induced pluripotency is a viable alternative to directing iPS cell differentiation into insulin secreting cells, which has great potential clinical applications in the treatment of T1DM and T2 DM.

Although significant progress has been made in differentiating pluripotent stem cells to -cells, several hurdles remain to be overcome. It is noted in several studies that the general efficiency of in vitro iPS cell differentiation into functional insulin-producing -like cells is low. Thus, it is highly essential to develop a safe, efficient, and easily scalable differentiation protocol before its clinical application. In addition, it is also important that insulin-producing b-like cells generated from the differentiation of iPS cells have an identical phenotype resembling that of adult human pancreatic cells in vivo.

Currently, the methodology of neural differentiation has been well established in human ES cells and shown that these methods can also be applied to iPS cells. Chambers et al. [65] demonstrated that the synergistic action of Noggin and SB431542 is sufficient to induce rapid and complete neural conversion of human ES and iPS cells under adherent culture conditions. Swistowsk et al. [66] used a completely defined (xenofree) system, that has efficiently differentiated human ES cells into dopaminergic neurons, to differentiate iPS cells. They showed that the process of differentiation into committed neural stem cells (NSCs) and subsequently into dopaminergic neurons was similar to human ES cells. Importantly, iPS cell derived dopaminergic neurons were functional as they survived and improved behavioral deficits in 6-hydroxydopamine-leasioned rats after transplantation. Lee et al. [67] provided detailed protocols for the step-wise differentiation of human iPS and human ES into neuroectodermal and neural crest cells using either the MS5 co-culture system or a defined culture system (Noggin with a small-molecule SB431542), NSB system. The average time required for generating purified human NSC precursors will be 25 weeks. The success of deriving neurons from human iPS cells provides a study model of normal development and impact of genetic disease during neural crest development.

Wernig et al., [68] showed that iPS cells can give rise to neuronal and glial cell types in culture. Upon transplantation into the fetal mouse brain, the cells differentiate into glia and neurons, including glutamatergic, GABAergic, and catecholaminergic subtypes. Furthermore, iPS cells were induced to differentiate into dopamine neurons of midbrain character and were able to improve behavior in a rat model of Parkinson's disease (PD) upon transplantation into the adult brain. This study highlights the therapeutic potential of directly reprogrammed fibroblasts for neural cell replacement in the animal model of Parkinsons disease.

Tsuji et al., [55] used pre-evaluated iPS cells derived for treatment of spinal cord injury. These cells differentiated into all three neural lineages, participated in remyelination and induced the axonal regrowth of host 5HT+ serotonergic fibers, promoting locomotor function recovery without forming teratomas or other tumors. This study suggests that iPS derived neural stem/progenitor cells may be a promising cell source for treatment of spinal cord injury.

Hargus et al., [69] demonstrated proof of principle of survival and functional effects of neurons derived from iPS cells reprogrammed from patients with PD. iPS cells from patients with Parkinsons disease were differentiated into dopaminergic neurons that could be transplanted without signs of neuro-degeneration into the adult rodent striatum. These cells survived and showed arborization, and mediated functional effects in an animal model of Parkinsons disease. This study suggests that disease specific iPS cells can be generated from patients with PD, which be used to study the PD development and in vitro drug screen for treatment of PD.

Reprogramming technology is being applied to derive patient specific iPS cell lines, which carry the identical genetic information as their patient donor cells. This is particularly interesting to understand the underlying disease mechanism and provide a cellular and molecular platform for developing novel treatment strategy.

Human iPS cells derived from somatic cells, containing the genotype responsible for the human disease, hold promise to develop novel patient-specific cell therapies and research models for inherited and acquired diseases. The differentiated cells from reprogrammed patient specific human iPS cells retain disease-related phenotypes to be an in vitro model of pathogenesis (Table ). This provides an innovative way to explore the molecular mechanisms of diseases.

Disease Modeling Using Human iPS Cells

Recent studies have reported the derivation and differentiation of disease-specific human iPS cells, including autosomal recessive disease (spinal muscular atrophy) [70], cardiac disease [71-75], blood disorders [13, 76], diabetes [77], neurodegenerative diseases (amyotrophic lateral sclerosis [78], Huntingtons disease [79]), and autonomic nervous system disorder (Familial Dysautonomia) [80]. Patient-specific cells make patient-specific disease modeling possible wherein the initiation and progression of this poorly understood disease can be studied.

Human iPS cells have been reprogrammed from spinal muscular atrophy, an autosomal recessive disease. Ebert et al., [70] generated iPS cells from skin fibroblast taken from a patient with spinal muscular atrophy. These cells expanded robustly in culture, maintained the disease genotype and generated motor neurons that showed selective deficits compared to those derived from the patients' unaffected relative. This is the first study to show that human iPS cells can be used to model the specific pathology seen in a genetically inherited disease. Thus, it represents a promising resource to study disease mechanisms, screen new drug compounds and develop new therapies.

Similarly, three other groups reported their findings on the use of iPS cells derived cardiomyocytes (iPSCMs) as disease models for LQTS type-2 (LQTS2). Itzhaki et al., [72] obtained dermal fibroblasts from a patient with LQTS2 harboring the KCNH2 gene mutation and showed that action potential duration was prolonged and repolarization velocity reduced in LQTS2 iPS-CMs compared with normal cardiomyocytes. They showed that Ikr was significantly reduced in iPS-CMs derived from LQTS2. They also tested the potential therapeutic effects of nifedipine and the KATP channel opener pinacidil (which augments the outward potassium current) and demonstrated that they shortened the action potential duration and abolished early after depolarization. Similarly, Lahti et al., [73] demonstrated a more pronounced inverse correlation between the beating rate and repolarization time of LQTS2 disease derived iPS-CMs compared with normal control cells. Prolonged action potential is present in LQT2-specific cardiomyocytes derived from a mutation. Matsa et al., [74] also successfully generated iPS-CMs from a patient with LQTS2 with a known KCNH2 mutation. iPS-CMs with LQTS2 displayed prolonged action potential durations on patch clamp analysis and prolonged corrected field potential durations on microelectrode array mapping. Furthermore, they demonstrated that the KATP channel opener nicorandil and PD-118057, a type 2 IKr channel enhancer attenuate channel closing.

LQTS3 has been recapitulated in mouse iPS cells [75]. Malan et al. [75] generated disease-specific iPS cells from a mouse model of a human LQTS3. Patch-clamp measurements of LQTS 3-specific cardiomyocytes showed the biophysical effects of the mutation on the Na+ current, withfaster recovery from inactivation and larger late currents than observed in normal control cells. Moreover, LQTS3-specific cardiomyocytes had prolonged action potential durations and early after depolarizations at low pacing rates, both of which are classic features of the LQTS3 mutation.

Human iPS cells have been used to recapitulate diseases of blood disorder. Ye et al. [13] demonstrated that human iPS cells derived from periphery blood CD34+ cells of patients with myeloproliferative disorders, have the JAK2-V617F mutation in blood cells. Though the derived iPS cells contained the mutation, they appeared normal in phenotypes, karyotype, and pluripotency. After hematopoietic differentiation, the iPS cell-derived hematopoietic progenitor (CD34+/CD45+) cells showed the increased erythropoiesis and expression of specific genes, recapitulating features of the primary CD34+ cells of the corresponding patient from whom the iPS cells were derived. This study highlights that iPS cells reprogrammed from somatic cells from patients with blood disease provide a prospective hematopoiesis model for investigating myeloproliferative disorders.

Raya et al., [76] reported that somatic cells from Fanconi anaemia patients can be reprogrammed to pluripotency after correction of the genetic defect. They demonstrated that corrected Fanconi-anaemia specific iPS cells can give rise to haematopoietic progenitors of the myeloid and erythroid lineages that are phenotypically normal. This study offers proof-of-concept that iPS cell technology can be used for the generation of disease-corrected, patient-specific cells with potential value for cell therapy applications.

Maehr et al., [77] demonstrated that human iPS cells can be generated from patients with T1DM by reprogramming their adult fibroblasts. These cells are pluripotent and differentiate into three lineage cells, including insulin-producing cells. These cells provide a platform to assess the interaction between cells and immunocytes in vitro, which mimic the pathological phenotype of T1DM. This will lead to better understanding of the mechanism of T1DM and developing effective cell replacement therapeutic strategy.

Lee et al., [80] reported the derivation of human iPS cells from patient with Familial Dysautonomia, an inherited disorder that affects the development and function of nerves throughout the body. They demonstrated that these iPS cells can differentiate into all three germ layers cells. However gene expression analysis demonstrated tissue-specific mis-splicing of IKBKAP in vitro, while neural crest precursors showed low levels of normal IKBKAP transcript. Transcriptome analysis and cell-based assays revealed marked defects in neurogenic differentiation and migration behavior. All these recaptured familial Dysautonomia pathogenesis, suggesting disease specificity of the with familial Dysautonomia human iPS cells. Furthermore, they validated candidate drugs in reversing and ameliorating neuronal differentiation and migration. This study illustrates the promise of disease specific iPS cells for gaining new insights into human disease pathogenesis and treatment.

Human iPS cells derived reprogrammed from patients with inherited neurodegenerative diseases, amyotrophic lateral sclerosis [78] and Huntingtons disease 79, have also been reported. Dimos et al., [78] showed that they generated iPS cells from a patient with a familial form of amyotrophic lateral sclerosis. These patient-specific iPS cells possess the properties of ES cells and were reprogrammed successfully to differentiate into motor neurons. Zhang et al., [79] derived iPS cells from fibroblasts of patient with Huntingtons disease. They demonstrated that striatal neurons and neuronal precursors derived from these iPS cells contained the same CAG repeat expansion as the mutation in the patient from whom the iPS cell line was established. This suggests that neuronal progenitor cells derived from Huntingtons disease cell model have endogenous CAG repeat expansion that is suitable for mechanistic studies and drug screenings.

Disease specific somatic cells derived from patient-specific human iPS cells will generate a wealth of information and data that can be used for genetically analyzing the disease. The genetic information from disease specific-iPS cells will allow early and more accurate prediction and diagnosis of disease and disease progression. Further, disease specific iPS cells can be used for drug screening, which in turn correct the genetic defects of disease specific iPS cells.

iPS cells appear to have the greatest promise without ethical and immunologic concerns incurred by the use of human ES cells. They are pluripotent and have high replicative capability. Furthermore, human iPS cells have the potential to generate all tissues of the human body and provide researchers with patient and disease specific cells, which can recapitulate the disease in vitro. However, much remains to be done to use these cells for clinical therapy. A better understanding of epigenetic alterations and transcriptional activity associated with the induction of pluripotency and following differentiation is required for efficient generation of therapeutic cells. Long-term safety data must be obtained to use human iPS cell based cell therapy for treatment of disease.

These works were supported by NIH grants HL95077, HL67828, and UO1-100407.

The authors confirm that this article content has no conflicts of interest.

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Abstract

Traumatic spinal cord injury (SCI) is a life changing neurological condition with substantial socioeconomic implications for patients and their care-givers. Recent advances in medical management of SCI has significantly improved diagnosis, stabilization, survival rate and well-being of SCI patients. However, there has been small progress on treatment options for improving the neurological outcomes of SCI patients. This incremental success mainly reflects the complexity of SCI pathophysiology and the diverse biochemical and physiological changes that occur in the injured spinal cord. Therefore, in the past few decades, considerable efforts have been made by SCI researchers to elucidate the pathophysiology of SCI and unravel the underlying cellular and molecular mechanisms of tissue degeneration and repair in the injured spinal cord. To this end, a number of preclinical animal and injury models have been developed to more closely recapitulate the primary and secondary injury processes of SCI. In this review, we will provide a comprehensive overview of the recent advances in our understanding of the pathophysiology of SCI. We will also discuss the neurological outcomes of human SCI and the available experimental model systems that have been employed to identify SCI mechanisms and develop therapeutic strategies for this condition.

Keywords: spinal cord injury, secondary injury mechanisms, clinical classifications and demography, animal models, glial and immune response, glial scar, chondroitin sulfate proteoglycans (CSPGs), cell death

Spinal cord injury (SCI) is a debilitating neurological condition with tremendous socioeconomic impact on affected individuals and the health care system. According to the National Spinal Cord Injury Statistical Center, there are 12,500 new cases of SCI each year in North America (1). Etiologically, more than 90% of SCI cases are traumatic and caused by incidences such as traffic accidents, violence, sports or falls (2). There is a reported male-to-female ratio of 2:1 for SCI, which happens more frequently in adults compared to children (2). Demographically, men are mostly affected during their early and late adulthood (3rd and 8th decades of life) (2), while women are at higher risk during their adolescence (1519 years) and 7th decade of their lives (2). The age distribution is bimodal, with a first peak involving young adults and a second peak involving adults over the age of 60 (3). Adults older than 60 years of age whom suffer SCI have considerably worse outcomes than younger patients, and their injuries usually result from falls and age-related bony changes (1).

The clinical outcomes of SCI depend on the severity and location of the lesion and may include partial or complete loss of sensory and/or motor function below the level of injury. Lower thoracic lesions can cause paraplegia while lesions at cervical level are associated with quadriplegia (4). SCI typically affects the cervical level of the spinal cord (50%) with the single most common level affected being C5 (1). Other injuries include the thoracic level (35%) and lumbar region (11%). With recent advancements in medical procedures and patient care, SCI patients often survive these traumatic injuries and live for decades after the initial injury (5). Reports on the clinical outcomes of patients who suffered SCI between 1955 and 2006 in Australia demonstrated that survival rates for those suffering from tetraplegia and paraplegia is 91.2 and 95.9%, respectively (5). The 40-year survival rate of these individuals was 47 and 62% for persons with tetraplegia and paraplegia, respectively (5). The life expectancy of SCI patients highly depends on the level of injury and preserved functions. For instance, patients with ASIA Impairment Scale (AIS) grade D who require a wheelchair for daily activities have an estimated 75% of a normal life expectancy, while patients who do not require wheelchair and catheterization can have a higher life expectancy up to 90% of a normal individual (6). Today, the estimated life-time cost of a SCI patient is $2.35 million per patient (1). Therefore, it is critical to unravel the cellular and molecular mechanisms of SCI and develop new effective treatments for this devastating condition. Over the past decades, a wealth of research has been conducted in preclinical and clinical SCI with the hope to find new therapeutic targets for traumatic SCI.

SCI commonly results from a sudden, traumatic impact on the spine that fractures or dislocates vertebrae. The initial mechanical forces delivered to the spinal cord at the time of injury is known as primary injury where displaced bone fragments, disc materials, and/or ligaments bruise or tear into the spinal cord tissue (79). Notably, most injuries do not completely sever the spinal cord (10). Four main characteristic mechanisms of primary injury have been identified that include: (1) Impact plus persistent compression; (2) Impact alone with transient compression; (3) Distraction; (4) Laceration/transection (8, 11). The most common form of primary injury is impact plus persistent compression, which typically occurs through burst fractures with bone fragments compressing the spinal cord or through fracture-dislocation injuries (8, 12, 13). Impact alone with transient compression is observed less frequently but most commonly in hyperextension injuries (8). Distraction injuries occur when two adjacent vertebrae are pulled apart causing the spinal column to stretch and tear in the axial plane (8, 12). Lastly, laceration and transection injuries can occur through missile injuries, severe dislocations, or sharp bone fragment dislocations and can vary greatly from minor injuries to complete transection (8). There are also distinct differences between the outcomes of SCI in military and civilian cases. Compared to civilian SCI, blast injury is the common cause of SCI in battlefield that usually involves multiple segments of the spinal cord (14). Blast SCI also results in higher severity scores and is associated with longer hospital stays (15). A study on American military personnel, who sustained SCI in a combat zone from 2001 to 2009, showed increased severity and poorer neurological recovery compared to civilian SCI (15). Moreover, lower lumbar burst fractures and lumbosacral dissociation happen more frequently in combat injuries (1). Regardless of the form of primary injury, these forces directly damage ascending and descending pathways in the spinal cord and disrupt blood vessels and cell membranes (11, 16) causing spinal shock, systemic hypotension, vasospasm, ischemia, ionic imbalance, and neurotransmitter accumulation (17). To date, the most effective clinical treatment to limit tissue damage following primary injury is the early surgical decompression (< 24 h post-injury) of the injured spinal cord (18, 19). Overall, the extent of the primary injury determines the severity and outcome of SCI (20, 21).

Functional classification of SCI has been developed to establish reproducible scoring systems by which the severity of SCI could be measured, compared, and correlated with the clinical outcomes (20). Generally, SCI can be classified as either complete or incomplete. In complete SCI, neurological assessments show no spared motor or sensory function below the level of injury (4). In the past decades, several scoring systems have been employed for clinical classification of neurological deficits following SCI. The first classification system, Frankel Grade, was developed by Frankel and colleagues in 1969 (22). They assessed the severity and prognosis of SCI using numerical sensory and motor scales (22). This was a 5-grade system in which Grade A was the most severe SCI with complete loss of sensory and motor function below the level of injury. Grade B represented complete motor loss with preserved sensory function and sacral sparing. Patients in Grade C and D had different degrees of motor function preservation and Grade E represented normal sensory and motor function. The Frankel Grade was widely utilized after its publication due to its ease of use. However, lack of clear distinction between Grades C and D and inaccurate categorization of motor improvements in patients over time, led to its replacement by other scoring systems (20).

Other classification methods followed Frankel's system. In 1987, Bracken et al. at Yale University School of Medicine classified motor and sensory functions separately in a 5 and 7-scale systems, respectively (23). However, this scoring system failed to account for sacral function (20). Moreover, integration of motor and sensory classifications was impossible in this system and it was abandoned due to complexity and impracticality in clinical settings (20). Several other scoring systems were developed in 1970' and 1980's by different groups such as Lucas and Ducker at the Maryland Institute for Emergency Medical Services in late 1970's (24), Klose and colleagues at the University of Miami Neuro-spinal Index (UMNI) in early 1980s (25) and Chehrazi and colleagues (Yale Scale) in 1981 (26). These scoring systems also became obsolete due to their disadvantage in evaluation of sacral functions, difficulty of use or discrepancies between their motor and sensory scoring sub-systems (20).

