DUBLIN, Feb. 27, 2020 /PRNewswire/ -- The "Stem Cell Banking - Market Analysis, Trends, and Forecasts" report has been added to ResearchAndMarkets.com's offering.

The growing interest in regenerative medicine which involves replacing, engineering or regenerating human cells, tissues or organs, will drive market growth of stem cells. Developments in stem cells bioprocessing are important and will be a key factor that will influence and help regenerative medicine research move into real-world clinical use. The impact of regenerative medicine on healthcare will be comparable to the impact of antibiotics, vaccines, and monoclonal antibodies in current clinical care. With the global regenerative medicine market poised to reach over US$45 billion by 2025, demand for stem cells will witness robust growth.

Another emerging application area for stem cells is in drug testing in the pharmaceutical field. New drugs in development can be safely, accurately, and effectively be tested on stem cells before commencing tests on animal and human models. Among the various types of stem cells, umbilical cord stem cells are growing in popularity as they are easy and safe to extract. After birth blood from the umbilical cord is extracted without posing risk either to the mother or the child. As compared to embryonic and fetal stem cells which are saddled with safety and ethical issues, umbilical cord is recovered postnatally and is today an inexpensive and valuable source of multipotent stem cells. Until now discarded as waste material, umbilical cord blood is today acknowledged as a valuable source of blood stem cells. The huge gap between newborns and available cord blood banks reveals huge untapped opportunity for developing and establishing a more effective banking system for making this type of stem cells viable for commercial scale production and supply. Umbilical cord and placenta contain haematopoietic blood stem cells (HSCs). These are the only cells capable of producing immune system cells (red cells, white cells and platelet).

HSCs are valuable in the treatment of blood diseases and successful bone marrow transplants. Also, unlike bone marrow stem cells, umbilical cord blood has the advantage of having 'off-the-shelf' uses as it requires no human leukocyte antigen (HLA) tissue matching. Developments in stem cell preservation will remain crucial for successful stem cell banking. Among the preservation technologies, cryopreservation remains popular. Development of additives for protecting cells from the stresses of freezing and thawing will also be important for the future of the market. The United States and Europe represent large markets worldwide with a combined share of 60.5% of the market. China ranks as the fastest growing market with a CAGR of 10.8% over the analysis period supported by the large and growing network of umbilical cord blood banks in the country. The Chinese government has, over the years, systematically nurtured the growth of umbilical cord blood (UCB) banks under the 'Developmental and Reproductive Research Initiation' program launched in 2008. Several hybrid public-private partnerships and favorable governmental licensing policies today are responsible for the current growth in this market.

Competitors identified in this market include:

Companies Mentioned

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Global Stem Cell Market Poised for Strong Growth as Global Regenerative Medicine Market Poised to Reach US$45 billion by 2025 - P&T Community

Bone Marrow Stem Cells Comments Off on Global Stem Cell Market Poised for Strong Growth as Global Regenerative Medicine Market Poised to Reach US$45 billion by 2025 – P&T Community | February 29th, 2020

A bone marrow donor registry drive on Texas A&M Universitys campus brought more than 400 students to Duncan Dining Hall on Friday.

Participants swabbed their mouths to provide DNA samples and have their names added to Be the Matchs national bone marrow registry, which will help people in need of bone marrow connect with donors. The event was a collaborative effort between the A&M Corps of Cadets, the Kidz1stFund and Be the Match.

If anyone is matched, Community Engagement Representative for Be the Match Gulf Coast Benita Davis said they will need to have additional blood work done before donating.

A&M senior and Cadet 1st Lt. Mitchell Moore said his attendance on Friday was motivated in part by his interest in the medical field, since he is aiming to go to medical school.

Its minimum to no risk for you and not too much time, Moore said about donating. A small time on your part can make a huge difference and extend someones life by years.

A&M football coach Jimbo Fisher and Candi Fisher started Kidz1stFund in 2011 to raise money for research about fanconi anemia, which is a rare blood disorder that their teenage son was diagnosed with. Fisher stopped by Fridays event to speak with organizers and meet participants.

Its amazing how many great people there are in this world who are willing to help other people, Fisher said. These young men and women out here are actually saving lives.

Davis said donating is not as painful as many people often think, especially since about 80% of donations can be made with the nonsurgical method of giving peripheral blood stem cells. The other 20% of donations involve marrow being removed from the hip while the donor is asleep under general anesthesia. According to the Be the Match website, about one in 430 people on the registry end up donating.

The cadets in attendance on Friday were a reflection of who they are as people and students, according to Amy Thompson, assistant commandant, marketing and communications for the Corps of Cadets.

Selfless service is an A&M core value its also a core value for the Corps of Cadets, Thompson said. The commandant really supports and encourages cadets to seek out opportunities to be leaders in selfless service. This is one of those opportunities where we can do that on a very large scale and make a huge impact.

Corps Squadron 1 commander and A&M senior Jacob Svetz donated using stem cells about two years ago. He said he thinks everyone should sign up for the registry.

The few pin pricks that you get Its such a miniscule amount of pain compared to what that family and individual are suffering through, Svetz said. To put yourself into a position to be able to help that for me, it doesnt make sense not to.

Be the Match On Campus President and A&M senior Paige Boone said the organization hosts drives every month. The next one will be March 25 at A&Ms Rudder Plaza. Home swab kits are also an option. To get started, visit join.bethematch.org or text CORPS to 61474. Anyone ages 18 to 44 can participate.

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Jimbo Fisher's Kidz1stFund, Aggie Corps of Cadets team up for Be the Match bone marrow donor registry drive - Bryan-College Station Eagle

Bone Marrow Stem Cells Comments Off on Jimbo Fisher’s Kidz1stFund, Aggie Corps of Cadets team up for Be the Match bone marrow donor registry drive – Bryan-College Station Eagle | February 29th, 2020

AgeX Therapeutics, Inc. ("AgeX"; NYSE American: AGE), a biotechnology company focused on developing therapeutics for human aging and regeneration, announced a new paper co-authored by two AgeX scientists that could lead to new insights into the fundamental mechanisms of aging and why super-centenarians not only live the longest, but also experience extraordinary healthspans; an extension of the healthy years of life that compresses morbidity to a very short period near the end of life. The paper, "Induced pluripotency and spontaneous reversal of cellular aging in supercentenarian donor cells," is published online in the peer-reviewed scientific journal "Biochemical and Biophysical Research Communications" from Elsevier. The senior author is Dana Larocca, PhD, VP of Discovery Research at AgeX, and the first author is Jieun Lee, PhD, Scientist at AgeX.

"Clearly, we can learn a lot about aging and longevity from the longest of the long-lived, the supercentenarians, and we hope that this paper accelerates such research," commented Dr. Larocca. "Now that we have converted the cells of one of the longest-lived people in history, a deceased 114-year-old American woman, to a young pluripotent state, researchers can do so with cells from other supercentenarians. The goal is to understand specifically how these "extreme agers" manage to avoid the major chronic illnesses of aging better than any other age group including centenarians. We can essentially put their cells in a time machine and revert them to an earlier state, then study their biology to help unlock the mysteries of super-longevity. Scientists have long wondered, and now we know that we can indeed reset the developmental state and cellular age in the oldest of the old."

By way of comparison, the paper also describes undertaking a similar process with cells from two other donors: an eight-year-old with a rapid-aging syndrome commonly known as Progeria, and a 43-year-old, healthy disease-free control (HDC) subject. The paper notes that the supercentenarians cells reverted to induced pluripotent stem (iPS) cells at the same rate as the HDC subject and the Progeria patient. However, there may be some negative impact of extreme age on telomere resetting as this did not occur as frequently in the supercentenarian as in the other two donors.

The donated cells were from "the longevity collection," a cell bank established by the NIHs National Institute on Aging.

About AgeX Therapeutics

AgeX Therapeutics, Inc. (NYSE American: AGE) is focused on developing and commercializing innovative therapeutics for human aging. Its PureStem and UniverCyte manufacturing and immunotolerance technologies are designed to work together to generate highly-defined, universal, allogeneic, off-the-shelf pluripotent stem cell-derived young cells of any type for application in a variety of diseases with a high unmet medical need. AgeX has two preclinical cell therapy programs: AGEX-VASC1 (vascular progenitor cells) for tissue ischemia and AGEX-BAT1 (brown fat cells) for Type II diabetes. AgeXs revolutionary longevity platform induced Tissue Regeneration (iTR) aims to unlock cellular immortality and regenerative capacity to reverse age-related changes within tissues. AGEX-iTR1547 is an iTR-based formulation in preclinical development. HyStem is AgeXs delivery technology to stably engraft PureStem cell therapies in the body. AgeX is developing its core product pipeline for use in the clinic to extend human healthspan and is seeking opportunities to establish licensing and collaboration agreements around its broad IP estate and proprietary technology platforms.

For more information, please visit http://www.agexinc.com or connect with the company on Twitter, LinkedIn, Facebook, and YouTube.

Forward-Looking Statements

Certain statements contained in this release are "forward-looking statements" within the meaning of the Private Securities Litigation Reform Act of 1995. Any statements that are not historical fact including, but not limited to statements that contain words such as "will," "believes," "plans," "anticipates," "expects," "estimates" should also be considered forward-looking statements. Forward-looking statements involve risks and uncertainties. Actual results may differ materially from the results anticipated in these forward-looking statements and as such should be evaluated together with the many uncertainties that affect the business of AgeX Therapeutics, Inc. and its subsidiaries particularly those mentioned in the cautionary statements found in more detail in the "Risk Factors" section of AgeXs Annual Report on Form 10-K and Quarterly Reports on Form 10-Q filed with the Securities and Exchange Commissions (copies of which may be obtained at http://www.sec.gov). Subsequent events and developments may cause these forward-looking statements to change. AgeX specifically disclaims any obligation or intention to update or revise these forward-looking statements as a result of changed events or circumstances that occur after the date of this release, except as required by applicable law.

View source version on businesswire.com: https://www.businesswire.com/news/home/20200228005122/en/

Contacts

Media Contact for AgeX:

Bill Douglass Gotham Communications, LLCbill@gothamcomm.com (646) 504-0890

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AgeX Therapeutics Researchers Publish Paper on the Age Reprogramming of Super-Centenarian Cells - Yahoo Finance

IPS Cell Therapy Comments Off on AgeX Therapeutics Researchers Publish Paper on the Age Reprogramming of Super-Centenarian Cells – Yahoo Finance | February 29th, 2020

INTRODUCTION

Severe combined immunodeficiency (SCID) is a heterogeneous group of genetic diseases characterized by severe T cell lymphopenia, causing increased susceptibility to viral, bacterial, and fungal infections since early in life (1). Most forms of SCID are due to genetic defects that are intrinsic to hematopoietic cells and can be successfully treated by allogeneic hematopoietic stem cell transplantation (HSCT). However, SCID may also be caused by genetic abnormalities that are intrinsic to thymic epithelium development and function; in such cases, thymus transplantation, but not hematopoietic cell transplantation, is required to cure the disease. Only a few genetic abnormalities, including complete DiGeorge syndrome, and pathogenic variants affecting FOXN1 or CHD7, are known to cause SCID as a result of abnormal thymic development in humans (1).