The ASIA scoring system is currently the most widely accepted and employed clinical scoring system for SCI. ASIA was developed in 1984 by the American Spinal Cord Injury Association and has been updated over time to improve its reliability (). In this system, sensory function is scored from 02 and motor function from 0 to 5 (20). The ASIA impairment score (AIS) ranges from complete loss of sensation and movement (AIS = A) to normal neurological function (AIS = E). The first step in ASIA system is to identify the neurological level of injury (NLI). In this assessment, except upper cervical vertebrae that closely overlay the underlying spinal cord segments, the anatomical relationship between the spinal cord segments and their corresponding vertebra is not reciprocally aligned along the adult spinal cord (20). At thoracic and lumbar levels, each vertebra overlays a spinal cord segment one or two levels below and as the result, a T11 vertebral burst fracture results in neurological deficit at and below L1 spinal cord segment. Hence, the neurological level of injury (NLI) is defined as the most caudal neurological level at which all sensory and motor functions are normal (20). Upon identifying the NLI, if the injury is complete (AIS = A), zone of partial preservation (ZPP) is determined (20). ZPP is defined as all the segments below the NLI that have some preserved sensory or motor function. A precise record of ZPP enables the examiners to distinguish spontaneous from treatment-induced functional recovery, thus, essential for evaluating the therapeutic efficacy of treatments (20). Complete loss of motor and preservation of some sensory functions below the neurological level of the injury is categorized as AIS B (20). If motor function is also partially spared below the level of the injury, AIS score can be C or D (20). The AIS is scored D when the majority of the muscle groups below the level of the injury exhibit strength level of 3 or higher (for more details see ). ASIA classification combines the assessments of motor, sensory and sacral functions, thus addressing the shortcomings of previous scoring systems (20). The validity and reproducibility of ASIA system combined with its accuracy in prediction of patients' outcome have made it the most accepted and reliable clinical scoring system utilized for neurological classification of SCI (20).

ASIA scoring for the neurological classification of the SCI. A sample scoring sheet used for ASIA scoring in clinical setting is provided (adopted from: http://asia-spinalinjury.org).

In clinical management of SCI, neurological outcomes are generally determined at 72 h after injury using ASIA scoring system (20, 27). This time-point has shown to provide a more precise assessment of neurological impairments after SCI (28). One important predictor of functional recovery is to determine whether the injury was incomplete or complete. As time passes, SCI patients experience some spontaneous recovery of motor and sensory functions. Most of the functional recovery occurs during the first 3 months and in most cases reaches a plateau by 9 months after injury (20). However, additional recovery may occur up to 1218 months post-injury (20). Long term outcomes of SCI are closely related to the level of the injury, the severity of the primary injury and progression of secondary injury, which will be discussed in this review.

Depending on the level of SCI, patients experience paraplegia or tetraplegia. Paraplegia is defined as the impairment of sensory or motor function in lower extremities (27, 28). Patients with incomplete paraplegia generally have a good prognosis in regaining locomotor ability (~76% of patients) within a year (27). Complete paraplegic patients, however, experience limited recovery of lower limb function if their NLI is above T9 (29). An NLI below T9 is associated with 38% chance of regaining some lower extremity function (29). In patients with complete paraplegia, the chance of recovery to an incomplete status is only 4% with only half of these patients regaining bladder and bowel control (29). Tetraplegia is defined as partial or total loss of sensory or motor function in all four limbs. Patients with incomplete tetraplegia will gain better recovery than complete tetra- and paraplegia (30). Unlike complete SCI, recovery from incomplete tetraplegia usually happens at multiple levels below the NLI (20). Patients generally reach a plateau of recovery within 912 months after injury (20). Regaining some motor function within the first month after the injury is associated with a better neurological outcome (20). Moreover, appearance of muscle flicker (a series of local involuntary muscle contractions) in the lower extremities is highly associated with recovery of function (31). Patients with complete tetraplegia, often (6690%) regain function at one level below the injury (28, 30). Importantly, initial muscle strength is an important predictor of functional recovery in these patients (20). Complete tetraplegic patients with cervical SCI can regain antigravity muscle function in 27% of the cases when their initial muscle strength is 0 on a 5-point scale (32). However, the rate of regaining antigravity muscle strength at one caudal level below the injury increases to 97% when the patients have initial muscle strength of 12 on a 5-point scale (33).

An association between sensory and motor recovery has been demonstrated in SCI where spontaneous sensory recovery usually follows the pattern of motor recovery (20, 34). Maintenance of pinprick sensation at the zone of partial preservation or in sacral segments has been shown as a reliable predictor of motor recovery (35). One proposed reason for this association is that pinprick fibers in lateral spinothalamic tract travel in proximity of motor fibers in the lateral corticospinal tract, and thus, preservation of sensory fibers can be an indicator of the integrity of motor fiber (20). Diagnosis of an incomplete injury is of great importance and failure to detect sensory preservation at sacral segments results in an inaccurate assessment of prognosis (20).

In the past few decades, various animal models have been developed to allow understanding the complex biomedical mechanisms of SCI and to develop therapeutic strategies for this condition. An ideal animal model should have several characteristics including its relevance to the pathophysiology of human SCI, reproducibility, availability, and its potential to generate various severities of injury (36).

Small rodents are the most frequently employed animals in SCI studies due to their availability, ease of use and cost-effectiveness compared to primates and larger non-primate models of SCI (36, 37). Among rodents, rats more closely mimic pathophysiological, electrophysiological, functional, and morphological features of non-primate and human SCI (38). In rat (39), cat (40), monkey (41), and human SCI (17), a cystic cavity forms in the center of the spinal cord, which is a surrounded by a rim of anatomically preserved white matter. A study by Metz and colleagues compared the functional and anatomical outcomes of rat contusive injuries and human chronic SCI (42). High resolution MRI assessments identified that SCI-induced neuroanatomical changes such as spinal cord atrophy and size of the lesion were significantly correlated with the electrophysiological and functional outcomes in both rat and human contusive injuries (42). Histological assessments in rats also showed a close correlation between the spared white matter and functional preservation following injury (42). These studies provide evidence that rat models of contusive SCI could serve as an adequate model to develop and evaluate the structural and functional benefits of therapeutic strategies for SCI (42).

Mice show different histopathology than human SCI in which the lesion site is filled with dense fibrous connective-like tissue (4346). Mouse SCI studies show the presence of fibroblast-like cells expressing fibronectin, collagen, CD11b, CD34, CD13, and CD45 within the lesion core of chronic SCI, while it is absent in the injured spinal cord of rats (47). Another key difference between rat and mice SCI is the time-point of inflammatory cell infiltration. While microglia/macrophage infiltration is relatively consistent between rat and mouse models of SCI (47), there is a temporal difference in infiltration of neutrophils and T cells between the two species (47, 48). In SCI rats, infiltration of neutrophils, the first responders, peaks at 6 h post injury, followed by a significant decline at 2448 h after SCI (48). Similarly, in mouse SCI, neutrophil infiltration occurs within 6 h following injury; however, their numbers continue to rise and do not peak until 314 days post injury (49). T cell infiltration also varies between rat and mouse SCI models (50). In rats, T cell infiltration occurs between 3 and 7 days post injury and declines by 50% in the following 2 weeks (47), whereas in mice, T cell infiltration is not detected until 14 days post injury and their number doubles between 2 and 6 weeks post injury (47). Regardless of their pathophysiological relevance, mice have been used extensively in SCI studies primarily due to the availability of transgenic and mutant mouse models that have allowed uncovering molecular and cellular mechanisms of SCI (38).

In recent years, there has been emerging interest in employment of non-human primates and other larger animals such as pig, dog and cat as intermediate pre-clinical models (5153) to allow more effective translation of promising treatments from rodent models to human clinical trials (50). Although rodents have served as invaluable models for studying SCI mechanisms and therapeutic development, larger mammals, in particular non-human primates, share a closer size, neuroanatomy, and physiology to humans. Importantly, their larger size provides a more relevant platform for drug development, bioengineering inventions, and electrophysiological and rehabilitation studies. Nonetheless, both small and large animal models of SCI have limitations in their ability to predict the outcome in human SCI. One important factor is high degree of variability in the nature of SCI incidence, severity and location of the injury in human SCI, while in laboratory animal models, these variabilities are less (36). Values acquired by clinical scoring systems such as ASIA or Frankel scoring systems lack the consistency of the data acquired from laboratory settings, which makes the translation of therapeutic interventions from experimental to clinical settings challenging (36). A significant effect from an experimental treatment in consistent laboratory settings may not be reproducible in clinical settings due to high variability and heterogeneity in human populations and their injuries (36). To date, several pharmacological and cellular preclinical discoveries have led to human clinical trials based on their efficacy in improving the outcomes of SCI in small animal models. However, the majority of these trials failed to reproduce the same efficacy in human SCI. Thus, in pre-clinical studies, animal models, and study designs should be carefully chosen to reflect the reality of clinical setting as closely as possible (36). Larger animals provide the opportunity to refine promising therapeutic strategies prior to testing in human SCI; however, their higher cost, need for specialized facilities and small subject (sample) size have limited their use in SCI research (50). Thus, rodents are currently the most commonly employed models for preclinical discoveries and therapeutic development, while the use of larger animals is normally pursued for late stage therapies that have shown efficacy and promise in small animal models. provides a summary of available SCI models.

Animal models are also classified based on the type of SCI. The following sections will provide an overview on the available SCI models that are developed based on injury mechanisms, their specifications and relevance to human SCI ().

A complete transection model of SCI is relatively easy to reproduce (51). However, this model is less relevant to human SCI as a complete transection of the spinal cord rarely happens (51). While they do not represent clinical reality of SCI, transection models are specifically suitable for studying axonal regeneration or developing biomaterial scaffolds to bridge the gap between proximal and distal stamps of the severed spinal cord (51). Due to complete disconnection from higher motor centers, this model is also suitable for studying the role of propriospinal motor and sensory circuits in recovery of locomotion following SCI (51, 80). Partial transection models including hemi-section, unilateral transection and dorsal column lesions are other variants of transection models (51). Partial transection models are valuable for investigation of nerve grafting, plasticity and where a comparison between injured and non-injured pathways is needed in the same animal (51). However, these models lead to a less severe injury and higher magnitude of spontaneous recovery rendering them less suitable for development and evaluation of new therapies (51).

Contusion is caused by a transient physical impact to the spinal cord and is clinically-relevant. There are currently three types of devices that can produce contusion injury in animal models: weight-drop apparatus, electromagnetic impactor, and a recently introduced air gun device (51). The impactor model was first introduced by Gruner at New York University (NYU) in 1992 (81). The original NYU impactor included a metal rod of specific weight (10 g) that could be dropped on the exposed spinal cord from a specific height to induce SCI (51). This model allowed induction of a defined severity of SCI by adjusting the height, which the rod fell on the spinal cord (81). Parameters such as time, velocity at impact and biomechanical response of the tissue can be recorded for analysis and verification (51). The NYU impactor was later renamed to Multicenter Animal Spinal Cord Injury Study (MASCIS) impactor, and conditions surrounding the study and use of the MASCIS impactor were standardized (51). Since its introduction, the MASCIS impactor has been updated twice. The most recent version, MACIS III, was introduced in 2012 and included both electromagnetic control and digital recording of the impact parameters (51). However, inability to control duration of impact and weight bounce, that could cause multiple impacts, have been known limitations of MASCIS impactors (51).

The Infinite Horizon (IH) impactor is another type of impactor that utilizes a stepping motor to generate force-controlled impact in contrast to free fall in the MASICS impactor (51). This feature allows for better control over the force of impact and prevents weight bounce as the computer-controlled metal impounder can be immediately retracted upon transmitting a desired force to the spinal cord (51). IH impactor can be set to different force levels to provide mild, moderate and severe SCI in rats (ex. 100, 150, and 200 kdyn) (51). A limitation with IH impactors is unreliability of their clamps in holding the spinal column firmly during the impact that can cause inconsistent parenchymal injury and neurological deficits (51).

Ohio State University (OSU) impactor is a computer controlled electromagnetic impactor that was originally invented in 1987 and refined in 1992 to improve reliability (58). As the OSU impactor is electromagnetically controlled, multiple strikes are avoided (51). Subsequently, a modified version of the OSU impactor was developed in 2000 for use in mice (43). However, the OSU impactor is limited by its inability to determine the precise initial contact point with the spinal cord due to displacement of CSF upon loading the device (51). To date, MASCIS, IH and OSU impactor devices have been employed extensively and successfully to induce SCI. These impactor devices are available for small and large animals such as mice, rats, marmosets, cats, and pigs (51, 82).

Compressive models of SCI have been also employed for several decades (61). While contusion injury is achieved by applying a force for a very brief period (milliseconds), the compression injury consists of an initial contusion for milliseconds followed by a prolonged compression through force application for a longer duration (seconds to minutes) (51). Thus, compression injury can be categorized as contusive-compressive models (51). Various models of compressive SCI are available.

Clip compression is the most commonly used compression model of SCI in rat and mice (51, 61, 62, 83). It was first introduced by Rivlin and Tator in 1978 (61). In this model, following laminectomy, a modified aneurism clip with a calibrated closing force is applied to the spinal cord for a specific duration of time (usually 1 min) to induce a contusive-compressive injury (51). The severity of injury can be calibrated and modified by adjusting the force of the clip and the duration of compression (51). For example, applying a 50 g clip for 1 min typically produces a severe SCI, while a 35 g clip creates a moderate to severe injury with the same duration (83). Aneurysm clips were originally designed for use in rat SCI, however, in recent years smaller and larger clips have been developed to accommodate its use in mice (62) and pig models (52). The clip compression model has several advantages compared to contusion models. This method is less expensive and easier to perform (51). Importantly, in contrast to the impactor injury that contusion is only applied dorsally to the spinal cord, the clip compression model provides contusion and compression simultaneously both dorsally and ventrally. Hence, clip compression model more closely mimics the most common form of human SCI, which is primarily caused by dislocation and burst compression fractures (83). Despite its advantages, clip compression model can create variabilities such as the velocity of closing and actual delivered force that cannot be measured precisely at the time of application (51).

Calibrated forceps compression has been also employed to induce SCI in rodents. This simple and inexpensive compressive model was first utilized in 1991 for induction of SCI in guinea pigs (64). In this method, a calibrated forceps with a spacer is used to compress the spinal cord bilaterally (51). This model lacks the initial impact and contusive injury, which is associated with most cases of human traumatic SCI. Accordingly, this model is not a clinically relevant model for reproducing human SCI pathology and therapeutic development (51).

Balloon Compression model has been also utilized extensively in primates and larger animals such as dogs and cats (8486). In this model, a catheter with an inflatable balloon is inserted in the epidural or subdural space. The inflation of the balloon with air or saline for a specific duration of time provides the force for induction of SCI (51). Generally, all compression models (clip, forceps, and balloon) have the same limitation as the velocity and amount of force are unmeasurable (51).

In conclusion, while existing animal models do not recapitulate all clinical aspects of human SCI, the compression and contusion models are considered to be the most relevant and commonly employed methods for understanding the secondary injury mechanisms and therapeutic development for SCI.

Secondary injury begins within minutes following the initial primary injury and continues for weeks or months causing progressive damage of spinal cord tissue surrounding the lesion site (7). The concept of secondary SCI was first introduced by Allen in 1911 (87). While studying SCI in dogs, he observed that removal of the post traumatic hematomyelia improved neurological outcome. He hypothesized that presence of some biochemical factors in the necrotic hemorrhagic lesion causes further damage to the spinal cord (87). The term of secondary injury is still being used in the field and is referred to a series of cellular, molecular and biochemical phenomena that continue to self-destruct spinal cord tissue and impede neurological recovery following SCI () (20).

Summary of secondary injury processes following traumatic spinal cord injury. Diagram shows the key pathophysiological events that occur after primary injury and lead to progressive tissue degeneration. Vascular disruption and ischemia occur immediately after primary injury that initiate glial activation, neuroinflammation, and oxidative stress. These acute changes results in cell death, axonal injury, matrix remodeling, and formation of a glial scar.

Secondary injury can be temporally divided into acute, sub-acute, and chronic phases. The acute phase begins immediately following SCI and includes vascular damage, ionic imbalance, neurotransmitter accumulation (excitotoxicity), free radical formation, calcium influx, lipid peroxidation, inflammation, edema, and necrotic cell death (7, 20, 88). As the injury progresses, the sub-acute phase of injury begins which involves apoptosis, demyelination of surviving axons, Wallerian degeneration, axonal dieback, matrix remodeling, and evolution of a glial scar around the injury site (). Further changes occur in the chronic phase of injury including the formation of a cystic cavity, progressive axonal die-back, and maturation of the glial scar (7, 8992). Here, we will review the key components of acute secondary injury that contribute to the pathophysiology of SCI (, ).