PAX1 is a member of the paired box (PAX) family of transcription factors and plays a critical role in pattern formation during embryogenesis. It is expressed in the pharyngeal pouches that give rise to the thymus, tonsils, parathyroid glands, thyroid, and middle ear development during human embryogenesis (2). Pax1 deficiency in mice is characterized by anomalies of the vertebral column and variable degrees of thymic hypoplasia and thymocyte number and maturation (35). In humans, a homozygous pathogenic PAX1 p.Gly166Val variant (6) and a homozygous frameshift insertion (c.1173_1174insGCCCG) (7) have been identified in patients with otofaciocervical syndrome type 2 (OTFCS2), a rare disorder characterized by facial dysmorphism, external ear anomalies with preauricular pits and hearing impairment, branchial cysts or fistulas, anomalies of the vertebrae and the shoulder girdle, and mild intellectual disability. Recently, another homozygous pathogenic PAX1 variant (p.Cys368*) has been reported in two affected children from a consanguineous family of North African descent, who presented with OTFCS2 associated with T B+ SCID (8). However, limited information was provided on the immunological phenotype of these patients, and the functional consequences of the PAX1 variant were not investigated. Here, we provide an in-depth clinical, biochemical, and immunological description of multiple patients with OTFCS2 associated with SCID who carried biallelic deleterious PAX1 variants. By performing transfection experiments, molecular modeling, molecular dynamics (MD) simulation, and in vitro differentiation of control- and patient-derived induced pluripotent stem cells (iPSCs) to thymic epithelial progenitor (TEP) cells, we sought to assess the effects of human PAX1 deficiency on thymus development and function.

Patient 1 (P1) is a male infant born to parents whose families were from the same rural region in Germany (Fig. 1A). Bilateral microtia, malar prominence, narrow alae nasi, cupid bow lip, and retrognathia were noticed at birth (fig. S1, A and B). Imaging studies demonstrated severely stenotic external auditory canal on the right side and narrow left auditory canal (fig. S1C), congenital kyphosis at C3-C4 and L3 levels, moderate spinal canal narrowing (fig. S1, D to F), and traction on the cauda equina (fig. S1G). Diffuse erythematous rash (fig. S1H), lymphadenopathy, elevated serum immunoglobulin E (IgE), and eosinophilia were present, consistent with Omenn syndrome. On chest x-ray, the thymus shadow was not visible, and split cervical vertebral bodies, hooked distal clavicles, and a shallow dysplastic glenoid fossa were seen (fig. S1I). This infection history during infancy included Staphylococcus aureus bacteremia, pneumonia, cellulitis, and diarrhea due to Clostridium difficile.

(A) Pedigrees and results of Sanger sequencing in patients with PAX1 variants and in healthy controls. For both family A and family B, results of Sanger sequencing in the heterozygous parents are also shown. (B) Schematic representation of the PAX1 protein and location of the variants identified in affected individuals.

P2 and P3 have been previously described (8) as patients V:1 and V:18, respectively, and are part of a large consanguineous family of Moroccan origin (Fig. 1A). At birth, P2 was noticed to have frontal and parietal bossing, hypertelorism, small nose with hypoplastic nasal root, low-set ears with agenesis of the left pinna and hypoplasia of the right pinna, scapular winging, and bilateral cryptorchidism. Imaging studies showed impaired development of internal auditory canals bilaterally and lack of a thymic shadow. P3 manifested similar facial dysmorphisms as P2, along with left facial nerve palsy, severe dorsal and lumbar scoliosis, and deafness. Imaging studies documented lack of thymic shadow, abnormal appearance of vertebrae, clavicles and shoulder blades, narrowing of both external auditory canals (fig. S1J), abnormalities of the middle ear, and presence of tubular structures with features of a dental element behind the mandibular condyle (fig. S1, K and L). Subject V:3 from the same family died early in life with a history of severe infections, but no formal medical records are available.

P4 and P7 are siblings born to consanguineous parents from Saudi Arabia. P7 was noticed to have severe bilateral microtia, postauricular sinuses, and micrognathia. He suffered from chronic diarrhea, recurrent respiratory infections, exfoliative dermatitis, regional dissemination of Bacille Calmette-Guerin (BCG-itis), and lymphadenopathy and died at 1 year of age.

P4 is a female with a history of chronic diarrhea, recurrent respiratory infections, and poor weight gain since the age of 1 month. Physical examination showed small malformed ears, a skin tag on the right ear, facial asymmetry, small nose with depressed nasal bridge, and small almond-shaped eyes. A skeletal survey showed wedge-shaped vertebral body at T11 and deficient posterior element of the sacrum at S4 and S5.

P5 and P6 were siblings born to consanguineous parents and belonged to the same extended family as P4 and P7. P5 had small, low-set malformed ears, triangular mouth, down-slanting palpebral fissures, a small nose with a depressed nasal bridge, and right facial palsy. She developed recurrent respiratory infections, chronic diarrhea, severe exfoliative dermatitis, and BCG-itis and was diagnosed with Omenn syndrome. She died at 8 months of age with progressive severe pneumonitis.

P6 was screened for immunodeficiency at birth because of the positive family history. She had malformed and low-set small ears, small chin, protruding forehead, and generalized eczema. A skeletal survey showed central depression of the vertebral bodies in the thoracic and lumbar spine. Her immunological workup was consistent with T B+ NK+ (natural killerpositive) SCID. She suffered from recurrent respiratory infections and chronic diarrhea and died at 9 months of age with respiratory syncytial virus (RSV) pneumonia.

The main immunological findings at presentation in P1 to P6 are shown in Table 1. In particular, P1 had significant T cell lymphopenia. His CD4+ lymphocytes were largely (98%) CD45R0+, no CD4+ CD45RA+ CD31+ cells were detected, and T cell proliferation to phytohemagglutinin (PHA) was impaired (fig. S2A). T cell receptor (TCR) excision circles (TRECs) were below the limit of detection, indicating lack of thymopoiesis. TCR V spectratyping revealed T cell oligoclonality (fig. S2B). Elevated serum IgE and eosinophilia were present, consistent with an Omenn syndrome presentation.