Pathophysiology of traumatic spinal cord injury. This schematic diagram illustrates the composition of normal and injured spinal cord. Of note, while these events are shown in one figure, some of the pathophysiological events may not temporally overlap and can occur at various phases of SCI, which are described here. Immediately after primary injury, activation of resident astrocytes and microglia and subsequent infiltration of blood-borne immune cells results in a robust neuroinflammatory response. This acute neuroinflammatory response plays a key role in orchestrating the secondary injury mechanisms in the sub-acute and chronic phases that lead to cell death and tissue degeneration, as well as formation of the glial scar, axonal degeneration and demyelination. During the acute phase, monocyte-derived macrophages occupy the epicenter of the injury to scavenge tissue debris. T and B lymphocytes also infiltrate the spinal cord during sub-acute phase and produce pro-inflammatory cytokines, chemokines, autoantibodies reactive oxygen and nitrogen species that contribute to tissue degeneration. On the other hand, M2-like macrophages and regulatory T and B cells produce growth factors and pro-regenerative cytokines such as IL-10 that foster tissue repair and wound healing. Loss of oligodendrocytes in acute and sub-acute stages of SCI leads to axonal demyelination followed by spontaneous remyelination in sub-acute and chronic phases. During the acute and sub-acute phases of SCI; astrocytes, OPCs and pericytes, which normally reside in the spinal cord parenchyma, proliferate and migrate to the site of injury and contribute to the formation of the glial scar. The glial scar and its associated matrix surround the injury epicenter and create a cellular and biochemical zone with both beneficial and detrimental roles in the repair process. Acutely, the astrocytic glial scar limits the spread of neuroinflammation from the lesion site to the healthy tissue. However, establishment of a mature longstanding glial scar and upregulation of matrix chondroitin sulfate proteoglycans (CSPGs) are shown to inhibit axonal regeneration/sprouting and cell differentiation in subacute and chronic phases.

Disruption of spinal cord vascular supply and hypo-perfusion is one of the early consequences of primary injury (93). Hypovolemia and hemodynamic shock in SCI patients due to excessive bleeding and neurogenic shock result in compromised spinal cord perfusion and ischemia (93). Larger vessels such as anterior spinal artery usually remain intact (94, 95), while rupture of smaller intramedullary vessels and capillaries that are susceptible to traumatic damage leads to extravasation of leukocytes and red blood cells (93). Increased tissue pressure in edematous injured spinal cord and hemorrhage-induced vasospasm in intact vessels further disrupts blood flow to the spinal cord (93, 95). In rat and monkey models of SCI, there is a progressive reduction in blood flow at the lesion epicenter within the first few hours after injury which remains low for up to 24 h (96). The gray matter is more prone to ischemic damage compared to the white matter as it has a 5-fold higher density of capillary beds and contains neurons with high metabolic demand (95, 97, 98). After injury, white matter blood flow typically returns to normal levels within 15 min post injury, whereas there are multiple hemorrhages in the gray matter and as a result, re-perfusion usually does not occur for the first 24 h (9, 99, 100). Vascular insult, hemorrhage and ischemia ultimately lead to cell death and tissue destruction through multiple mechanisms, including oxygen deprivation, loss of adenosine triphosphate (ATP), excitotoxicity, ionic imbalance, free radical formation, and necrotic cell death. Cellular necrosis and release of cytoplasmic content increase the extracellular level of glutamate causing glutamate excitotoxicity (93, 101). Moreover, re-establishment of blood flow in ischemic tissue leads to further damage through generating free radicals and eliciting an inflammatory response (93, 102) that will be discussed in this review.

Within few minutes after primary SCI, the combination of direct cellular damage and ischemia/hypoxia triggers a significant rise of extracellular glutamate, the main excitatory neurotransmitter in the CNS (7). Glutamate binds to ionotropic (NMDA, AMPA, and Kainate receptors) as well as metabotropic receptors resulting in calcium influx inside the cells (103105) (93). The effect of glutamate is not restricted to neurons as its receptors are vastly expressed on the surface of all glia and endothelial cells (103106). Astrocytes can also release excess glutamate extracellularly upon elevation of their intracellular Ca2+ levels. Reduced ability of activated astrocytes for glutamate re-uptake from the interstitial space due to lipid peroxidation results in further accumulation of glutamate in the SCI milieu (93). Using microdialysis, elevated levels of glutamate have been detected in the white matter in the acute stage of injury (107). Based on a study by Panter and colleagues, glutamate increase is detected during the first 2030 min post SCI and returns to the basal levels after 60 min (108).

Under normal condition, concentration of free Ca2+ can considerably vary in different parts of the cell (109). In the cytosol, Ca2+ ranges from 50100 nM while it approaches 0.51.0 mM in the lumen of endoplasmic reticulum (110112). A long-lasting abnormal increase in Ca2+ concentration in cytosol, mitochondria or endoplasmic reticulum has detrimental consequences for the cell (109113). Mitochondria play a central role in calcium dependent neuronal death (113). In neurons, during glutamate induced excitotoxicity, NMDA receptor over-activity leads to mitochondrial calcium overload, which can cause apoptotic or necrotic cell death (113). Shortly after SCI, Ca2+ enters mitochondria through the mitochondrial calcium uniporter (MCU) (114). While the amount of mitochondrial calcium is limited during the resting state of a neuron, they can store a high amount of Ca2+ following stimulation (113). Calcium overload also activates a host of protein kinases and phospholipases that results in calpain mediated protein degradation and oxidative damage due to mitochondrial failure (93). In the injured white matter, astrocytes, oligodendrocytes and myelin are also damaged by the increased release of glutamate and Ca2+-dependent excitotoxicity (115). Within the first few hours after injury, oligodendrocytes show signs of caspase-3 activation and other apoptotic features, and their density declines (116). Interestingly, while glutamate excitotoxicity is triggered by ionic imbalance in the white matter, in the gray matter, it is largely associated with the activity of neuronal NMDA receptors (117, 118). Altogether, activation of NMDA receptors and consequent Ca2+ overload appears to induce intrinsic apoptotic pathways in neurons and oligodendrocytes and causes cell death in the first week of SCI in the rat (119, 120). Administration of NMDA receptor antagonist (MK-801) shortly following SCI has been associated with improved functional recovery and reduced edema (121).

Mitochondrial calcium overload also impedes mitochondrial respiration and results in ATP depletion disabling Na+/K+ ATPase and increasing intracellular Na+ (119, 122124). This reverses the function of the Na+ dependent glutamate transporter that normally utilizes Na+ gradient to transfer glutamate into the cells (119, 125, 126). Moreover, the excess intracellular Na+ reverses the activity of Na+/Ca2+ exchanger allowing more Ca+ influx (127). Cellular depolarization activates voltage gated Na+ channels that results in entry of Cl and water into the cells along with Na+ causing swelling and edema (128). Increased Na+ concentration over-activates Na+/H+ exchanger causing a rise in intracellular H+ (101, 129). Resultant intracellular acidosis increases membrane permeability to Ca2+ that exacerbates the injury-induced ionic imbalance (101, 129). Axons are more susceptible to the damage caused by ionic imbalance due to their high concentration of voltage gated Na+ channels in the nodes of Ranvier (7). Accumulating evidence shows that administration of Na+ channel blockers such as Riluzole attenuates tissue damage and improves functional recovery in SCI underlining sodium as a key player in secondary injury mechanisms (130133).

SCI results in production of free radicals and nitric oxide (NO) (114). Mitochondrial Ca2+ overload activates NADPH oxidase (NOX) and induces generation of superoxide by electron transport chain (ETC) (114). Reactive oxygen and nitrogen species (ROS and RNS) produced by the activity of NOX and ETC activates cytosolic poly (ADP ribose) polymerase (PARP). PARP consumes and depletes NAD+ causing failure of glycolysis, ATP depletion and cell death (114). Moreover, PAR polymers produced by PARP activity, induce the release of apoptosis inducing factor (AIF) from mitochondria and induce cell death (114). On the other hand, acidosis caused by SCI results in the release of intracellular iron from ferritin and transferrin (93). Spontaneous oxidation of Fe2+ to Fe3+ gives rise to more superoxide radicals (93). Subsequently, the Fenton reaction between Fe3+ and hydrogen peroxide produces highly reactive hydroxyl radicals (134). The resultant ROS and RNS react with numerous targets including lipids in the cell membrane with the most deleterious effects (93, 135). Because free radicals are short-lived and difficult to assess, measurements of their activity and final products, such as Malondialdehyde (MDA), are more reliable following SCI. Current evidence indicates that MDA levels are elevated as early as 1 h and up to 1 week after SCI (136, 137).

Oxidation of lipids and proteins is one of the key mechanisms of secondary injury following SCI (93). Lipid peroxidation starts when ROSs interact with polyunsaturated fatty acids in the cell membrane and generate reactive lipids that will then form lipid peroxyl radicals upon interacting with free superoxide radicals (138, 139). Each lipid peroxyl radical can react with a neighboring fatty acid, turn it into an active lipid and start a chain reaction that continues until no more unsaturated lipids are available or terminates when the reactive lipid quenches with another radical (93). The final products of this termination step of the lipid peroxidation is 4-hydroxynonenal (HNE) and 2-propenal, which are highly toxic to the cells (138140). Lipid peroxidation is also an underlying cause of ionic imbalance through destabilizing cellular membranes such as cytoplasmic membrane and endoplasmic reticulum (93). Moreover, lipid peroxidation leads to Na+/K+ ATPase dysfunction that exacerbates the intracellular Na+ overload (141). In addition to ROS associated lipid peroxidation, amino acids are subject to significant RNS associated oxidative damage following SCI (93). RNSs (containing ONOO) can nitrate the tyrosine residues of amino acids to form 3-nitrotyrosine (3-NT), a marker for peroxynitrite (ONOO) mediated protein damage (139). Lipid and protein oxidation following SCI has a number of detrimental consequences at cellular level including mitochondrial respiratory and metabolic failure as well as DNA alteration that ultimately lead to cell death (141).

Cell death is a major event in the secondary injury mechanisms that affects neurons and glia after SCI (142145). Cell death can happen through various mechanisms in response to various injury-induced mediators. Necrosis and apoptosis were originally identified as two major cell death mechanisms following SCI (146148). However, recent research has uncovered additional forms of cell death. In 2012, the Nomenclature Committee on Cell Death (NCCD) NCCD defined 12 different forms of cell death such as necroptosis, pyroptosis, and netosis (149). Among the identified modes of cell death, to date, necrosis, necroptosis, apoptosis, and autophagy have been studied more extensively in the context of SCI and will be discussed in this review.

Following SCI, neurons and glial cells die through necrosis as the result of mechanical damage at the time of primary injury that also continues to the acute and subacute stages of injury (7, 150). Necrosis occurs due to a multitude of factors including accumulation of toxic blood components (151), glutamate excitotoxicity and ionic imbalance (152), ATP depletion (153), pro-inflammatory cytokine release by neutrophils and lymphocytes (154, 155), and free radical formation (142, 156158). It was originally thought that necrosis is caused by a severe impact on a cell that results in rapid cell swelling and lysis. However, follow up evidence showed that in the case of seizure, ischemia and hypoglycemia, necrotic neurons show signs of shrunken, pyknotic, and condensed nuclei, with swollen, irreversibly damaged mitochondria and plasma membrane that are surrounded by astrocytic processes (159). Moreover, necrosis was conventionally viewed as instantaneous energy-independent non-programmed cell death (142, 156). However, recent research has identified another form of necrosis, termed as necroptosis, that is executed by regulated mechanisms.

Programmed necrosis or necroptosis has been described more recently as a highly regulated, caspase-independent cell death with similar morphological characteristics as necrosis (160). Necroptosis is a receptor-mediated process. It is induced downstream of the TNF receptor 1 (TNFR1) and is dependent on the activity of the receptor interacting protein kinase 1 (RIPK1) and RIPK3. Recent studies has uncovered a key role for RIPK1 as the mediator of necroptosis and a regulator of the innate immune response involved in both inflammation and cell death (161). Evidence from SCI studies show that lysosomal damage can potentiate necroptosis by promoting RIPK1 and RIPK3 accumulation (161). Interestingly, inhibition of necroptosis by necrostatin-1, a RIPK1 inhibitor, improves functional outcomes after SCI (150). These initial findings suggest that modulation of necroptosis pathways seems to be a promising target for neuroprotective strategies after SCI.

Apoptosis is the most studied mechanism of cell death after SCI. Apoptosis represents a programmed, energy dependent mode of cell death that begins within hours of primary injury (7). This process takes place in cells that survive the primary injury but endure enough insult to activate their apoptotic pathways (142). In apoptosis, the cell shrinks and is eventually phagocytosed without induction of an inflammatory response (156). Apoptosis typically occurs in a delayed manner in areas more distant to the injury site and most abundantly affects oligodendrocytes. In rat SCI, apoptosis happens as early as 4 h after the injury and reaches a peak at 7 day (156). At the site of injury majority of oligodendrocytes are lost within 7 days after SCI (162). However, apoptosis can be observed at a diminished rate for weeks after SCI (162, 163). Microglia and astrocytes also undergo apoptosis (156, 164). Interestingly, apoptotic cell death occurs in the chronically injured spinal cord in rat, monkey and human models of SCI, which is thought to be due to loss of trophic support from degenerating axons (146, 165).

Apoptosis is induced through extrinsic and intrinsic pathways based on the triggering mechanism (166). The extrinsic pathway is triggered by activation of death receptors such as FAS and TNFR1, which eventually activates caspase 8 (167). The intrinsic pathway, however, is regulated through a balance between intracellular pro- and anti-apoptotic proteins and is triggered by the release of cytochrome C from mitochondria and activating caspase 9 (167). In SCI lesion, apoptosis primarily happens due to injury induced Ca2+ influx, which activates caspases and calpain; enzymes involved in breakdown of cellular proteins (7). Moreover, it is believed that the death of neurons and oligodendrocytes in remote areas from the lesion epicenter can be mediated through cytokines such as TNF-, free radical damage and excitotoxicity since calcium from damaged cells within the lesion barely reaches these remote areas (8, 168). Fas mediated cell death has been suggested as a key mechanism of apoptosis following SCI (144, 169172). Post-mortem studies on acute and chronic human SCI and animal models revealed that Fas mediated apoptosis plays a role in oligodendrocyte apoptosis and inflammatory response at acute and subacute stages of SCI (173). Fas deficient mice exhibit a significant reduction in apoptosis and inflammatory response evidenced by reduced macrophage infiltration and inflammatory cytokine expression following SCI (173). Interestingly, Fas deficient mice show a significantly improved functional recovery after SCI (173) suggesting the promise of anti-apoptotic strategies for SCI.

SCI also results in a dysregulated autophagy (174). Normally, autophagy plays an important role in maintaining the homeostasis of cells by aiding in the turnover of proteins and organelles. In autophagy, cells degrade harmful, defective or unnecessary cytoplasmic proteins and organelles through a lysosomal dependent mechanism (175, 176). The process of autophagy starts with the formation of an autophagosome around the proteins and organelles that are tagged for autophagy (176). Next, fusion of the phagosome with a lysosome form an autolysosome that begins a recycling process (176). In response to cell injury and endoplasmic reticulum (ER) stress, autophagy is activated and limits cellular loss (177, 178). Current evidence suggests a neuroprotective role for autophagy after SCI (175, 179). Dysregulation of autophagy contributes to neuronal loss (174, 180). Accumulation of autophagosomes in ventral horn motor neurons have been detected acutely following SCI (181). Neurons with dysregulated autophagy exhibit higher expression of caspase 12 and become more prone to apoptosis (174). Moreover, blocking autophagy has been associated with neurodegenerative diseases such as Parkinson's and Alzheimer's disease (182184). Autophagy promotes cell survival through elimination of toxic proteins and damaged mitochondria (185, 186). Interestingly, autophagy is crucial in cytoskeletal remodeling and stabilizes neuronal microtubules by degrading SCG10, a protein involved in microtubule disassembly (179). Pharmacological induction of autophagy in a hemi-section model of SCI in mice has been associated with improved neurite outgrowth and axon regeneration, following SCI (179). Altogether, although further studies are needed, autophagy is currently viewed as a beneficial mechanism in SCI.

Neuroinflammation is a key component of the secondary injury mechanisms with local and systemic consequences. Inflammation was originally thought to be detrimental for the outcome of SCI (187). However, now it is well-recognized that inflammation can be both beneficial and detrimental following SCI, depending on the time point and activation state of immune cells (188). There are multiple cell types involved in the inflammatory response following injury including neutrophils, resident microglia, and astrocytes, dendritic cells (DCs), blood-born macrophages, B- and T-lymphocytes (189) (). The first phase of inflammation (02 days post injury) involves the recruitment of resident microglia and astrocytes and blood-born neutrophils to the injury site (190). The second phase of inflammation begins approximately 3 days post injury and involves the recruitment of blood-born macrophages, B- and T-lymphocytes to the injury site (189, 191193). T lymphocytes become activated in response to antigen presentation by macrophages, microglia and other antigen presenting cells (APCs) (194). CD4+ helper T cells produce cytokines that stimulate B cell antibody production and activate phagocytes (195) (). In SCI, B cells produce autoantibodies against injured spinal cord tissue, which exacerbate neuroinflammation and cause tissue destruction (196). While inflammation is more pronounced in the acute phase of injury, it continues in subacute and chronic phase and may persist for the remainder of a patients' life (193). Interestingly, composition and phenotype of inflammatory cells change based on the injury phase and the signals present in the injury microenvironment. It is established that microglia/macrophages, T cells, B cells are capable of adopting a pro-inflammatory or an anti-inflammatory pro-regenerative phenotype in the injured spinal cord (191, 197199). The role of each immune cell population in the pathophysiology of SCI will be discussed in detail in upcoming sections.

Immune response in spinal cord injury. Under normal circumstances, there is a balance between pro-inflammatory effects of CD4+ effector T cells (Teff) and anti-inflammatory effects of regulatory T and B cells (Treg and Breg). Treg and Breg suppress the activation of antigen specific CD4+ Teff cells through production of IL-10 and TGF-. Injury disrupts this balance and promote a pro-inflammatory environment. Activated microglia/macrophages release pro-inflammatory cytokines and chemokines and present antigens to CD4+ T cells causing activation of antigen specific effector T cells. Teff cells stimulate antigen specific B cells to undergo clonal expansion and produce autoantibodies against spinal cord tissue antigens. These autoantibodies cause neurodegeneration through FcR mediated phagocytosis or complement mediated cytotoxicity. M1 macrophages/microglia release pro-inflammatory cytokines and reactive oxygen species (ROS) that are detrimental to neurons and oligodendrocytes. Breg cells possess the ability to promote Treg development and restrict Teff cell differentiation. Breg cells could also induce apoptosis in Teff cells through Fas mediate mechanisms.

Astrocytes are not considered an immune cell per se; however, they play pivotal roles in the neuroinflammatory processes in CNS injury and disease. Their histo-anatomical localization in the CNS has placed them in a strategic position for participating in physiological and pathophysiological processes in the CNS (200). In normal CNS, astrocytes play major roles in maintaining CNS homeostasis. They contribute to the structure and function of blood-brain-barrier (BBB), provide nutrients and growth factors to neurons (200), and remove excess fluid, ions, and neurotransmitters such as glutamate from synaptic spaces and extracellular microenvironment (200). Astrocytes also play key roles in the pathologic CNS by regulating BBB permeability and reconstruction as well as immune cell activity and trafficking (201). Astrocytes contribute to both innate and adaptive immune responses following SCI by differential activation of their intracellular signaling pathways in response to environmental signals (201).

Astrocytes react acutely to CNS injury by increasing cytokine and chemokine production (202). They mediate chemokine production and recruitment of neutrophils through an IL-1R1-Myd88 pathway (202). Activation of the nuclear factor kappa b (NF-B) pathway, one of the key downstream targets of interleukin (IL)1R-Myd88 axis, increases expression of intracellular adhesion molecule (ICAM) and vascular cell adhesion molecule (VCAM), which are necessary for adhesion and extravasation of leukocytes in inflammatory conditions such as SCI (201, 202). Within minutes of injury, production of IL-1 is significantly elevated in astrocytes and microglia (203). Moreover, chemokines such as monocyte chemoattractant protein (MCP)-1, chemokine C-C motif ligand 2 (CCL2), C-X-C motif ligand 1 (CXCL1), and CXCL2 are produced by astrocytes, and enhance the recruitment of neutrophils and pro-inflammatory macrophages following injury (201, 202). Astrocytes also promote pro-inflammatory M1-like phenotype in microglia/macrophages in the injured spinal cord through their production of TNF-, IL-12, and IFN- (204206). Interestingly, astrocytes also produce anti-inflammatory cytokines, such as TGF- and IL-10, which can promote a pro-regenerative M2-like phenotype in microglia/macrophages (201, 207, 208).

Immunomodulatory role of astrocytes is defined by activity of various signaling pathways through a wide variety of surface receptors (200). For example, gp130, a member of IL-6 cytokine family, activates SHP2/Ras/Erk signaling cascade in astrocytes and limits neuroinflammation in autoimmune rodent models (209). TGF- signaling in astrocytes has been implicated in modulation of neuroinflammation through inhibition of NF-B activity and nuclear translocation (201, 210). STAT3 is another key signaling pathway in astrocytes with beneficial properties in neuroinflammation. Increase in STAT3 phosphorylation enhances astrocytic scar formation and restricts the expansion of inflammatory cells in mouse SCI, which is associated with improved functional recovery (211). Detrimental signaling pathways in astrocytes are known to be activated by cytokines, sphingolipids and neurotrophins (200). As an example, IL-17 is a key pro-inflammatory cytokine produced by effector T cells that can bind to IL-17R on the astrocyte surface (200). Activation of IL-17R results in the activation of NF-B, which enhances expression of pro-inflammatory mediators, activation of oxidative pathways and exacerbation of neuroinflammation (200, 212). This evidence shows the significance of astrocytes in the inflammatory processes following SCI and other neuroinflammatory diseases of the CNS.

Neutrophils infiltrate the spinal cord from the bloodstream within the first few hours after injury (213). Their population increases acutely in the injured spinal cord tissue and reaches a peak within 24 h post-injury (214). The presence of neutrophils is mostly limited to the acute phase of SCI as they are rarely found sub-acutely in the injured spinal cord (214). The role of neutrophils in SCI pathophysiology is controversial. Evidence shows that neutrophils contribute to phagocytosis and clearance of tissue debris (48). They release inflammatory cytokines, proteases and free radicals that degrade ECM, activate astrocytes and microglia and initiate neuroinflammation (48). Although neutrophils have been conventionally associated with tissue damage (48, 215), their elimination compromises the healing process and impedes functional recovery (216).

To elucidate the role of neutrophils in SCI, Stirling and colleagues used a specific antibody to reduce circulating LyG6/Gr1+ neutrophils in a mouse model of thoracic contusive SCI (216). This approach significantly reduced neutrophil infiltration in the injured spinal cord by 90% at 24 and 48 h after SCI (216). Surprisingly, neutrophil depletion aggravated the neurological and structural outcomes in the injured animals suggesting a beneficial role for neutrophils in the acute phase of injury (216). It is shown that simulated neutrophils release IL-1 receptor antagonist that can exert neuroprotective effects following SCI (217). Moreover, ablation of neutrophils results in altered expression of cytokines and chemokines and downregulation of growth factors such as fibroblast growth factors (FGFs), vascular endothelial growth factors (VEGFs) and bone morphogenetic proteins (BMPs) in the injured spinal cord that seemingly disrupt the normal healing process (216). Altogether, neutrophils play important roles in regulating neuroinflammation at the early stage of SCI that shapes the immune response and repair processes at later stages. While neutrophils were originally viewed as being detrimental in SCI, emerging evidence shows their critical role in the repair process. Further investigations are required to elucidate the role of neutrophils in SCI pathophysiology.

Following neutrophil invasion, microglia/macrophages populate the injured spinal cord within 23 days post-SCI. Macrophage population is derived from invading blood-borne monocytes or originate from the CNS resident macrophages that reside in the perivascular regions within meninges and subarachnoid space (218, 219). The population of microglia/macrophages reaches its peak at 710 days post-injury in mouse SCI, followed by a decline in the subacute and chronic phases (20, 220). While macrophages and microglia share many functions and immunological markers, they have different origins. Microglia are resident immune cells of the CNS that originate from yolk sac during the embryonic period (221). Macrophages are derived from blood monocytes, which originate from myeloid progeny in the bone marrow (222, 223). Upon injury, acute disruption of brain-spinal cord barrier (BSB) enables monocytes, to infiltrate the spinal cord tissue and transform into macrophages (222). Macrophages populate the injury epicenter, while resident microglia are mainly located in the perilesional area (222). Once activated, macrophages, and microglia are morphologically and immunohistologically indistinguishable (224). Macrophages and microglia play a beneficial role in CNS regeneration. They promote the repair process by expression of growth promoting factors such as nerve growth factor (NGF), neurotrophin-3 (NT-3) and thrombospondin (225, 226). Macrophages and microglia are important for wound healing process following SCI due to their ability for phagocytosis and scavenging damaged cells and myelin debris following SCI (222, 227).

Based on microenvironmental signals, macrophages/microglia can be polarized to either pro-inflammatory (M1-like) or anti-inflammatory pro-regenerative (M2-like) phenotype, and accordingly contribute to injury or repair processes following SCI (191, 224, 228230). Whether both microglia and macrophages possess the ability to polarize or it is mainly the property of monocyte derived macrophages is still a matter of debate and needs further elucidation (231233). Some evidence show that Proinflammatory M1-like microglia/macrophages can be induced by exposure to Th1 specific cytokine, interferon (IFN)- (224, 230). Moreover, the SCI microenvironment appears to drive M1 polarization of activated macrophages (231). SCI studies have revealed that increased level of the proinflammatory cytokine, TNF-, and intracellular accumulation of iron drives an M1-like proinflammatory phenotype in macrophages after injury (231). Importantly, following SCI, activated M1-like microglia/macrophages highly express MHCII and present antigens to T cells and contribute to the activation and regulation of innate and adaptive immune response () (224, 228). Studies on acute and subacute SCI and experimental autoimmune encephalomyelitis (EAE) models have shown that M1-like macrophages are associated with higher expression of chondroitin sulfate proteoglycans (CSPGs) and increased EAE severity and tissue damage (234237). In vitro, addition of activated M1-like macrophages to dorsal root ganglion (DRG) neuron cultures leads to axonal retraction and failure of regeneration as the expression of CSPGs is much higher in M1-like compared to M2-like macrophages (237, 238). M1-like macrophages also produce other repulsive factors such as repulsive guidance molecule A (RGMA) that is shown to induce axonal retraction following SCI (239, 240). Interestingly, recent evidence shows that IFN- and TNF polarized M1 microglia show reduced capacity for phagocytosis (241), a process that is critical for tissue repair after SCI.

Pro-regenerative M2-like microglia/macrophages, are polarized by Th2 cytokines, IL-4 and IL-13 and exhibit a high level of IL-10, TGF-, and arginase-1 with reduced NF-B pathway activity (224). IL-10 is a potent immunoregulatory cytokine with positive roles in repair and regeneration following CNS injury (242244). IL-10 knock-out mice show higher production of pro-inflammatory and oxidative stress mediators after SCI (245). Lack of IL-10 is also correlated with upregulated levels of pro-apoptotic factors such as Bax and reduced expression of anti-apoptotic factors such as Bcl-2 (245). SCI mice that lacked IL-10 exhibited poorer recovery of function compared to wild-type mice (245). Our recent studies show that IL-10 polarized M2 microglia show enhanced capacity for phagocytosis (241). We have also found that M2 polarized microglia enhance the ability of neural precursor cells for oligodendrocyte differentiation through IL-10 mediated mechanisms (241). In addition to immune modulation, M2-like microglia/macrophages promote axonal regeneration (224). However, similar to the detrimental effects of prolonged M1 macrophage response, excessive M2-like activity promotes fibrotic scar formation through the release of factors such as TGF-, PDGF, VEGF, IGF-1, and Galectin-3 (224, 246248). Hence, a balance between proinflammatory M1 and pro-regenerative M2 macrophage/microglia response is beneficial for the repair of SCI (249).

T and B lymphocytes play pivotal role in the adaptive immune response after SCI (194). Lymphocytes infiltrate the injured spinal cord acutely during the first week of injury and remain chronically in mouse and rat SCI (47, 193, 194, 196). In contrast to the innate immune response that can be activated directly by foreign antigens, the adaptive immune response requires a complex signaling process in T cells elicited by antigen presenting cells (250). Similar to other immune cells, T and B lymphocytes adopt different phenotypes and contribute to both injury and repair processes in response to microenvironmental signals (194, 251). SCI elicits a CNS-specific autoimmune response in T and B cells, which remains active chronically (196). Autoreactive T cells can exert direct toxic effects on neurons and glial cells (194, 252). Moreover, T cells can indirectly affect neural cell function and survival through pro-inflammatory cytokine and chemokine production (e.g. IL-1, TNF-, IL-12, CCL2, CCL5, and CXCL10) (194, 252). Genetic elimination of T cells (in athymic nude rats) or pharmacological inhibition of T cells (using cyclosporine A and tacrolimus) leads to improved tissue preservation and functional recovery after SCI (194, 253) signifying the impact of T cells in SCI pathophysiology and repair.

Under normal circumstances, systemic autoreactive effector CD4+ helper T cells (Teff) are suppressed by CD4+FoxP3+ regulatory T cells (Treg) () (194, 254). This inhibition is regulated through various mechanisms such as release of anti-inflammatory cytokines IL-10 and TGF- by the Treg cells () (194). Moreover, it is known that Treg mediated inhibition of antigen presentation by dendritic cells (DCs) prevent Teff cell activation (194). Following SCI, this Treg -Teff regulation is disrupted. Increased activity of autoreactive Teff cells contributes to tissue damage through production of pro-inflammatory cytokines and chemokines, promoting M1-like macrophage phenotype and induction of Fas mediated neuronal and oligodendroglial apoptosis () (173). Moreover, autoreactive Teff cells promote activation and differentiation of antigen specific B cells to autoantibody producing plasma cells that contribute to tissue damage after SCI (255). In SCI and MS patients, myelin specific proteins such as myelin basic protein (MBP) significantly increase the population of circulating T cells (256, 257). Moreover, serological assessment of SCI patients has shown high levels of CNS reactive IgM and IgG isotypes confirming SCI-induced autoimmune activity of T and B cells () (196, 258, 259). In animal models of SCI, serum IgM level increases acutely followed by an elevation in the levels of IgG1 and IgG2a at later time-points (196). In addition to autoantibody production, autoreactive B cells contribute to CNS injury through pro-inflammatory cytokines that stimulate and maintain the activation states of Teff cells (194, 260). B cell knockout mice (BCKO) that have no mature B cell but with normal T cells, show a reduction in lesion volume, lower antibody levels in the cerebrospinal fluid and improved recovery of function following SCI compared to wild-type counterparts (255). Of note, antibody mediated injury is regulated through complement activation as well as macrophages/microglia that express immunoglobulin receptors (193, 255).

The effect of SCI on systemic B cell response is controversial. Evidence shows that SCI can suppress B cell activation and antibody production (261). Studies in murine SCI have shown that B cell function seems to be influenced by the level of injury (262). While injury to upper thoracic spinal cord (T3) suppresses the antibody production, a mid-thoracic (T9) injury has no effect on B cell antibody production (262). An increase in the level of corticosterone in serum together with elevation of splenic norepinephrine found to be responsible for the suppression of B cell function acutely following SCI (261). Elevated corticosterone and norepinephrine leads to upregulation of lymphocyte beta-2 adrenergic receptors eliciting lymphocyte apoptosis (194). This suggests a critical role for sympathetic innervation of peripheral lymphoid tissues in regulating B cell response following CNS injury (261). Despite their negative roles, B cells also contribute to spinal cord repair following injury through their immunomodulatory Breg phenotype () (263). Breg cells control antigen-specific T cell autoimmune response through IL-10 production (264).

Detrimental effects of SCI-induced autoimmunity are not limited to the spinal cord. Autoreactive immune cells contribute to the exacerbation of post-SCI sequelae such as cardiovascular, renal and reproductive dysfunctions (194). For example, presence of an autoantibody against platelet prostacyclin receptor has been associated with a higher incidence of coronary artery disease in SCI patients (265). Collectively, evidence shows the critical role of adaptive immune system in SCI pathophysiology and repair. Thus, treatments that harness the pro-regenerative properties of the adaptive immune system can be utilized to reduce immune mediated tissue damage, improve neural tissue preservation and facilitate repair following SCI.

Traumatic SCI triggers the formation of a glial scar tissue around the injury epicenter (266, 267). The glial scar is a multifactorial phenomenon that is contributed f several populations in the injured spinal cord including activated astrocytes, NG2+ oligodendrocyte precursor cells (OPCs), microglia, fibroblasts, and pericytes (268271). The heterogeneous scar forming cells and associated ECM provides a cellular and biochemical zone within and around the lesion () (272). Resident and infiltrating inflammatory cells contribute to the process of glial activation and scar formation by producing cytokines (e.g., IL-1 and IL-6) chemokines and enzymes that activate glial cells or disrupt BSB (267). Activated microglia/macrophages produce proteolytic enzymes such as matrix metalloproteinases (MMPs) that increase vascular permeability and further disruption of the BSB (273). Inhibition of MMPs improves neural preservation and functional recovery in animal models of SCI (273275). In addition to glial and immune cells, fibroblasts, pericytes and ependymal cells also contribute to the structure of the glial scar (267). In penetrating injuries where meninges are compromised, meningeal fibroblasts infiltrate the lesion epicenter (276). Fibroblasts contribute to the production of fibronectin, collagen, and laminin in the ECM of the inured spinal cord (267) and are a source of axon-repulsing molecules such as semaphorins that influence axonal regeneration following SCI (277). Fibroblasts have also been found in contusive injuries where meninges are intact (268, 270). Studies using genetic fate mapping in these injuries have unraveled that perivascular pericytes and fibroblasts migrate to the injury site and form a fibrotic core in the scar which matures within 2 weeks post-injury (268, 270). SCI also triggers proliferation and migration of the stem/progenitor cell pool of the spinal cord parenchyma and ependyma. These cells can give rise to new scar forming astrocytes and OPCs (278280). In a mature glial scar, activated microglia/macrophages occupy the innermost portion closer to the injury epicenter surrounded by NG2+ OPCs () (267), while reactive astrocytes reside in the injury penumbra and form a cellular barrier (267). Of note, in human SCI, the glial scar begins to form within the first hours after the SCI and remains chronically in the spinal cord tissue (281). The glial scar has been found within the injured human spinal cord up to 42 years after the injury (267).

Activated astrocytes play a leading role in the formation of the glial scar (267). Following injury, astrocytes increase their expression of intermediate filaments, GFAP, nestin and vimentin, and become hypertrophied (282, 283). Reactive astrocytes proliferate and mobilize to the site of injury and form a mesh like structure of intermingled filamentous processes around the injury epicenter (284, 285). The astrocytic glial scar has been shown to serve as a protective barrier that prevents the spread of infiltrating immune cells into the adjacent segments (267, 284, 286). Attenuating astrocyte reactivity and scar formation by blockade of STAT3 activation results in poorer outcomes in SCI (211, 286). Reactive astrogliosis is also essential for reconstruction of the BBB, and blocking this process leads to exacerbated leukocyte infiltration, cell death, myelin damage, and reduced functional recovery (211, 285, 286). Despite the protective role of the astrocytic glial scar in acute SCI, its evolution and persistence in the sub-acute and chronic stages of injury has been considered as a potent inhibitor for spinal cord repair and regeneration (267, 287). A number of inhibitory molecules have been associated with activated astrocytes and their secreted products such as proteoglycans and Tenascin-C (288). Thus, manipulation of the astrocytic scar has been pursued as a promising treatment strategy for SCI (267, 289).