AEoC, absolute eosinophil count; ALC, absolute lymphocyte count; ANC, absolute neutrophil count; n.d.: not done; cpm, counts per minute.

Laboratory investigations in P2 at 2 weeks of age revealed profound T cell lymphopenia, markedly reduced proliferative response to mitogens, and increased serum IgE. An inguinal lymph node biopsy showed severe lymphoid depletion, with primary follicles without germinal centers, associated with nearly complete absence of CD3+ T cells, but presence of B and NK cells and sparse plasma cells, and increased number of CD68+ histiocytes and eosinophils (fig. S3). A diagnosis of T B+ NK+ SCID was established.

Severe T cell lymphopenia was observed in P3, P4, and P6, associated with virtually absent in vitro T cell proliferation to PHA in P4 and P6, consistent with a diagnosis of T B+ NK+ SCID (Table 1). Last, P5 was diagnosed as having Omenn syndrome based on generalized erythroderma, lymphocytosis, eosinophilia, hypogammaglobulinemia, increased IgE, and severely reduced in vitro T cell proliferation to PHA.

Because of severe immunological abnormalities, HSCT was attempted in P1 to P4 before the gene defect was known. Details of transplant, chimerism, and immune reconstitution are shown in Table 2. In all cases, a conditioning regimen was used. Two patients (P1 and P4) attained full donor chimerism. P2 failed to engraft, developed interstitial pneumonitis, and died 5.5 months after HSCT. In P3, initial engraftment was followed by secondary graft failure, and a second HSCT was performed, resulting in mixed chimerism. Although three of the patients attained either full or mixed donor chimerism, none of them achieved reconstitution of the T cell compartment. In P1, who exhibits full donor chimerism, all T cells have a CD45R0+ phenotype and therefore likely represent donor-derived T cells contained in the graft that have undergone peripheral expansion. P3 attained mixed chimerism but remained with persistent severe T cell lymphopenia. She developed Pneumocystis jiroveci pneumonia, recurrent gastrointestinal infections, and liver failure and died of septic shock at the age of 4 years and 7 months. P4, who attained full chimerism but failed to reconstitute T cells, developed severe autoimmune hemolytic anemia, requiring multiple courses of rituximab and immunosuppressive therapy. Together, these data indicate that HSCT was unable to correct the profound T cell immunodeficiency of this disease.

ATG, anti-thymocyte globulin; PBSC, peripheral blood stem cells; URD, unrelated donor.

Before HSCT, karyotype analysis revealed no cytogenetic abnormalities in P1, P2, and P3. No evidence for copy number variation (CNV) was found by chromosomal microarray analysis in P1, and search for 22q11 deletion in P2 by in situ fluorescence hybridization was negative. No pathogenic variants in any of the known SCID-causing genes were identified in P4 by a targeted next-generation sequencing primary immunodeficiency gene panel. In an attempt to define the molecular mechanisms of the disease, whole-exome sequencing (WES) was performed in P1, P2, and P4 independently (fig. S4 and table S1). In P1, a total of 153,376 variants were identified. Assuming autosomal recessive inheritance, and upon filtering for homozygous, rare, nonsynonymous changes in coding regions and splice sites, 38 variants were considered. Among these, functional annotation identified the PAX1 NM_006192.3 c.463_465del variant, predicted to cause an in-frame deletion of asparagine at position 155 (p.Asn155del) of the PAX1 protein, as the most likely cause of the disease. In P2, 87,423 variants were detected. Assuming an autosomal recessive inheritance, and upon filtering for homozygous, nonsynonymous, and rare (minor allele frequency < 0.01) variants falling in coding regions or splice sites, 18 such variants were considered. Functional filtering of these revealed the PAX1 c.1104C>A variant, predicted to cause a premature termination at codon 368 (p.Cys368*), as the most likely cause of the disease. In P4, 60,772 variants were detected. Upon filtering for homozygous, nonsynonymous, rare (in-house Saudi variant database <0.005) variants, which were restricted to exonic or splice sites, contained in an autozygome region identified on chromosome 20 by high-density genotyping, and shared with P5 and P6, only two variants were identified, including the PAX1 c.439G>C variant, predicted to cause a p.Val147Leu amino acid change.

Sanger sequencing confirmed homozygosity for the suspected pathogenic PAX1 variants in P1 to P6 (Fig. 1A). The Val147 and the Asn155 amino acid residues are in the DNA-binding paired box domain, and the Cys368 residue is in the transactivation domain of the PAX1 protein (Fig. 1B). All these positions are evolutionarily conserved (fig. S5). The scaled CADD (combined annotation dependent depletion) score (CADD-Phred) for the p.Val147Leu, p.Asn155del, and p.Cys368* variants is 28.1, 21.2, and 38, respectively, significantly higher than the mutation significance cutoff (MSC) score (9), which for the PAX1 gene is 12.06. Together, these data strongly support a pathogenic role of the PAX1 variants identified. Of note, while molecular and cellular studies to confirm the pathogenic role of the PAX1 variants were under way, another group independently attempted WES in P3 and in other family members (but not in P2) and reported the occurrence of the p.Cys368* variant in P3 (8).

To examine the effects of the PAX1 variants at the protein level, we transfected 293T cells with plasmids encoding for either wild-type (WT) or mutant PAX1 complementary DNA (cDNA) and analyzed protein expression by Western blot. In this assay, we also included the PAX1 p.Gly166Val variant, which had been previously reported in a patient with OTFCS2 (6). As shown in Fig. 2A, all mutant proteins were expressed at similar levels as WT PAX1, with the p.Cys368* mutant migrating as a lower molecular weight product, as predicted. To check whether the identified variants altered the subcellular localization of the PAX1 protein, 293T cells were transfected with PAX1 constructs with an N-terminal HA tag, and immunofluorescence was performed with tetramethyl rhodamine isothiocyanate (TRITC)conjugated anti-HA antibody. As shown in Fig. 2B, both WT and mutant PAX proteins were detected in the nucleus, indicating that these variants do not affect subcellular localization.