Chondroitin sulfate proteoglycans (CSPGs) are well-known for their contribution to the inhibitory role of the glial scar in axonal regeneration (290295), sprouting (296299), conduction (300302), and remyelination (241, 303307). In normal condition, basal levels of CSPGs are expressed in the CNS that play critical roles in neuronal guidance and synapse stabilization (90, 308). Following injury, CSPGs (neurocan, versican, brevican, and phosphacan) are robustly upregulated and reach their peak of expression at 2 weeks post-SCI and remain upregulated chronically (309, 310). Mechanistically, disruption of BSB and hemorrhage following traumatic SCI triggers upregulation of CSPGs in the glial scar by exposing the scar forming cells to factors in plasma such as fibrinogen (311). Studies in cortical injury have shown that fibrinogen induces CSPG expression in astrocytes through TGF/Smad2 signaling pathway (311). The authors show that intracellular Smad2 translocation is essential for Smad2 signal transduction process and its inhibition reduces scar formation (312). In contrast, another study has identified that TGF induces CSPGs production in astrocytes through a SMAD independent pathway (313). This study showed a significant upregulation of CSPGs in SMAD2 and SMAD4 knockdown astrocytes. Interestingly, CSPG upregulation was found to be mediated by the activation of the phosphoinositide 3-kinase (PI3K)/Akt and mTOR axis (313). Further studies are required to confirm these findings.

Extensive research in the past few decades has demonstrated the inhibitory effect of CSPGs on axon regeneration (314, 315). The first successful attempt on improving axon outgrowth and/or sprouting by enzymatic degradation of CSPGs using chondroitinase ABC (ChABC) in a rat SCI model was published in 2002 by Bradbury and colleagues (291). This study showed significant improvement in recovery of locomotor and proprioceptive functions following intrathecal delivery of ChABC in a rat model of dorsal column injury (291). This observation was followed by several other studies demonstrating the promise of CSPGs degradation in improvement of axon regeneration and sprouting of the serotonergic (295, 297, 299, 303), sensory (293, 298, 316), corticospinal (291, 297, 303, 317), and rubrospinal fibers (318) in animal models of CNS injury. Additionally, ChABC treatment is shown to be neuroprotective by preventing CSPG induced axonal dieback and degeneration (303, 319, 320). Studies by our group also showed that degradation of CSPGs using ChABC attenuates axonal dieback in corticospinal fibers in chronic SCI model in the rat (303). ChABC also blocks macrophage-mediated axonal degeneration in neural cultures and after SCI (238).

The inhibitory effects of astrocytic glial scar on axonal regeneration has been recently challenged after SCI (321). Using various transgenic mouse models, a study by Sofroniew's and colleagues has shown that spontaneous axon regrowth failed to happen following the ablation or prevention of astrocytic scar in acute and chronic SCI. They demonstrated that when the intrinsic ability of dorsal root ganglion (DRG) neurons for growth was enhanced by pre-conditioning injury as well as local delivery of a combination of axon growth promoting factors into the SCI lesion, the axons grew to the wall of the glial scar and CSPGs within the lesion. However, when astrocyte scarring was attenuated, the pre-conditioned/growth factor stimulated DRG neurons showed a reduced ability for axon growth (321). From these observations, the authors suggested a positive role for the astrocytic scar in axonal regeneration following SCI (321). Overall, this study points to the importance of reactive and scar forming astrocytes and their pivotal role in the repair process following SCI (322). This is indeed in agreement with previous studies by the same group that showed a beneficial role for activated astrocytes in functional recovery after SCI by limiting the speared of infiltrated inflammatory cells and tissue damage in SCI (285). It is also noteworthy that the glial scar is contributed by various cell populations and not exclusively by astrocytes (269, 271). Therefore, the outcomes of this study need to be interpreted in the context of astrocytes and astrocytic scar. Moreover, the reduced capacity of the injured spinal cord for regeneration is not solely driven by the glial scar as other factors including inflammation and damaged myelin play important inhibitory role in axon regeneration (323, 324). Taken together, further investigation is needed to delineate the mechanisms of the glial scar including the contribution of astrocyte-derived factors on axon regeneration in SCI.

While CSPGs were originally identified as an inhibitor of axon growth and plasticity within the glial scar, emerging evidence has also identified them as an important regulator of endogenous cell response. Emerging evidence has identified CSPGs as an inhibitor of oligodendrocytes (241, 272, 306). Replacement of oligodendrocytes is an important repair process in SCI and other demyelinating conditions such as MS (90). SCI and MS triggers activation of endogenous OPCs and their mobilization to the site of injury (143, 162, 306, 325). In vitro and in vivo evidence shows that CSPGs limit the recruitment of NPCs and OPCs to the lesion and inhibit oligodendrocyte survival, differentiation and maturation (145, 272, 305, 306, 326). Our group and others have shown that targeting CSPGs by ChABC administration or xyloside, or through inhibition of their signaling receptors enhances the capacity of NPCs and OPCs for proliferation, oligodendrocyte differentiation and remyelination following SCI and MS-like lesions (145, 303, 304, 306).

Mechanistically, the inhibitory effects of CSPGs on axon growth and endogenous cell differentiation is mainly governed by signaling through receptor protein tyrosine phosphatase sigma (RPTP) and leukocyte common antigen-related phosphatase receptor (LAR) (327). RPTP is the main receptor mediating the inhibition of axon growth by CSPGs (327, 328). Improved neuronal regeneration has been demonstrated in RPTP/ mice model of SCI and peripheral nerve injury (328, 329). Blockade of RPTP and LAR by intracellular sigma peptide (ISP) and intracellular LAR peptide (ILP), facilitates axon regeneration following SCI (327, 330). Inhibition of RPTP results in significant improvement in locomotion and bladder function associated with serotonergic re-innervation below the level of injury in rat SCI (327). Our group has also shown that CSPGs induce caspase-3 mediated apoptosis in NPCs and OPCs in vitro and in oligodendrocytes in the injured spinal cord that is mediated by both RPTP and LAR (241). Inhibition of LAR and RPTP sufficiently attenuates CSPG-mediated inhibition of oligodendrocyte maturation and myelination in vitro and attenuated oligodendrocyte cell death after SCI (241).

CSPGs have been implicated in regulating immune response in CNS injury and disease. Interestingly, our recent studies indicated that CSPGs signaling appears to restrict endogenous repair by promoting a pro-inflammatory immune response in SCI (241, 331). Inhibition of LAR and RPTP enhanced an anti-inflammatory environment after SCI by promoting the populations of pro-regenerative M2-like microglia/macrophages and regulatory T cells (241) that are known to promote repair process (224). These findings are also in agreement with recent studies in animal models of MS that unraveled a pro-inflammatory role for CSPGs in autoimmune demyelinating conditions (332). In MS and EAE, studies by Stephenson and colleagues have shown that CSPGs are abundant within the leucocyte-containing perivascular cuff, the entry point of inflammatory cells to the CNS tissue (332). Presence of CSPGs in these perivascular cuffs promotes trafficking of immune cells to induce a pro-inflammatory response in MS condition. In contrast to these new findings, early studies in SCI described that preventing CSPG formation with xyloside treatment at the time of injury results in poor functional outcome, while manipulation of CSPGs at 2 days after SCI was beneficial for functional recovery (333). These differential outcomes were associated with the modulatory role of CSPGs in regulating the response of macrophages/microglia. Disruption in CSPG formation immediately after injury promoted an M1 pro-inflammatory phenotype in macrophages/microglia, whereas delayed manipulation of CSPGs resulted in a pro-regenerative M2 phenotype (333). In EAE, by products of CSPG degradation also improve the outcomes by attenuating T cell infiltration and their expression of pro-inflammatory cytokines IFN- and TNF (334).

These emerging findings suggest an important immunomodulatory role for CSPGs in CNS injury and disease; further investigations are needed to elucidate CSPG mechanisms in regulating neuroinflammation. Altogether, current evidence has identified a multifaceted inhibitory role for CSPGs in regulating endogenous repair mechanisms after SCI, suggesting that targeting CSPGs may present a promising treatment strategy for SCI.

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Traumatic Spinal Cord Injury: An Overview of ...

Abstract

Stem cells or mother or queen of all cells are pleuropotent and have the remarkable potential to develop into many different cell types in the body. Serving as a sort of repair system for the body, they can theoretically divide without limit to replenish other cells as long as the person or animal is alive. When a stem cell divides, each new cell has the potential to either remain a stem cell or become another type of cell with a more specialized function, such as a muscle cell, a red blood cell, or a brain cell. Stem cells differ from other kinds of cells in the body. All stem cells regardless of their source have three general properties:

They are unspecialized; one of the fundamental properties of a stem cell is that it does not have any tissue-specific structures that allow it to perform specialized functions.

They can give rise to specialized cell types. These unspecialized stem cells can give rise to specialized cells, including heart muscle cells, blood cells, or nerve cells.

They are capable of dividing and renewing themselves for long periods. Unlike muscle cells, blood cells, or nerve cells which do not normally replicate themselves - stem cells may replicate many times. A starting population of stem cells that proliferates for many months in the laboratory can yield millions of cells. Today, donated organs and tissues are often used to replace those that are diseased or destroyed. Unfortunately, the number of people needing a transplant far exceeds the number of organs available for transplantation. Pleuropotent stem cells offer the possibility of a renewable source of replacement cells and tissues to treat a myriad of diseases, conditions, and disabilities including Parkinsons and Alzheimers diseases, spinal cord injury, stroke, Cerebral palsy, Battens disease, Amyotrophic lateral sclerosis, restoration of vision and other neuro degenerative diseases as well.

Stem cells may be the persons own cells (a procedure called autologous transplantation) or those of a donor (a procedure called allogenic transplantation). When the persons own stem cells are used, they are collected before chemotherapy or radiation therapy because these treatments can damage stem cells. They are injected back into the body after the treatment.

The sources of stem cells are varied such as pre-implantation embryos, children, adults, aborted fetuses, embryos, umbilical cord, menstrual blood, amniotic fluid and placenta

New research shows that transplanted stem cells migrate to the damaged areas and assume the function of neurons, holding out the promise of therapies for Alzheimers disease, Parkinsons, spinal cord injury, stroke, Cerebral palsy, Battens disease and other neurodegenerative diseases.

The therapeutic use of stem cells, already promising radical new treatments for cancer, immune-related diseases, and other medical conditions, may someday be extended to repairing and replenishing the brain. In a study published in the February 19, 2002, Proceedings of the National Academy of Sciences, researchers exposed the spinal cord of a rat to injury, paralyzing the animals hind limbs and lower body. Stem cells grown in exponential numbers in the laboratory were then injected into the site of the injury. It was seen that week after the injury, motor function improved dramatically,

The following diseases have been treated by various stem cell practitioners with generally positive results and the spectrum has ever since been increasing.

Cerebral palsy is a disorder caused by damage to the brain during pregnancy, delivery or shortly after birth. It is often accompanied by seizures, hearing loss, difficulty speaking, blindness, lack of co-ordination and/or mental retardation. Studies in animals with experimentally induced strokes or traumatic injuries have indicated that benefit is possible by stem cell therapy. The potential to do these transplants via injection into the vasculature rather than directly into the brain increases the likelihood of timely human studies. As a result, variables appropriate to human experiments with intravascular injection of cells, such as cell type, timing of the transplant and effect on function, need to be systematically performed in animal models Studies in animals with experimentally induced strokes or traumatic injuries have indicated that benefit is possible with injury, with the hope of rapidly translating these experiments to human trials.(1)

Cerebral palsy produces chronic motor disability in children. The causes are quite varied and range from abnormalities of brain development to birth-related injuries to postnatal brain injuries. Due to the increased survival of very premature infants, the incidence of cerebral palsy may be increasing. While premature infants and term infants who have suffered neonatal hypoxic-ischemic (HI) injury represent only a minority of the total cerebral palsy population, this group demonstrates easily identifiable clinical findings, and much of their injury is to oligodendrocytes and the white matter (2)

Alzheimers is a complex, fatal disease involving progressive cell degeneration, beginning with the loss of brain cells that control thought, memory and language. The disease, which currently has no cure, was first described by German physician Dr. Alzheimer, who discovered amyloid plaques and neurofibrillary tangles in the brain of a woman who died of an unusual mental illness. A compound similar to the components of DNA may improve the chances that stem cells transplanted from a patients bone marrow to the brain will take over the functions of damaged cells and help treat Alzheimers disease and other neurological illnesses. A research team led by University of Central Florida professor Kiminobu Sugaya found that treating bone marrow cells in laboratory cultures with bromodeoxyuridine, a compound that becomes part of DNA, made adult human stem cells more likely to develop as brain cells after they were implanted in adult rat brains.

It has long been recognized that Alzheimers disease (AD) patients present an irreversible decline of cognitive functions as consequence of cell deterioration in a structure called nucleus basalis of Meynert The reduction of the number of cholinergic cells causes interference in several aspects of behavioral performance including arousal, attention, learning and emotion. It is also common knowledge that AD is an untreatable degenerative disease with very few temporary and palliative drug therapies. Neural stem cell (NSC) grafts present a potential and innovative strategy for the treatment of many disorders of the central nervous system including AD, with the possibility of providing a more permanent remedy than present drug treatments. After grafting, these cells have the capacity to migrate to lesioned regions of the brain and differentiate into the necessary type of cells that are lacking in the diseased brain, supplying it with the cell population needed to promote recovery. (3)

Malignant multiple sclerosis (MS) is a rare but clinically important subtype of MS characterized by the rapid development of significant disability in the early stages of the disease process. These patients are refractory to conventional immunomodulatory agents and the mainstay of their treatment is plasmapheresis or immunosuppression with mitoxantrone, cyclophosphamide, cladribine or, lately, bone marrow transplantation. A report on the case of a 17-year old patient with malignant MS who was treated with high-dose chemotherapy plus anti-thymocyte globulin followed by autologous stem cell transplantation. This intervention resulted in an impressive and long-lasting clinical and radiological response (4).

In other experiment treatment of 24 patients (14 women, 10 men) with relapsing-remitting Multiple Sclerosis, in the course of 28 years was done For treatment, used were embryonic stem cell suspensions (ESCS) containing stem cells of mesenchymal and ectodermal origin obtained from active growth zones of 48 weeks old embryonic cadavers organs. Suspensions were administered in the amount of 13 ml, cell count being 0,1-100x105/ml. In the course of treatment, applied were 24 different suspensions, mode of administration being intracavitary, intravenous, and subcutaneous. After treatment, syndrome of early post-transplant improvement was observed in 70% of patients, its main manifestations being decreased weakness, improved appetite and mood, decreased depression. In the course of first post-treatment months, positive dynamics was observed in the following aspects: Nystagmus, convergence disturbances, spasticity, and coordination. In such symptoms as dysarthria, dysphagia, and ataxia, positive changes occurred at much slower rate. In general, the treatment resulted in improved range and quality of motions in the extremities, normalized muscle tone, decreased fatigue and general weakness, and improved quality of life. Forth, 87% of patients reported no exacerbations, no aggravation of neurological symptoms, and no further progression of disability. MRI performed in 12 years after the initial treatment, showed considerable subsidence of focal lesions, mean by 31%, subsidence of gadolinium enhanced lesions by 48%; T2-weighted images showed marked decrease of the focis relative density.

Doctors firstly isolated adult stem cells from the patients brain, they were then cultured in vitro and encouraged to turn into dopamine-producing neurons. As soon as tests showed that the cells were producing dopamine they were then re-injected into the mans brain. After the transplant, the mans condition was seen to improve and he experienced a reduction in the trembling and muscle rigidity associated with the disease. Brain scans taken 3-months after the transplant revealed that dopamine production had increased by 58%, however it later dropped but the Parkinsons symptoms did not return. The study is the first human study to show that stem cell transplants can help to treat Parkinsons.

The use of fetal-derived neural stem cells has shown significant promise in rodent models of Parkinsons disease, and the potential for tumorigenicity appears to be minimal. The authors report that undifferentiated human neural stem cells (hNSCs) transplanted into severely Parkinsonian 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated primates could survive, migrate, and induce behavioral recovery of Parkinsonian symptoms, which were directly related to reduced dopamine levels in the nigrostriatal system(5). Working with these cells, the researchers created dopamine neurons deficient in DJ-1, a gene mutated in an inherited form of Parkinsons. They report that DJ-1-deficient cells -- and especially DJ-1-deficient dopamine neurons -- display heightened sensitivity to oxidative stress, caused by products of oxygen metabolism that react with and damage cellular components like proteins and DNA. In a second paper, they link DJ-1 dysfunction to the aggregation of alpha-synuclein, a hallmark of Parkinsons neuropathology. (6,7)

In summary most of studies using aborted human embryonic tissue indicate that:

Clinical benefit does occur; however, the benefit is not marked and there is a delay of many months before the clinical change.

Postmortem examinations show that tissue grafts do survive and innervate the striatum.

PET scans show that there is an increase in dopamine uptake after transplantation.

Followup studies show that long term benefit does occur with transplantation.(8)

During and after a stroke, certain cellular events take place that lead to the death of brain cells. Compounds that inhibit a group of enzymes called histone deacetylases can modulate gene expression, and in some cases produce cellular proteins that are actually neuroprotective -- they are able to block cell death. Great deal of research has gone into developing histone deacetylase inhibitors as novel therapeutics (9)

One Mesenchymal stem cell (MSC) transplantation improves recovery from ischemic stroke in animals. The Researchers examined the feasibility, efficacy, and safety of cell therapy using culture-expanded autologous MSCs in patients with ischemic stroke. They prospectively and randomly allocated 30 patients with cerebral infarcts within the middle cerebral arterial territory Serial evaluations showed no adverse cell-related, serological, or imaging-defined effects. In patients with severe cerebral infarcts, the intravenous infusion of autologous MSCs appears to be a feasible and safe therapy that may improve functional recovery.(10)

Early intravenous stem cell injection displayed anti-inflammatory functionality that promoted neuroprotection, mainly by interrupting splenic inflammatory responses after intra cranial Haemorrage.