(A) Western blot showing expression of WT and mutant human PAX1 proteins upon transient transfection in 293T cells. (B) Left: Intracellular protein localization upon transfection of HA-tagged WT and mutant PAX1 constructs into 293T cells, followed by staining with TRITC anti-HA. Right: Counterstaining with DAPI, demonstrating that the mutant PAX1 protein retains nuclear translocation capacity. Scale bar, 10 m. (C) Results of a luciferase reporter assay demonstrating reduced transcriptional activity of mutant PAX1 proteins, corresponding to the PAX1 variants detected in patients. The promoter region of Nkx3-2 was used to drive luciferase expression. Results of six independent experiments (each run in triplicate) are shown (means SEM). P value was calculated with one-way ANOVA and adjusted by Dunnetts multiple comparisons test. **P < 0.01; ***P < 0.0001.

Next, we tested the transcriptional activity of the PAX1 mutant proteins. Little is known on transcriptional targets of human PAX1; however, the Nkx3-2 promoter has been identified as a PAX1 target in mice (10). Therefore, we generated a reporter system in which luciferase expression is driven by the mouse Nkx3-2 promoter. In parallel, we generated both WT (Pax1WT) and mutant (Pax1Val138Leu, Pax1Asn146del, Pax1Cys359*, and Pax1Gly157Val) N-terminal HA-tagged mouse Pax1 constructs, which encode for mouse mutant PAX1 proteins corresponding to the human p.Val147Leu, p.Asn155del, p.Cys368*, and p.Gly166Val variants, respectively. Western blot analysis confirmed that the mutant mouse PAX1 proteins were expressed at similar levels as WT PAX1 (fig. S6). Upon cotransfection of the Nkx3.2-luciferase reporter plasmid and of either WT or mutant PAX1 expression plasmids into 293T cells, analysis of luciferase activity showed that the p.Val138Leu, p.Asn146del, and p.Cys359* PAX1 mutant proteins had significantly reduced reporter expression when compared with WT PAX1 (Fig. 2C and data file S1). A similar defect was also observed for the p.Gly157Val mutant, confirming previous findings (6). These data suggest that the human p.Val147Leu, p.Asn155del, and p.Cys368* variants do not affect protein stability or subcellular localization but alter PAX1 transcriptional activity.

The structure of the human PAX1 protein has not been solved experimentally. However, a crystal structure is available for the paired box domain of the highly homologous PAX6 protein (11). Sequence alignment between the paired box domain of PAX6 and PAX1 proteins reveals a high level of conservation with a similarity of 71%, with a 100% coverage of the region to be modeled as calculated with the BLOSUM80 matrix from PSI-BLAST (E = 1.3691 1020). As reported by Kelm et al. (12), this degree of homology often yields a model for the target (PAX1) with an accuracy of less than 1 root mean square deviation (RMSD) of atomic mobility to the experimentally solved structure of the template (PAX6). Because the p.Val147Leu and p.Asn155del mutants fall within the paired box domain of the protein, we assessed whether the reduced functional activity of the mouse p.Val138Leu and p.Asn146del (and by inference, the human p.Val147Leu and p.Asn155del) variants results from an altered structure and/or abnormal DNA binding. To do this, we first developed a structural model of the paired box domain of WT and mutant PAX1 bound to DNA, based on its homology to the published crystal structure of PAX6 [Protein Data Bank (PDB): 6PAX] (11) by the satisfaction of spatial restraints method using Modeler (13). Structural alignment revealed that the paired box domains of the PAX1 and PAX6 proteins are almost identical with a template modeling (TM) score of 0.99963 and RMSD of 0.08 as measured by the TM align algorithm (14). In addition, the high quality of the model is reflected by the fact that 99% of the residues are in the allowed regions of the (phi) versus (psi) angles of the Ramachandran plot, as shown in fig. S7 (15). Therefore, we used this model to derive a corresponding model for the p.Val147Leu and p.Asn155del variants and for the previously described p.Gly166Val PAX1 variant (6), using in silico site-directed mutagenesis and energy minimization refinement as previously described (16). As shown in Fig. 3A, the paired box domain of all three mutant PAX1 proteins retains a structure composed of two globular domains separated by a linker. These structural models were then used in MD simulations for both their free and DNA-bound forms to define how they differ in both structure and time-dependent dynamic behavior from the canonical WT PAX1 protein.

(A) Molecular modeling of the paired box domain of WT and mutant PAX1 proteins, showing the presence of two globular domain separated by a linker. Note that the asparagine residue at position 155 is adjacent to linker domain, and its deletion results in shortening of the last turn of the third helix in the first globular domain of the paired box domain. (B) Molecular superimposition of WT (in light blue) and mutant PAX1 variants after MD simulation, showing that both the Val147Leu and Asn155del variants predominantly affect the conformation of the C-terminal globular domain, whereas both globular domains are affected by the Gly166Val variant. (C) RMSF values of WT PAX1 and of the Val147Leu, Asn155del, and Gly166Val variants during MD simulations. RMSF values are used here as a measure of the flexibility of different regions of the protein during the MD simulations. The Y axes indicate the magnitude of the fluctuation, whereas the X axes indicate the specific location of each amino acid within the paired box domain.

Because the p.Val147Leu variant is located in the first globular domain, the p.Asn155del is also located in this domain and adjacent to the highly flexible linker, and the p.Gly166Val variant is within the linker, we initially performed 200-ps MD simulations of PAX1 in the absence of DNA to capture potential alterations of the rapid movement of this region of the protein in relationship to the N- and C-terminal helix-loop-helix domains. To gain additional insights into the behavior of the protein, we extended these simulations to 10 ns, in the absence or presence of DNA. When a harmonic restraint is applied to reduce the conformational changes in both globular domains during the 200-ps simulation, the linker is observed to move freely. In this situation, the molecular movement of WT PAX1 paired box domain resembles a barbell-shaped harmonic oscillator, where the globular domains move relative to each other without forming bonds that lock them together in space.

At the end of the 200 ps, in the absence of DNA, the linker of PAX1 shortens and the protein populates a conformational landscape where the globular domains come in close proximity to each other, with the linker located between the N-terminal helix 3 (H3) and the C-terminal helix 1 (H1), respectively (fig. S8). In the most extended conformation of the linker, the interglobular domain distance measured from the Gly158 -C to the Pro175 -C shortens from an original 38.946 to 21.414 (SD = 2.421, P = 0.0001). This shortening contributes to the differences in the RMSD curve, where in the first part of the simulation we observed significant changes due to this shortening, whereas the difference in conformational sampling decreases toward the end of the run. Identical results were obtained in 10-ns simulations. Thus, this H3-Linker-H1 state is likely the one that the PAX1 binding domain adopts when in conformational equilibrium before binding to DNA. In this manner, the linker would be free to contact the minor groove of the DNA and extend in a manner that allows the positioning of both globular domains for full binding. These results led us to set up simulations that would enable gathering information on potential differences in DNA binding among the WT and mutant PAX1 variants.

To investigate whether alterations in the structure or the dynamics of the PAX1 variants have the potential to affect the protein function as a transcription factors, we modeled these proteins in complex with DNA. For this purpose, we again used the bound form of PAX6 as a template. Figure S9 shows the energy-minimized structure of these models before MD simulations. Because the variants identified in the patients either change the sequence of the linker (p.Gly166Val) or the N-terminal globular domain (p.Val147Leu and p.Asn155del), we compared the structures of these variants with WT PAX1 after MD simulation. Because the structure of the DNA interacting with WT or mutant PAX1 proteins was the same in all models shown in fig. S9, we removed it to facilitate the observation of changes that occur in the PAX1 polypeptide chain. When compared with WT PAX1, the p.Val147Leu and the p.Asn155del variants associated with OTFCS2 + SCID differ in particular at the C-terminal second globular domain, as shown by molecular superimposition (Fig. 3B). This result is consistent with the measured root mean square fluctuation (RMSF) values, which shows that the second globular domain is highly flexible in the p.Val147Leu and p.Asn155del mutant proteins (Fig. 3C). By contrast, RMSF values in the first globular domain were lower in all mutant proteins (and especially so in the p.Asn155del and p.Gly166Val mutants) as compared with WT PAX1. Considering these changes, we evaluated potential alterations in the ability of these proteins to recognize and bind to DNA in silico. For this purpose, we analyzed the PAX1-DNA interface. As shown in Fig. 4, as compared with WT PAX1, a lower number of amino acid residues contacting DNA were present within the paired box domain of the p.Val147Leu and p.Asn155del PAX1 mutants. These alterations are more pronounced for the C-terminal region of the domain, which contacts the 3 half of the oligonucleotide and is necessary to maintain appropriate binding to DNA. This altered pattern of interaction with DNA observed in silico may contribute to the altered transcriptional activity of the PAX1 mutant proteins.

Nucleotide residues, in which the paired box domain of either WT or PAX1 mutant proteins establishes interaction, are shown in black. The amino acids contacting nucleotides of target DNA are indicated on the Y axis for each PAX1 protein. The red and green colors indicate loss and gain of DNA binding, respectively.

To gain insights into how pathogenic PAX1 variants may perturb the developmental program of thymic epithelial cells (TECs), we reprogrammed fibroblasts from a healthy control, P1, and P4 to iPSCs and subsequently differentiated these to TEP cells using a previously published protocol (17) with some modifications (see Materials and Methods). Quantitative real-time polymerase chain reaction (qRT-PCR) showed a comparable stemness profile in both control and patient iPSCs (fig. S10), and cytogenetic analysis confirmed their karyotypic integrity. iPSCs were then exposed in vitro to a cocktail of growth factors and molecules that provide essential cues to allow differentiation into definitive endoderm (DE) and eventually into TEP cells (fig. S11A).

To assess changes in the gene expression profile of cells during differentiation, we performed RNA sequencing (RNA-seq) in control cells collected in triplicate at iPS [day 0 (d0)], DE (d5), and TEP (d14) stages of cell differentiation. For each condition, between 15 and 20 million reads were obtained per well. As shown in fig. S11B, during differentiation of control iPSCs to DE and TEPs, we observed progressive changes of gene expression profile, with increased expression of stemness (OCT4, MYC, SOX2, TERT, DNMT3B, and NANOG), endoderm (EOMES, CXCR4, and SOX17), and epithelial (KRT8, CLDN1, EPCAM, LAMA1, and KRT19) genes at iPS, DE, and TEP stages, respectively. In addition, expression of ASXL1, HES1, SHH, GATA3, HOXA3, PSEN1, ZBTB1, HAND2, and MAFB genes, which are all part of the gene set Thymus development, was up-regulated at TEP stage (fig. S11B). Gene set enrichment analysis (GSEA) confirmed differential expression of genes involved in somatic cell maintenance and endoderm development, as well as in other pathways related to differentiation of tissues derived from the third and fourth pharyngeal pouches (fig. S11C).