In summary, early intravenous NSC injection displayed anti-inflammatory functionality that neural stem cell (NSC) transplantation has been investigated as a means to reconstitute the damaged brain after stroke. In this study, however, was investigated the effect on acute cerebral and peripheral inflammation after intracerebral haemorrhage (ICH). STEM CELLS from fetal human brain were injected intravenously (NSCs-iv, 5 million cells) or intracerebrally (NSCs-ic, 1 million cells) at 2 or 24 h after collagenase-induced ICH in a rat model. Only NSCs-iv-2 h resulted in fewer initial neurologic deteriorations and reduced brain edema formation, inflammatory infiltrations and apoptosis. (11)

Emerging cell therapies for the restoration of sight have focused on two areas of the eye that are critical for visual function, the cornea and the retina. The relatively easy access of the cornea, the homogeneity of the cells forming the different layers of the corneal epithelium and the improvement of cell culture protocols are leading to considerable success in corneal epithelium restoration. Rebuilding the entire cornea is however still far from reality. The restoration of the retina has recently been achieved in different animal models of retinal degeneration using immature photoreceptors (12)

Bone marrow contains stem cells, which have the extraordinary abilities to home in on injuries and possibly regenerate other cell types in the body. In this case, the cells were transplanted to confirm that bone marrow does regenerate the injured RPE. Damage to RPE is present in many diseases of the retina, including age-related macular degeneration, which affects more than 1.75 million people in the United States. (13)

Neural stem cells (NSCs) offer the potential to replace lost tissue after nervous system injury. Thus, stem cells can promote host neural repair in part by secreting growth factors, and their regeneration-promoting activities can be modified by gene delivery.

Attempted repair of human spinal cord injury by transplantation of stem cells depends on complex biological interactions between the host and graft

Extrapolating results from experimental therapy in animals to humans with spinal cord injury requires great caution.

There is great pressure on surgeons to transplant stem cells into humans with spinal cord injury. However, as the efficacy of and exact indications for this therapy are still uncertain, and morbidity (such as rejection or late tumour development) may result, only carefully designed studies based on sound experimental work which attempts to eliminate placebo effects should proceed.

Premature application of stem cell transplantation in humans with spinal cord injury should be discouraged. 14, 15, 16)

Attempted repair of human spinal cord injury by transplantation of stem cells depends on complex biological interactions between the host and graft

Extrapolating results from experimental therapy in animals to humans with spinal cord injury requires great caution.

There is great pressure on surgeons to transplant stem cells into humans with spinal cord injury. However, as the efficacy of and exact indications for this therapy are still uncertain, and morbidity (such as rejection or late tumour development) may result, only carefully designed studies based on sound experimental work which attempts to eliminate placebo effects should proceed.

Premature application of stem cell transplantation in humans with spinal cord injury should be discouraged.

Mesenchymal stem cells have also been identified and are currently being developed for bone, cartilage, muscle, tendon, and ligament repair and regeneration. These MSCs are typically harvested, isolated, and expanded from bone marrow or adipose tissue, and they have been isolated from rodents, dogs, and humans. Interestingly, these cells can undergo extensive sub cultivation in vitro without differentiation, magnifying their potential clinical use.(17) Human MSCs can be directed toward osteoblastic differentiation by adding dexamethasone, ascorbic acid, and -glycerophosphate to the tissue culture media. This osteoblastic commitment and differentiation can be clearly documented by analyzing alkaline phosphatase activity, the expression of bone matrix proteins, and the mineralization of the extracellular matrix.(18)

Children with Battens disease suffer seizures, motor control disturbances, blindness and communication problems. As many as 600 children in the US are currently diagnosed with the condition.(19)

Death can occur in children as young as 8 years old. The children lack an enzyme for breaking down complex fat and protein compounds in the brain, explains Robert Steiner, vice chair of paediatric research at the hospital. The material accumulates and interferes with tissue function, ultimately causing brain cells to die. Tests on animals demonstrated that stem cells injected into the brain secreted the missing enzyme. And the stem cells were found to survive well in the rodent brain. Once injected, the purified neural cells may develop into neurons or other nervous system tissue, including oligodendrocytes, or glial cells, which support the neurons(20).

In a study that demonstrates the promise of cell-based therapies for diseases that have proved intractable to modern medicine, a team of scientists from the University of Wisconsin-Madison has shown it is possible to rescue the dying neurons characteristic of amyotrophic lateral sclerosis (ALS), a fatal neuromuscular disorder also known as Lou Gehrigs disease. Previously there was no effective treatments for ALS, which afflicts roughly 40,000 people in the United States and which is almost always fatal within three to five years of diagnosis. Patients gradually experience progressive muscle weakness and paralysis as the motor neurons that control muscles are destroyed by the disease

In the new Wisconsin study, nascent brain cells known as neural progenitor cells derived from human fetal tissue were engineered to secrete a chemical known as glial cell line derived neurotrophic factor (GDNF), an agent that has been shown to protect neurons but that is very difficult to deliver to specific regions of the brain. The engineered cells were then implanted in the spinal cords of rats afflicted with a form of ALS. The implanted cells, in fact, demonstrated an affinity for the areas of the spinal cord where motor neurons were dying. The cells after being injected to the area of damage where they just sit and release GDNF. At the early stages of disease, almost 100 percent protection of motor neurons was seen. (21)

In other study MSCs were isolated from bone marrow of 9 patients with definite ALS. Growth kinetics, immunophenotype, telomere length and karyotype were evaluated during in vitro expansion. No significant differences between donors or patients were observed. The patients received intraspinal injections of autologous MSCs at the thoracic level and monitored for 4 years. No significant acute or late side effects were evidenced. No modification of the spinal cord volume or other signs of abnormal cell proliferation were observed. The results seem to demonstrate that MSCs represent a good chance for stem cell cell-based therapy in ALS and that intraspinal injection of MSCs is safe also in the long term. A new phase 1 study is carried out to verify these data in a larger number of patients. (22)

Stem-cell-based technology offers amazing possibilities for the future. These include the ability to reproduce human tissues and potentially repair damaged organs (such as the brain, spinal cord, vertebral column the eye), where, at present, we mainly provide supportive care to prevent the situation from becoming worse. This potential almost silences the sternest critics of such technology, but the fact remains that the ethical challenges are daunting. It is encouraging that, in tackling these challenges, we stand to reflect a great deal about the ethics of our profession and our relationships with patients, industry, and each other. The experimental basis of stem-cell or OEC transplantation should be sound before these techniques are applied to humans with neurological disorders.

1. Stem cell therapy for cerebral palsy. Bartley J, Carroll JE. Department of Pediatrics of the Medical College of Georgia, Augusta, Georgia, USA

8. Department of Neurology, Mt. Sinai School of Medicine, New York, NY, Medscape journal. Stem Cell Transplantation for Parkinsons Disease

9. Journal of Medicinal Chemistry. Future Therapies For Stroke May Block Cell Death 16 Jun 2007

10. Neurosurg Focus. 2005;19(6) 2005 American Association of Neurological Surgeons

11. Brain Advance Access originally published online on December 20, 2007 Brain 2008 Anti-inflammatory mechanism of intravascular neural stem cell transplantation in haemorrhagic stroke.

13. University of Florida(2006, June 8). Bone Marrow May Restore Cells Lost In Vision Diseases. ScienceDaily.

18. Autologous mesenchymal stem cell transplantation in stroke patients Oh Young Bang, MD, PhD 1, Jin Soo Lee, MD Department of Neurology, School of Medicine, Ajou University, Suwon, South Korea Brain Disease Research Center, School of Medicine, Ajou University, Suwon, South Korea.

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Role of Stem Cells in Treatment of Neurological Disorder

Every week there are numerous scientific studies published. Heres a look at some of the more interesting ones.

T-Cell Immune Response to COVID-19 Vaccines and Natural Infections

Much of the discussions and news reports about immune responses to vaccines and COVID-19 revolve around antibody levels. Much less has been said about T-cells, which provide longer-term protection. Researchers atGladstone Institutesconducteda detailed T-cell survey before and after COVID-19 immunization, which they published ineLife. They concluded that the Pfizer-BioNTech and Moderna mRNA vaccines create long-term populations of T-cells that recognize multiple SARS-COV-2 virus variants. They also found key differences in the T-cell responses in people who had COVID-19 infections before vaccination compared to people who had never been infected.

Overall, our data support the idea that vaccines are eliciting a very robust T-cell response in healthy individuals, said Nadia Roan, senior author of the study and Gladstone Associate Investigator. But they also suggest there may be some ways to improve them further, by getting more of the vaccine-elicited T-cells to park themselves in the respiratory tract.

Antibodies produced by B-cells quickly recognize viruses, target them, and prevent infection by destroying the viruses. T-cells, however, identify and destroy cells that are already infected. Antibodies are better at stopping initial infection, but T-cells typically last longer after an initial infection or vaccine. At that point they are better at fighting off disease in its early stages, which prevents severe symptoms. But T-cells are very diverse and difficult to analyze. Some subsets respond differently to infected cells and behave differently, while others have different functions within the overall T-cell immune response.

One key finding was that in people who had not been previously infected, the T-cell responses become stronger in quantity and quality after the second dose of the vaccine. But in people who had previously had COVID-19, there was not much of a change between the first and second vaccine dose.

Blood Biomarkers Provide Warning Signs of Dementia

Investigators at theGerman Center for Neurodegenerative Diseases (DZNE)identifiedmolecules in the blood that potentially warn of impending dementia. The research study included several university hospitals across Germany. The biomarkers were based on measuring levels of microRNAs. They say that the technique isnt ready yet for practical use, but they hope to develop a simple blood test. MicroRNAs have regulatory properties, influencing protein production and metabolism. In tests in humans, mice and cell cultures, they found three microRNAs whose levels were linked to mental performance. The three microRNAs also influence neuro-inflammation and neuroplasticity, including the ability of neurons to establish connections with each other.

Stem Cell Population Essential for Bone Regeneration

Researchers at theUniversity of Tsukuba, Japan,identifieda subpopulation of mesenchymal stem cells that play a major role in bone healing. The stem cells are found in the bone marrow and express the marker CD73. When a bone fracture heals, it moves through a series of stages, including clotted blood forming at the fracture. This clot is replaced by fibrous tissue and cartilage, then by a hard bony callus. The bone is then remodeled, with regular bone replacing the hard callus. They found that the generation of the callus is critically dependent on recruiting MSCs from the surrounding tissue and bone marrow. They observed the CD73-positive MSCs migrating toward the fracture site and forming new cartilage and bone cells. When they grafted CD73-positive MSCs into the fracture, they noted enhanced healing processes.

Antiviral Molecule Prevents SARS-CoV-2 from Entering Cells

Scientists atWashington University School of Medicinein St. Louisdevelopeda compound that prevents the SARS-CoV-2 virus, which causes COVID-19, from entering cells. The compound is called MM3122 and has been studied in cell cultures and in mice. MM3122 targets a key human protein called transmembrane serine protease 2 (TMPRSS2), which coronaviruses use to enter and infect human cells. Once the virus attached onto a cell in the epithelia of the airway, the TMPRSS2 protein cuts the viral spike protein, which activates the spike protein to mediate fusion of the viral and cellular membranesstarting the infection process. In cell cultures, MM3122 protected cells from viral damage better than remdesivir,Gilead Sciences antiviral against COVID-19; and an acute safety assay in mice demonstrated that large doses of MM3122 given for seven days did not cause noticeable issues. The compound also was effective against SARS-CoV, the virus behind SARS, and MERS-CoV, the coronavirus that causes MERS. The researchers are now working with researchers at the NIH to test it in animal models of COVID-19. They are also working on an oral version of the injectable compound.

Specific Personality Traits Might Signal Pending Alzheimers

Researchers atFlorida State Universityfoundthat specific changes in the brain linked with Alzheimers disease are often visible earlier in people with personality traits associated with the disease. The research focused on two traits: neuroticism, or a predisposition for negative emotions, and conscientiousness, linked to a tendency to be careful, organized, goal-directed and responsible. They found that people with amyloid and tau deposits, proteins linked to Alzheimers disease in the brain, were identified in participants who scored higher in neuroticism levels and lower in conscientiousness. The study suggests that personality traits might help protect against Alzheimers and other brain diseases by delaying or preventing the neuropathology for people strong in conscientiousness and low in neuroticism.

Why We Overeat

Astudyfrom theUniversity of Washington School of Medicine/UW Medicinereported on the function of glutamatergic neurons in mice. These neurons communicate to the lateral habenula, a brain region associated with the pathophysiology of depression, and the ventral tegmental area, which is involved in motivation, reward and addiction. They found that when mice are eating, the neurons in the lateral habenula are more responsive than the neurons in the ventral tegmental area. They suggest that these neurons might play a bigger role in guiding feeding. In addition, they studied the influence of the leptin and ghrelin hormones, which are believed to regulate behavior via the mesolimbic dopamine system, part of the reward pathway. The research adds additional insight into satiety and why people or at least mice overeat.

We found these cells are not a monolithic group, and that different flavors of these cells do different things, said Garret Stuber, a joint UW professor of anesthesiology and pain medicine and pharmacology, the papers senior author.

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Research Roundup: T-Cell Immune Response to COVID-19 Vaccines and More - BioSpace

NEW YORK, Oct. 14, 2021 /PRNewswire/ --BrainStorm Cell Therapeutics Inc. (NASDAQ: BCLI), a leading developer of cellular therapies for neurodegenerative diseases, will present findings from a multicenter, open label clinical trial of NurOwn in progressive multiple sclerosis. The study, "Phase 2 Safety and Efficacy Study of Intrathecal MSC-NTF cells in Progressive Multiple Sclerosis," will be delivered in an oral presentation today at the fully digital37thCongress of the European Committee for Treatment and Research in Multiple Sclerosis (ECTRIMS).

The Phase 2 clinical trial was designed to evaluate intrathecal administration of NurOwn (autologous MSC-NTF cells) in participants with progressive MS. The study achieved the primary endpoint of safety and tolerability. It demonstrated a reduction of neuroinflammatory biomarkers and an increase in neuroprotective biomarkers in the cerebrospinal fluid (CSF) and consistent improvement across MS functional outcome measures, including measures of walking, upper extremity function, vision and cognition.

"We were pleased that this study demonstrated safety, preliminary evidence of efficacy and relevant biomarker outcomes in patients with progressive multiple sclerosis, in an area of high unmet need," said Jeffrey Cohen, M.D., Director of Experimental Therapeutics at the Cleveland Clinic Mellen Center for MS and principal investigator for the trial. "These results should be confirmed in a randomized placebo-controlled trial."

The study was sponsored by Brainstorm Cell Therapeutics with additional financial support for biomarker analyses from the National Multiple Sclerosis Society Fast-Forward Program. It was conducted at four U.S. MS centers of excellence:

"We very much appreciate the tremendous collaboration among many premier organizations, for their generous sharing of expertise, support and data, which enabled the important balance between scientific rigor and ethical treatment of progressive MS participants in the trial," said Ralph Kern, M.D., MHSc., President and Chief Medical Officer, Brainstorm Cell Therapeutics. "We are holding discussions with key MS experts, and seeking guidance from the FDA to determine next steps for the development of NurOwn in progressive MS."

"The National MS Society is pleased to support the biomarker portion of this study through our commercial funding program Fast Forward," said Mark Allegretta, Ph.D., Vice President, Research. "We're encouraged to see evidence that the biomarker analysis showed proof of concept for detecting neuroprotection and reduced inflammation."

About the trial

The Phase 2 open-label studyevaluated the safety and efficacy of intrathecal administration of autologous MSC-NTF cells in patients with primary or secondary progressive MS. The primary study endpoint was safety and tolerability. Secondary efficacy endpoints included: timed 25-foot walk (T25FW); 9-Hole Peg Test (9-HPT); Low Contrast Letter Acuity (LCLA); Symbol Digit Modalities Test (SDMT); 12 item MS Walking Scale (MSWS-12); as well as cerebrospinal fluid (CSF) and blood biomarkers. Clinical efficacy outcomes were compared with matched (n=48) participants in the Comprehensive Longitudinal Investigation of Multiple Sclerosis (CLIMB) registry, Tanuja Chitnis, MD Brigham and Women's Hospital and the Ann Romney Center for Neurologic Diseases, and 255 patient randomized double blind placebo controlled NN-102 SPRINT-MS Study, courtesy NIH/NINDS, PI: Robert J. Fox, MD, MS, FAAN, Cleveland Clinic, CTR: NCT01982942. Baseline characteristics from these two cohorts were similar allowing for comparison of efficacy results, comparisons with SPRINT-MS were with the placebo arm of this study.

Mean age of participants was 47 years, 56% were female, and mean baseline EDSS score was 5.4. 18 participants were treated, 16 (80%) received all 3 treatments and completed the entire study; 2 study discontinuations were due to procedure-related adverse events. No deaths or treatment-related adverse events due to worsening of MS were observed.

In responder analyses, 14% and 13% of MSC-NTF treated participants showed at least a 25% improvement in T25FW and 9-HPT (combined hands) respectively, compared to 5% and 0% in matched CLIMB patients and 9% and 3% in SPRINT. Twenty-seven percent (27%) showed at least an 8-letter improvement in LCLA (binocular, 2.5% threshold) and 67% showed at least a 3-point improvement in SDMT, compared to 6% and 18% in CLIMB and 13% and 35% in SPRINT, respectively.