To assess the reproducibility of the differentiation protocol, we differentiated the same control iPS line twice to TEP cells (named C1 and C2, respectively) in parallel to differentiation of P1 and P4 iPSCs to TEP cells in two distinct differentiation experiments. As shown in Fig. 5A, a similar pattern of changes in the gene expression profile was observed when differentiating control (C1) and P1 iPSCs or control (C2) and P4 iPSCs to TEP cells. In both experiments, control and patient cells showed increased expression of stemness genes at the iPS stage, whereas enhanced expression of epithelial marker genes and of other genes included in the Thymus development gene set was detected at TEP stage. Furthermore, immunohistochemistry analysis confirmed that both control and P1 TEP cells expressed cytokeratin 8 (KRT8), a marker of TECs (fig. S12) (18).

(A) Heatmap of differentially expressed genes between iPS and TEP stage as determined by RNA-seq. Each heatmap shows the top 3000 genes, which were differentially expressed between iPS and TEP cells, with a significance (q < 0.01) by the two-group comparison (t test). Genes whose expression was found to be up-regulated at the TEP stage included epithelial cell markers (EPCAM, KRT8, and KRT19) as well as several genes (PSEN1, HES1, ASXL1, HOXA3, HAND2, EPHB3, and GATA3), which appeared at the leading edge of GSEA of thymus development in (B). (B and C) GSEA on thymus development gene set by preranked genes according to signed log10 adjusted P value. The adjusted P value was acquired by DEseq2 analysis using normalized read count of RNA seq data. FDR, false discovery rate. (D) qRT-PCR analysis of FOXN1 and DLL4 expression at TEP stage of differentiation. Results are from five independent experiments for control and P1, and four independent experiments for control and P4, with triplicates in each case (mean SEM). The P value was calculated with two-tailed paired t test. P < 0.05 was considered to be significant. (E) Thymus development genes with evidence of differential expression between patient and control cells (adjusted P < 0.1 and concordant pattern of expression in both RNA-seq experiments). For this comparison, we considered genes that were part of the Thymus development gene set in MSigDB v7.0, and in the top 30 FOXN1 target genes reported in (19). The values displayed are the signed log10 adjusted P value for differential expression.

GSEA confirmed that upon differentiation of control iPSCs to TEP cells, genes involved in thymus development were more abundantly expressed at the TEP stage both in control and in PAX1 mutant cells (Fig. 5B). Despite similar changes in gene expression profile during differentiation of control- and patient-derived iPSCs to TEP cells, GSEA demonstrated that genes involved in thymus development were more abundantly expressed in control than in patient TEP cells (Fig. 5C). To gain additional mechanistic insights into the severe T cell immunodeficiency of P1 and P4, we performed multiple rounds of differentiation of control and patient iPSCs to TEP cells (five times for control and P1 and four times for control and P4 cells, respectively) and used qRT-PCR to analyze the expression of FOXN1, a master regulator of TEC development (19, 20), and to its target DLL4, a Notch ligand that plays a critical role in T cell commitment (21). FOXN1 expression was significantly reduced in P1 and P4 TEPs as compared with control cells, and a similar trend was observed for DLL4, although the latter significance was reached only when comparing P1 with control TEPs (Fig. 5D and data file S1). Analysis of RNA-seq data revealed several other genes that showed concordantly reduced expression in P1 and P4 TEPs versus control TEPs, reaching statistical significance in at least one of the patients TEP lines (Fig. 5E and table S2). These included STC2, CD83, ZAR1, and ANKMY1, which are known FOXN1 target genes (19); TP63, a regulator of TEC proliferation and aging (22, 23); BMP4, which has been implied in thymus development (24, 25) and in maintenance of TEPs (26, 27); and EYA1 and PAX9, which are involved in patterning of pharyngeal endoderm (28, 29). Together, these data indicate that multiple mechanisms contribute to the thymic defects associated with PAX1 deficiency. Consistent with this, and with the syndromic features manifested by the patients, we observed that several genes included in the Neural crest cell differentiation, Ear development, Cartilage development, Pharyngeal system development, and Skeletal system development gene sets also manifested differential expression in P1 and P4 versus control TEPs (fig. S13).

We have studied six patients from three unrelated families in whom biallelic, loss-of-function PAX1 variants underlie a clinical phenotype characterized by OTFCS2 and severe T cell immunodeficiency. The first example of a biallelic, rare PAX1 variant (p.Gly166Val) in a patient with autosomal recessive OTFCS2 was provided by Pohl et al. (6), who also showed reduced transcriptional activity of the mutant PAX1 protein. However, no data on the patients immunological phenotype were provided. More recently, Patil et al. (7) have described two siblings with a homozygous frameshift PAX1 variant causing OTFCS2; one of them lacked a thymic shadow on chest x-ray. Last, the clinical features of OTFCS2 and SCID have been recently reported by Paganini et al. (8) in two of the patients studied here (P2 and P3), but no immunological or mechanistic characterization was provided.

Several mouse models of PAX1 deficiency, due to distinct variants in the Pax1 gene, have been described, including the undulated (un), undulated extensive (unex), undulated short-tail (unS), and undulated intermediate (un-i) models (30). All of these mutant strains display thymic abnormalities, which are more severe in the unS model (30); however, none of them results in complete athymia. A more profound phenotype, with lack of thymus and parathyroids, associated with craniofacial and skeletal abnormalities, has been observed in Pax9/ mice (31). No cases have been reported of humans with biallelic PAX9 pathogenic variants, and heterozygous PAX9 variants in humans are associated with hypodontia but not with thymic defects (32). Together, these data suggest that the impact of PAX1 and PAX9 on thymus development may be different in humans and mice.