Mean improvements of +0.10 ft/sec in T25FW and -0.23 sec in 9-HPT (combined hands), were observed in MSC-NTF treated participants, compared to a mean worsening of -0.07 ft/sec and +0.49 sec in CLIMB and -0.06 ft/sec and +0.28 sec in SPRINT, respectively. MSC-NTF treated participants showed a mean improvement of +3.3 letters in LCLA (binocular, 2.5% threshold) and 3.8 points in SDMT, compared to a mean worsening of -1.07 letters in LCLA (binocular, 2.5% threshold) and mean improvement of +0.10 in SDMT, in CLIMB and -0.6 and -0.1 in SPRINT. In addition the MSFC-4 Composite Z-score of T25W, 9-HPT, SDMT and LCLA showed a 0.18 point improvement in MSC-NTF treated participants, while CLIMB and SPRINT showed decreases of -0.02 and -0.05.

Furthermore, 38% of treated patients showed at least a 10-point improvement in the MSWS-12 a patient reported outcome that evaluates the impact of MS on walking function, whereas this outcome was not evaluated in CLIMB or SPRINT.

CSF biomarkers obtained at 3 consecutive time points, showed increases in neuroprotective molecules (VEGF, HGF, NCAM-1,Follistatin, Fetuin-A) and decreases in neuroinflammatory biomarkers (MCP-1, SDF-1, sCD27 and Osteopontin).

About NurOwn

The NurOwntechnology platform (autologous MSC-NTF cells) represents a promising investigational therapeutic approach to targeting disease pathways important in neurodegenerative disorders. MSC-NTF cells are produced from autologous, bone marrow-derived mesenchymal stem cells (MSCs) that have been expanded and differentiated ex vivo. MSCs are converted into MSC-NTF cells by growing them under patented conditions that induce the cells to secrete high levels of neurotrophic factors (NTFs). Autologous MSC-NTF cells are designed to effectively deliver multiple NTFs and immunomodulatory cytokines directly to the site of damage to elicit a desired biological effect and ultimately slow or stabilize disease progression.

About BrainStorm Cell Therapeutics Inc.

BrainStorm Cell Therapeutics Inc. is a leading developer of innovative autologous adult stem cell therapeutics for debilitating neurodegenerative diseases. The Company holds the rights to clinical development and commercialization of the NurOwntechnology platform used to produce autologous MSC-NTF cells through an exclusive, worldwide licensing agreement. Autologous MSC-NTF cells have received Orphan Drug designation status from the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) for the treatment of amyotrophic lateral sclerosis (ALS). BrainStorm has completed a Phase 3 pivotal trial in ALS (NCT03280056); this trial investigated the safety and efficacy of repeat-administration of autologous MSC-NTF cells and was supported by a grant from the California Institute for Regenerative Medicine (CIRM CLIN2-0989). BrainStorm completed under an investigational new drug application a Phase 2 open-label multicenter trial (NCT03799718) of autologous MSC-NTF cells in progressive multiple sclerosis (MS) and was supported by a grant from the National MS Society (NMSS).

For more information, visit the company's website atwww.brainstorm-cell.com.

Safe-Harbor Statement

Statements in this announcement other than historical data and information, including statements regarding future NurOwnmanufacturing and clinical development plans, constitute "forward-looking statements" and involve risks and uncertainties that could cause BrainStorm Cell Therapeutics Inc.'s actual results to differ materially from those stated or implied by such forward-looking statements. Terms and phrases such as "may," "should," "would," "could," "will," "expect,""likely," "believe," "plan," "estimate," "predict," "potential," and similar terms and phrases are intended to identify these forward-looking statements. The potential risks and uncertainties include, without limitation, BrainStorm's need to raise additional capital, BrainStorm's ability to continue as a going concern, the prospects for regulatory approval of BrainStorm's NurOwntreatment candidate, the initiation, completion, and success of BrainStorm's product development programs and research, regulatory and personnel issues, development of a global market for our services, the ability to secure and maintain research institutions to conduct our clinical trials, the ability to generate significant revenue, the ability of BrainStorm's NurOwntreatment candidate to achieve broad acceptance as a treatment option for ALS or other neurodegenerative diseases, BrainStorm's ability to manufacture, or to use third parties to manufacture, and commercialize the NurOwntreatment candidate, obtaining patents that provide meaningful protection, competition and market developments, BrainStorm's ability to protect our intellectual property from infringement by third parties, heath reform legislation, demand for our services, currency exchange rates and product liability claims and litigation; and other factors detailed in BrainStorm's annual report on Form 10-K and quarterly reports on Form 10-Q available athttp://www.sec.gov. These factors should be considered carefully, and readers should not place undue reliance on BrainStorm's forward-looking statements. The forward-looking statements contained in this press release are based on the beliefs, expectations and opinions of management as of the date of this press release. We do not assume any obligation to update forward-looking statements to reflect actual results or assumptions if circumstances or management's beliefs, expectations or opinions should change, unless otherwise required by law. Although we believe that the expectations reflected in the forward-looking statements are reasonable, we cannot guarantee future results, levels of activity, performance or achievements.

Contacts:

Investor Relations:Eric GoldsteinLifeSci Advisors, LLCPhone: +1 (646) 791-9729egoldstein@lifesciadvisors.com

Media:Mariesa Kemble kemblem@mac.com

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KENILWORTH, N.J. & TOKYO--(BUSINESS WIRE)--Merck (NYSE: MRK), known as MSD outside the United States and Canada, and Eisai today announced that the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency has adopted positive opinions recommending approval of the combination of KEYTRUDA, Mercks anti-PD-1 therapy, plus LENVIMA (marketed as KISPLYX in the European Union [EU] for the treatment of advanced renal cell carcinoma [RCC]), the orally available multiple receptor tyrosine kinase inhibitor discovered by Eisai, for two different indications. One positive opinion is for the first-line treatment of adult patients with advanced RCC, and the other is for the treatment of adult patients with advanced or recurrent endometrial carcinoma (EC) who have disease progression on or following prior treatment with a platinum-containing therapy in any setting and are not candidates for curative surgery or radiation. Decisions on the CHMPs recommendations will be given by the European Commission for marketing authorization in the EU, and are expected in the fourth quarter of 2021. If approved, this would be the first combination of an anti-PD-1 therapy with a tyrosine kinase inhibitor approved for the treatment of two different types of cancer in the EU.

The positive CHMP opinions are based on data from two pivotal Phase 3 trials: CLEAR (Study 307)/KEYNOTE-581 evaluating the combination in adult patients with advanced RCC and KEYNOTE-775/Study 309 evaluating the combination in certain patients with advanced EC.

In CLEAR/KEYNOTE-581, KEYTRUDA plus LENVIMA demonstrated statistically significant improvements versus sunitinib in the efficacy outcome measures of overall survival (OS), reducing the risk of death by 34% (HR=0.66 [95% CI, 0.49-0.88]; p=0.0049) versus sunitinib, and progression-free survival (PFS), reducing the risk of disease progression or death by 61% (HR=0.39 [95% CI, 0.32-0.49]; p<0.0001) with a median PFS of 23.9 months versus 9.2 months for sunitinib. Additionally, the confirmed objective response rate was 71% (95% CI: 66-76) (n=252) for patients who received KEYTRUDA plus LENVIMA versus 36% with sunitinib (95% CI: 31-41) (n=129).

In KEYNOTE-775/Study 309, KEYTRUDA plus LENVIMA demonstrated statistically significant improvements in the studys dual efficacy outcome measures of OS, reducing the risk of death by 38% (HR=0.62 [95% CI, 0.51-0.75]; p<0.0001) with a median OS of 18.3 months versus 11.4 months for chemotherapy (investigators choice of doxorubicin or paclitaxel), and PFS, reducing the risk of disease progression or death by 44% (HR=0.56 [95% CI, 0.47-0.66]; p<0.0001), with a median PFS of 7.2 months versus 3.8 months for chemotherapy (investigators choice of doxorubicin or paclitaxel).

KEYTRUDA plus LENVIMA demonstrated a survival benefit for advanced renal cell carcinoma in the first-line setting and represents an important potential new treatment option for these patients. Additionally, KEYTRUDA plus LENVIMA is the first anti-PD-1 and tyrosine kinase inhibitor combination to demonstrate a survival benefit in advanced endometrial carcinoma patients, and the benefit was shown regardless of mismatch repair status, said Dr. Gregory Lubiniecki, Vice President, Clinical Research, Merck Research Laboratories. We are pleased that the CHMP has recognized the important role of the combination therapy in these difficult-to-treat cancers.

We appreciate the positive opinions rendered by the EU CHMP recommending approval of KEYTRUDA plus LENVIMA in advanced renal cell carcinoma and advanced endometrial carcinoma, underscoring the potential significance of the outcomes observed in the CLEAR/KEYNOTE-581 and KEYNOTE-775/Study 309 trials, said Dr. Takashi Owa, President, Oncology Business Group at Eisai. We are grateful to the patients who participated in these studies, their families and clinicians. Their commitment made these meaningful milestones possible.

The safety of KEYTRUDA in combination with axitinib or LENVIMA in advanced RCC, and in combination with LENVIMA in advanced EC has been evaluated in a total of 1,456 patients with advanced RCC or advanced EC. In these patient populations, the most frequent adverse reactions were diarrhea (58%), hypertension (54%), hypothyroidism (46%), fatigue (41%), decreased appetite and nausea (40% each), arthralgia (30%), vomiting, weight decreased, dysphonia and abdominal pain (28% each), proteinuria (27%), palmar plantar erythrodysesthesia syndrome and rash (26% each), stomatitis and constipation (25% each), musculoskeletal pain and headache (23% each) and cough (21%).

About Renal Cell Carcinoma (RCC)

Worldwide, it is estimated there were more than 431,000 new cases of kidney cancer diagnosed and more than 179,000 deaths from the disease in 2020. In Europe, it is estimated there were more than 138,000 new cases of kidney cancer diagnosed and more than 54,000 deaths from the disease in 2020. Renal cell carcinoma is by far the most common type of kidney cancer; about nine out of 10 kidney cancer diagnoses are RCC. Renal cell carcinoma is about twice as common in men as in women. Most cases of RCC are discovered incidentally during imaging tests for other abdominal diseases. Approximately 30% of patients with RCC will have metastatic disease at diagnosis. Survival is highly dependent on the stage at diagnosis, and the five-year survival rate is 13% for patients diagnosed with metastatic disease.

About Endometrial Cancer

Endometrial cancer begins in the inner lining of the uterus, which is known as the endometrium and is the most common type of cancer in the uterus. Worldwide, it was estimated there were more than 417,000 new cases and more than 97,000 deaths from uterine body cancers in 2020 (these estimates include both endometrial cancers and uterine sarcomas; more than 90% of uterine body cancers occur in the endometrium, so the actual numbers for endometrial cancer cases and deaths are slightly lower than these estimates). In Europe, it is estimated there were more than 130,000 new cases and more than 29,000 deaths in 2020. The five-year relative survival rate for metastatic endometrial cancer (stage IV) is estimated to be approximately 17%.

About KEYTRUDA (pembrolizumab) Injection, 100 mg

KEYTRUDA is an anti-programmed death receptor-1 (PD-1) therapy that works by increasing the ability of the bodys immune system to help detect and fight tumor cells. KEYTRUDA is a humanized monoclonal antibody that blocks the interaction between PD-1 and its ligands, PD-L1 and PD-L2, thereby activating T lymphocytes which may affect both tumor cells and healthy cells.

Merck has the industrys largest immuno-oncology clinical research program. There are currently more than 1,600 trials studying KEYTRUDA across a wide variety of cancers and treatment settings. The KEYTRUDA clinical program seeks to understand the role of KEYTRUDA across cancers and the factors that may predict a patient's likelihood of benefitting from treatment with KEYTRUDA, including exploring several different biomarkers.

Selected KEYTRUDA (pembrolizumab) Indications in the U.S.

Melanoma

KEYTRUDA is indicated for the treatment of patients with unresectable or metastatic melanoma.

KEYTRUDA is indicated for the adjuvant treatment of patients with melanoma with involvement of lymph node(s) following complete resection.

Non-Small Cell Lung Cancer

KEYTRUDA, in combination with pemetrexed and platinum chemotherapy, is indicated for the first-line treatment of patients with metastatic nonsquamous non-small cell lung cancer (NSCLC), with no EGFR or ALK genomic tumor aberrations.

KEYTRUDA, in combination with carboplatin and either paclitaxel or paclitaxel protein-bound, is indicated for the first-line treatment of patients with metastatic squamous NSCLC.

KEYTRUDA, as a single agent, is indicated for the first-line treatment of patients with NSCLC expressing PD-L1 [tumor proportion score (TPS) 1%] as determined by an FDA-approved test, with no EGFR or ALK genomic tumor aberrations, and is:

KEYTRUDA, as a single agent, is indicated for the treatment of patients with metastatic NSCLC whose tumors express PD-L1 (TPS 1%) as determined by an FDA-approved test, with disease progression on or after platinum-containing chemotherapy. Patients with EGFR or ALK genomic tumor aberrations should have disease progression on FDA-approved therapy for these aberrations prior to receiving KEYTRUDA.

Head and Neck Squamous Cell Cancer

KEYTRUDA, in combination with platinum and fluorouracil (FU), is indicated for the first-line treatment of patients with metastatic or with unresectable, recurrent head and neck squamous cell carcinoma (HNSCC).

KEYTRUDA, as a single agent, is indicated for the first-line treatment of patients with metastatic or with unresectable, recurrent HNSCC whose tumors express PD-L1 [combined positive score (CPS 1)] as determined by an FDA-approved test.

KEYTRUDA, as a single agent, is indicated for the treatment of patients with recurrent or metastatic HNSCC with disease progression on or after platinum-containing chemotherapy.

Classical Hodgkin Lymphoma

KEYTRUDA is indicated for the treatment of adult patients with relapsed or refractory classical Hodgkin lymphoma (cHL).

KEYTRUDA is indicated for the treatment of pediatric patients with refractory cHL, or cHL that has relapsed after 2 or more lines of therapy.

Primary Mediastinal Large B-Cell Lymphoma

KEYTRUDA is indicated for the treatment of adult and pediatric patients with refractory primary mediastinal large B-cell lymphoma (PMBCL), or who have relapsed after 2 or more prior lines of therapy. KEYTRUDA is not recommended for treatment of patients with PMBCL who require urgent cytoreductive therapy.

Urothelial Carcinoma

KEYTRUDA is indicated for the treatment of patients with locally advanced or metastatic urothelial carcinoma (mUC):

Non-muscle Invasive Bladder Cancer

KEYTRUDA is indicated for the treatment of patients with Bacillus Calmette-Guerin (BCG)-unresponsive, high-risk, non-muscle invasive bladder cancer (NMIBC) with carcinoma in situ (CIS) with or without papillary tumors who are ineligible for or have elected not to undergo cystectomy.

Microsatellite Instability-High or Mismatch Repair Deficient Cancer

KEYTRUDA is indicated for the treatment of adult and pediatric patients with unresectable or metastatic microsatellite instability-high (MSI-H) or mismatch repair deficient (dMMR) solid tumors that have progressed following prior treatment and who have no satisfactory alternative treatment options.

This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials. The safety and effectiveness of KEYTRUDA in pediatric patients with MSI-H central nervous system cancers have not been established.

Microsatellite Instability-High or Mismatch Repair Deficient Colorectal Cancer

KEYTRUDA is indicated for the treatment of patients with unresectable or metastatic MSI-H or dMMR colorectal cancer (CRC).

Gastric Cancer

KEYTRUDA, in combination with trastuzumab, fluoropyrimidine- and platinum-containing chemotherapy, is indicated for the first-line treatment of patients with locally advanced unresectable or metastatic HER2-positive gastric or gastroesophageal junction (GEJ) adenocarcinoma.

This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

KEYTRUDA, as a single agent, is indicated for the treatment of patients with recurrent locally advanced or metastatic gastric or gastroesophageal junction (GEJ) adenocarcinoma whose tumors express PD-L1 (CPS 1) as determined by an FDA-approved test, with disease progression on or after 2 or more prior lines of therapy including fluoropyrimidine- and platinum-containing chemotherapy and if appropriate, HER2/neu-targeted therapy.

This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Esophageal Cancer

KEYTRUDA is indicated for the treatment of patients with locally advanced or metastatic esophageal or GEJ (tumors with epicenter 1 to 5 centimeters above the GEJ) carcinoma that is not amenable to surgical resection or definitive chemoradiation either:

Cervical Cancer

KEYTRUDA, in combination with chemotherapy, with or without bevacizumab, is indicated for the treatment of patients with persistent, recurrent, or metastatic cervical cancer whose tumors express PD-L1 (CPS 1) as determined by an FDA-approved test.

KEYTRUDA, as a single agent, is indicated for the treatment of patients with recurrent or metastatic cervical cancer with disease progression on or after chemotherapy whose tumors express PD-L1 (CPS 1) as determined by an FDA-approved test.

Hepatocellular Carcinoma

KEYTRUDA is indicated for the treatment of patients with hepatocellular carcinoma (HCC) who have been previously treated with sorafenib. This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Merkel Cell Carcinoma

KEYTRUDA is indicated for the treatment of adult and pediatric patients with recurrent locally advanced or metastatic Merkel cell carcinoma (MCC). This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials.

Renal Cell Carcinoma

KEYTRUDA, in combination with axitinib, is indicated for the first-line treatment of adult patients with advanced renal cell carcinoma (RCC).

KEYTRUDA, in combination with LENVIMA, is indicated for the first-line treatment of adult patients with advanced RCC.

Endometrial Carcinoma

KEYTRUDA, in combination with LENVIMA, is indicated for the treatment of patients with advanced endometrial carcinoma that is not MSI-H or dMMR, who have disease progression following prior systemic therapy in any settings and are not candidates for curative surgery or radiation.

Tumor Mutational Burden-High Cancer

KEYTRUDA is indicated for the treatment of adult and pediatric patients with unresectable or metastatic tumor mutational burden-high (TMB-H) [10 mutations/megabase] solid tumors, as determined by an FDA-approved test, that have progressed following prior treatment and who have no satisfactory alternative treatment options. This indication is approved under accelerated approval based on tumor response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in the confirmatory trials. The safety and effectiveness of KEYTRUDA in pediatric patients with TMB-H central nervous system cancers have not been established.

Cutaneous Squamous Cell Carcinoma

KEYTRUDA is indicated for the treatment of patients with recurrent or metastatic cutaneous squamous cell carcinoma (cSCC) or locally advanced cSCC that is not curable by surgery or radiation.

Triple-Negative Breast Cancer

KEYTRUDA is indicated for the treatment of patients with high-risk early-stage triple-negative breast cancer (TNBC) in combination with chemotherapy as neoadjuvant treatment, and then continued as a single agent as adjuvant treatment after surgery.

KEYTRUDA, in combination with chemotherapy, is indicated for the treatment of patients with locally recurrent unresectable or metastatic TNBC whose tumors express PD-L1 (CPS 10) as determined by an FDA-approved test.

Selected Important Safety Information for KEYTRUDA

Severe and Fatal Immune-Mediated Adverse Reactions

KEYTRUDA is a monoclonal antibody that belongs to a class of drugs that bind to either the PD-1 or the PD-L1, blocking the PD-1/PD-L1 pathway, thereby removing inhibition of the immune response, potentially breaking peripheral tolerance and inducing immune-mediated adverse reactions. Immune-mediated adverse reactions, which may be severe or fatal, can occur in any organ system or tissue, can affect more than one body system simultaneously, and can occur at any time after starting treatment or after discontinuation of treatment. Important immune-mediated adverse reactions listed here may not include all possible severe and fatal immune-mediated adverse reactions.

Monitor patients closely for symptoms and signs that may be clinical manifestations of underlying immune-mediated adverse reactions. Early identification and management are essential to ensure safe use of antiPD-1/PD-L1 treatments. Evaluate liver enzymes, creatinine, and thyroid function at baseline and periodically during treatment. For patients with TNBC treated with KEYTRUDA in the neoadjuvant setting, monitor blood cortisol at baseline, prior to surgery, and as clinically indicated. In cases of suspected immune-mediated adverse reactions, initiate appropriate workup to exclude alternative etiologies, including infection. Institute medical management promptly, including specialty consultation as appropriate.

Withhold or permanently discontinue KEYTRUDA depending on severity of the immune-mediated adverse reaction. In general, if KEYTRUDA requires interruption or discontinuation, administer systemic corticosteroid therapy (1 to 2 mg/kg/day prednisone or equivalent) until improvement to Grade 1 or less. Upon improvement to Grade 1 or less, initiate corticosteroid taper and continue to taper over at least 1 month. Consider administration of other systemic immunosuppressants in patients whose adverse reactions are not controlled with corticosteroid therapy.

Immune-Mediated Pneumonitis

KEYTRUDA can cause immune-mediated pneumonitis. The incidence is higher in patients who have received prior thoracic radiation. Immune-mediated pneumonitis occurred in 3.4% (94/2799) of patients receiving KEYTRUDA, including fatal (0.1%), Grade 4 (0.3%), Grade 3 (0.9%), and Grade 2 (1.3%) reactions. Systemic corticosteroids were required in 67% (63/94) of patients. Pneumonitis led to permanent discontinuation of KEYTRUDA in 1.3% (36) and withholding in 0.9% (26) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 23% had recurrence. Pneumonitis resolved in 59% of the 94 patients.

Pneumonitis occurred in 8% (31/389) of adult patients with cHL receiving KEYTRUDA as a single agent, including Grades 3-4 in 2.3% of patients. Patients received high-dose corticosteroids for a median duration of 10 days (range: 2 days to 53 months). Pneumonitis rates were similar in patients with and without prior thoracic radiation. Pneumonitis led to discontinuation of KEYTRUDA in 5.4% (21) of patients. Of the patients who developed pneumonitis, 42% interrupted KEYTRUDA, 68% discontinued KEYTRUDA, and 77% had resolution.

Immune-Mediated Colitis

KEYTRUDA can cause immune-mediated colitis, which may present with diarrhea. Cytomegalovirus infection/reactivation has been reported in patients with corticosteroid-refractory immune-mediated colitis. In cases of corticosteroid-refractory colitis, consider repeating infectious workup to exclude alternative etiologies. Immune-mediated colitis occurred in 1.7% (48/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (1.1%), and Grade 2 (0.4%) reactions. Systemic corticosteroids were required in 69% (33/48); additional immunosuppressant therapy was required in 4.2% of patients. Colitis led to permanent discontinuation of KEYTRUDA in 0.5% (15) and withholding in 0.5% (13) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 23% had recurrence. Colitis resolved in 85% of the 48 patients.

Hepatotoxicity and Immune-Mediated Hepatitis

KEYTRUDA can cause immune-mediated hepatitis. Immune-mediated hepatitis occurred in 0.7% (19/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.4%), and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 68% (13/19) of patients; additional immunosuppressant therapy was required in 11% of patients. Hepatitis led to permanent discontinuation of KEYTRUDA in 0.2% (6) and withholding in 0.3% (9) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, none had recurrence. Hepatitis resolved in 79% of the 19 patients.

KEYTRUDA with Axitinib

KEYTRUDA in combination with axitinib can cause hepatic toxicity. Monitor liver enzymes before initiation of and periodically throughout treatment. Consider monitoring more frequently as compared to when the drugs are administered as single agents. For elevated liver enzymes, interrupt KEYTRUDA and axitinib, and consider administering corticosteroids as needed. With the combination of KEYTRUDA and axitinib, Grades 3 and 4 increased alanine aminotransferase (20%) and increased aspartate aminotransferase (13%) were seen at a higher frequency compared to KEYTRUDA alone. Fifty-nine percent of the patients with increased ALT received systemic corticosteroids. In patients with ALT 3 times upper limit of normal (ULN) (Grades 2-4, n=116), ALT resolved to Grades 0-1 in 94%. Among the 92 patients who were rechallenged with either KEYTRUDA (n=3) or axitinib (n=34) administered as a single agent or with both (n=55), recurrence of ALT 3 times ULN was observed in 1 patient receiving KEYTRUDA, 16 patients receiving axitinib, and 24 patients receiving both. All patients with a recurrence of ALT 3 ULN subsequently recovered from the event.

Immune-Mediated Endocrinopathies

Adrenal Insufficiency

KEYTRUDA can cause primary or secondary adrenal insufficiency. For Grade 2 or higher, initiate symptomatic treatment, including hormone replacement as clinically indicated. Withhold KEYTRUDA depending on severity. Adrenal insufficiency occurred in 0.8% (22/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.3%), and Grade 2 (0.3%) reactions. Systemic corticosteroids were required in 77% (17/22) of patients; of these, the majority remained on systemic corticosteroids. Adrenal insufficiency led to permanent discontinuation of KEYTRUDA in <0.1% (1) and withholding in 0.3% (8) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Hypophysitis

KEYTRUDA can cause immune-mediated hypophysitis. Hypophysitis can present with acute symptoms associated with mass effect such as headache, photophobia, or visual field defects. Hypophysitis can cause hypopituitarism. Initiate hormone replacement as indicated. Withhold or permanently discontinue KEYTRUDA depending on severity. Hypophysitis occurred in 0.6% (17/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.3%), and Grade 2 (0.2%) reactions. Systemic corticosteroids were required in 94% (16/17) of patients; of these, the majority remained on systemic corticosteroids. Hypophysitis led to permanent discontinuation of KEYTRUDA in 0.1% (4) and withholding in 0.3% (7) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Thyroid Disorders

KEYTRUDA can cause immune-mediated thyroid disorders. Thyroiditis can present with or without endocrinopathy. Hypothyroidism can follow hyperthyroidism. Initiate hormone replacement for hypothyroidism or institute medical management of hyperthyroidism as clinically indicated. Withhold or permanently discontinue KEYTRUDA depending on severity. Thyroiditis occurred in 0.6% (16/2799) of patients receiving KEYTRUDA, including Grade 2 (0.3%). None discontinued, but KEYTRUDA was withheld in <0.1% (1) of patients.

Hyperthyroidism occurred in 3.4% (96/2799) of patients receiving KEYTRUDA, including Grade 3 (0.1%) and Grade 2 (0.8%). It led to permanent discontinuation of KEYTRUDA in <0.1% (2) and withholding in 0.3% (7) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement. Hypothyroidism occurred in 8% (237/2799) of patients receiving KEYTRUDA, including Grade 3 (0.1%) and Grade 2 (6.2%). It led to permanent discontinuation of KEYTRUDA in <0.1% (1) and withholding in 0.5% (14) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement. The majority of patients with hypothyroidism required long-term thyroid hormone replacement. The incidence of new or worsening hypothyroidism was higher in 1185 patients with HNSCC, occurring in 16% of patients receiving KEYTRUDA as a single agent or in combination with platinum and FU, including Grade 3 (0.3%) hypothyroidism. The incidence of new or worsening hypothyroidism was higher in 389 adult patients with cHL (17%) receiving KEYTRUDA as a single agent, including Grade 1 (6.2%) and Grade 2 (10.8%) hypothyroidism.

Type 1 Diabetes Mellitus (DM), Which Can Present With Diabetic Ketoacidosis

Monitor patients for hyperglycemia or other signs and symptoms of diabetes. Initiate treatment with insulin as clinically indicated. Withhold KEYTRUDA depending on severity. Type 1 DM occurred in 0.2% (6/2799) of patients receiving KEYTRUDA. It led to permanent discontinuation in <0.1% (1) and withholding of KEYTRUDA in <0.1% (1) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement.

Immune-Mediated Nephritis With Renal Dysfunction

KEYTRUDA can cause immune-mediated nephritis. Immune-mediated nephritis occurred in 0.3% (9/2799) of patients receiving KEYTRUDA, including Grade 4 (<0.1%), Grade 3 (0.1%), and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 89% (8/9) of patients. Nephritis led to permanent discontinuation of KEYTRUDA in 0.1% (3) and withholding in 0.1% (3) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, none had recurrence. Nephritis resolved in 56% of the 9 patients.

Immune-Mediated Dermatologic Adverse Reactions

KEYTRUDA can cause immune-mediated rash or dermatitis. Exfoliative dermatitis, including Stevens-Johnson syndrome, drug rash with eosinophilia and systemic symptoms, and toxic epidermal necrolysis, has occurred with antiPD-1/PD-L1 treatments. Topical emollients and/or topical corticosteroids may be adequate to treat mild to moderate nonexfoliative rashes. Withhold or permanently discontinue KEYTRUDA depending on severity. Immune-mediated dermatologic adverse reactions occurred in 1.4% (38/2799) of patients receiving KEYTRUDA, including Grade 3 (1%) and Grade 2 (0.1%) reactions. Systemic corticosteroids were required in 40% (15/38) of patients. These reactions led to permanent discontinuation in 0.1% (2) and withholding of KEYTRUDA in 0.6% (16) of patients. All patients who were withheld reinitiated KEYTRUDA after symptom improvement; of these, 6% had recurrence. The reactions resolved in 79% of the 38 patients.

Other Immune-Mediated Adverse Reactions

The following clinically significant immune-mediated adverse reactions occurred at an incidence of <1% (unless otherwise noted) in patients who received KEYTRUDA or were reported with the use of other antiPD-1/PD-L1 treatments. Severe or fatal cases have been reported for some of these adverse reactions. Cardiac/Vascular: Myocarditis, pericarditis, vasculitis; Nervous System: Meningitis, encephalitis, myelitis and demyelination, myasthenic syndrome/myasthenia gravis (including exacerbation), Guillain-Barr syndrome, nerve paresis, autoimmune neuropathy; Ocular: Uveitis, iritis and other ocular inflammatory toxicities can occur. Some cases can be associated with retinal detachment. Various grades of visual impairment, including blindness, can occur. If uveitis occurs in combination with other immune-mediated adverse reactions, consider a Vogt-Koyanagi-Harada-like syndrome, as this may require treatment with systemic steroids to reduce the risk of permanent vision loss; Gastrointestinal: Pancreatitis, to include increases in serum amylase and lipase levels, gastritis, duodenitis; Musculoskeletal and Connective Tissue: Myositis/polymyositis, rhabdomyolysis (and associated sequelae, including renal failure), arthritis (1.5%), polymyalgia rheumatica; Endocrine: Hypoparathyroidism; Hematologic/Immune: Hemolytic anemia, aplastic anemia, hemophagocytic lymphohistiocytosis, systemic inflammatory response syndrome, histiocytic necrotizing lymphadenitis (Kikuchi lymphadenitis), sarcoidosis, immune thrombocytopenic purpura, solid organ transplant rejection.

Infusion-Related Reactions

KEYTRUDA can cause severe or life-threatening infusion-related reactions, including hypersensitivity and anaphylaxis, which have been reported in 0.2% of 2799 patients receiving KEYTRUDA. Monitor for signs and symptoms of infusion-related reactions. Interrupt or slow the rate of infusion for Grade 1 or Grade 2 reactions. For Grade 3 or Grade 4 reactions, stop infusion and permanently discontinue KEYTRUDA.

Complications of Allogeneic Hematopoietic Stem Cell Transplantation (HSCT)

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Merck and Eisai Receive Positive EU CHMP Opinions for KEYTRUDA (pembrolizumab) Plus LENVIMA (lenvatinib) in Two Different Types of Cancer - Business...

Every person, every mouse, every dog, has one unmistakable sign of aging: hair loss. But why does that happen?

Rui Yi, a professor of pathology at Northwestern University, set out to answer the question.

A generally accepted hypothesis about stem cells says they replenish tissues and organs, including hair, but they will eventually be exhausted and then die in place. This process is seen as an integral part of aging.

Instead Dr. Yi and his colleagues made a surprising discovery that, at least in the hair of aging animals, stem cells escape from the structures that house them.

Its a new way of thinking about aging, said Dr. Cheng-Ming Chuong, a skin cell researcher and professor of pathology at the University of Southern California, who was not involved in Dr. Yis study, which was published on Monday in the journal Nature Aging.

The study also identifies two genes involved in the aging of hair, opening up new possibilities for stopping the process by preventing stem cells from escaping.

Charles K.F. Chan, a stem cell researcher at Stanford University, called the paper very important, noting that in science, everything about aging seems so complicated we dont know where to start. By showing a pathway and a mechanism for explaining aging hair, Dr. Yi and colleagues may have provided a toehold.

Stem cells play a crucial role in the growth of hair in mice and in humans. Hair follicles, the tunnel-shaped miniature organs from which hairs grow, go through cyclical periods of growth in which a population of stem cells living in a specialized region called the bulge divide and become rapidly growing hair cells.

Sarah Millar, director of the Black Family Stem Cell Institute at the Icahn School of Medicine at Mount Sinai, who was not involved in Dr. Yis paper, explained that those cells give rise to the hair shaft and its sheath. Then, after a period of time, which is short for human body hair and much longer for hair on a persons head, the follicle becomes inactive and its lower part degenerates. The hair shaft stops growing and is shed, only to be replaced by a new strand of hair as the cycle repeats.

But while the rest of the follicle dies, a collection of stem cells remains in the bulge, ready to start turning into hair cells to grow a new strand of hair.

Dr. Yi, like most scientists, had assumed that with age the stem cells died in a process known as stem cell exhaustion. He expected that the death of a hair follicles stem cells meant that the hair would turn white and, when enough stem cells were lost, the strand of hair would die. But this hypothesis had not been fully tested.

Together with a graduate student, Chi Zhang, Dr. Yi decided that to understand the aging process in hair, he needed to watch individual strands of hair as they grew and aged.

Ordinarily, researchers who study aging take chunks of tissue from animals of different ages and examine the changes. There are two drawbacks to this approach, Dr. Yi said. First, the tissue is already dead. And it is not clear what led to the changes that are observed or what will come after them.

He decided his team would use a different method. They watched the growth of individual hair follicles in the ears of mice using a long wavelength laser that can penetrate deep into tissue. They labeled hair follicles with a green fluorescent protein, anesthetized the animals so they did not move, put their ear under the microscope and went back again and again to watch what was happening to the same hair follicle.

What they saw was a surprise: When the animals started to grow old and gray and lose their hair, their stem cells started to escape their little homes in the bulge. The cells changed their shapes from round to amoeba-like and squeezed out of tiny holes in the follicle. Then they recovered their normal shapes and darted away.

Sometimes, the escaping stem cells leapt long distances, in cellular terms, from the niche where they lived.

If I did not see it for myself I would not have believed it, Dr. Yi said. Its almost crazy in my mind.

The stem cells then vanished, perhaps consumed by the immune system.

Dr. Chan compared an animal's body to a car. If you run it long enough and dont replace parts, things wear out, he said. In the body, stem cells are like a mechanic, providing replacement parts, and in some organs like hair, blood and bone, the replacement is continual.

But with hair, it now looks as if the mechanic the stem cells simply walks off the job one day.

But why? Dr. Yi and his colleagues next step was to ask if genes are controlling the process. They discovered two FOXC1 and NFATC1 that were less active in older hair follicle cells. Their role was to imprison stem cells in the bulge. So the researchers bred mice that lacked those genes to see if they were the master controllers.

By the time the mice were 4 to 5 months old, they started losing hair. By age 16 months, when the animals were middle-aged, they looked ancient: They had lost a lot of hair and the sparse strands remaining were gray.

Now the researchers want to save the hair stem cells in aging mice.

This story of the discovery of a completely unexpected natural process makes Dr. Chuong wonder what remains to be learned about living creatures.

Nature has endless surprises waiting for us, he said. You can see fantastic things.

Excerpt from:
Losing Your Hair? You Might Blame the Great Stem Cell Escape. - The New York Times