To gain insights into the molecular mechanisms by which PAX1 deficiency may cause syndromic SCID in humans, we have first investigated the expression, subcellular localization, and transactivation activity of PAX1 mutant proteins using transient transfection and luciferase reporter studies. Although transient transfection may result in protein overexpression and therefore cannot be directly compared with protein expression in vivo, the PAX1 p.Val147Leu, p.Asn155del, and p.Cys368* mutant proteins retained the capacity to translocate to the nucleus, and the equivalent murine mutant proteins showed decreased transcription factor activity in vitro. Similar results were obtained for the PAX1 p.Gly166Val (and the mouse equivalent p.Gly157Val) variants, confirming previous observations (6). To further investigate the mechanisms underlying the impaired transcriptional activity of the mutant PAX1 proteins, we have performed structural modeling, using the crystal structure of the PAX6 paired box domain as a template. The results suggest that the structural behavior of the paired box domain (consisting of two globular domains interconnected by a linker) was retained in the p.Val147Leu, p.Asn155del, and p.Gly166Val mutants. MD simulation studies have demonstrated that these variants alter the flexibility of the paired box domain and are predicted to alter binding of PAX1 to its target DNA. Our in silico studies suggest that the mutants differ in their ability to gain or lose binding to distinct nucleotides, with possible impact on the severity of clinical and immunological phenotype. Fine characterization of the molecular mechanisms underlying such heterogeneity will require resolution of the crystal structure of the PAX1 paired box domain and precise identification of its human DNA target sequence(s).

By exposing control- and patient-derived iPSCs to defined differentiation cues, we have successfully differentiated iPSCs to TEPs. Comparison of gene expression profile in control- and patient-derived cells at the TEP stage of in vitro differentiation demonstrated altered expression of genes involved in thymus development in patient cells. In particular, qRT-PCR analysis revealed reduced expression of FOXN1, a master gene of thymus development, and of several FOXN1 target genes, including DLL4. Biallelic FOXN1 pathogenic variants in humans are responsible for a syndromic form of SCID that is the equivalent to what is observed in the nude mouse (33, 34). We have recently reported that FOXN1 haploinsufficiency in humans causes severe T cell lymphopenia at birth (35). The reduced levels of FOXN1 expression observed in patient TEPs (and, by inference, in the patients thymus) may therefore play a direct role in the severe T cell lymphopenia observed in these patients. However, analysis of gene expression profile in patient and control TEPs suggests that other mechanisms, besides reduced FOXN1 expression, may also contribute to impaired thymic development associated with PAX1 deficiency. In particular, reduced TP63 expression may cause impaired TEC proliferation and hence thymic hypoplasia. Moreover, we observed that both P1 and P4 TEPs displayed significantly reduced expression of BMP4 as compared with control TEPs. Conditional deletion of Bmp4 from the pharyngeal endoderm before Foxn1 expression disrupts thymus morphogenesis in mice (24). Furthermore, recent studies have indicated that BMP4 plays a critical role in maintenance of TEC progenitors (27), and reduced BMP4 expression might alter replenishment of the TEC compartment. Future studies based on precise enumeration of TEPs generated in vitro from patient- and control-derived iPSCs may help test this hypothesis. In any case, these data suggest that PAX1 deficiency causes early and more global effects on the development of tissues derived from the third and fourth pharyngeal pouches, including the thymus. Consistent with this hypothesis, patient TEPs were concordant in the abnormal expression of a number of genes involved in skeletal, cartilage, pharyngeal, neural crest, and ear development. Abnormalities in these pathways during differentiation of tissues derived from the third and fourth pharyngeal pouches are likely to contribute to the broad range of malformations observed in the patients reported here.

Last, we have reported that HSCT, which was attempted in four of the six patients, failed to correct the T cell immunodeficiency, despite engraftment in three of them. PAX1 deficiency should be added to the list of severe T cell immunodeficiencies characterized by a primary thymic defect, which also includes complete DiGeorge syndrome, CHARGE syndrome, and FOXN1 deficiency (1). Thymus transplantation represents the treatment of choice to correct the immunodeficiency in these disorders (3638). By contrast, use of unmanipulated HSCT may allow engraftment of donor-derived postthymic T cells that may expand in the recipient, as also observed in P1 in this study, but does not permit de novo generation of a polyclonal repertoire of nave T cells (39). In summary, we have provided mechanistic insights into the pathophysiology of OTFCS2 associated with severe T cell immunodeficiency, an autosomal recessive condition caused by PAX1 variants, and have demonstrated the thymic-intrinsic nature of the immunodeficiency of this condition.

The scope of the study was to identify the molecular basis of a syndromic form of SCID and to perform genomic, molecular, biochemical, structural modeling, and in vitro disease modeling studies to analyze deleterious effects of the PAX1 variants identified. All patients provided written informed consent, according to protocols approved by the local Institutional Review Boards (IRBs). Research studies were performed under National Institutes of Health (NIH) IRB-approved protocol 16-I-N139. For P4, public disclosure of secondary genomic findings was not permitted by the protocol and consent form approved by the local IRB.

WES was performed on P1 and his healthy parents and on P2 and P4 without parental samples. Detailed methods for capture, library preparation, and bioinformatic analysis are described in the Supplementary Materials. Candidate variants were confirmed by Sanger sequencing and described according to Human Genome Variation Society (HGVS) guidelines. For P1 and P2, WES data have been deposited to the National Center for Biotechnology Information (NCBI) Sequence Read Archive (SRA) Submission Portal, with the following ID: PRJNA601119.

Flow cytometry studies were performed on either a 10-color Gallios (Beckman Coulter, Brea, CA) or an 8-color Canto II (BD Biosciences, San Jose, CA) cytometer, and results were analyzed using Kaluza software v1.5 (Beckman Coulter, Brea, CA). T cell proliferation studies were performed using Edu-based (Thermo Fisher Scientific, Waltham, MA) flow cytometry method in P1, and tritiated thymidine (3HTdR) incorporation in P2, P4, P5, and P6. TCR V repertoire spectratyping was carried out using a fragment length method on a capillary electrophoresis system (ABI 3730xl DNA Sequencer, Applied Biosystems Inc., Thermo Fisher, Waltham, MA), and data were analyzed using the GeneMarker (v.2.4.0) software (SoftGenetics, State College, PA). All reference values for interpretation were established in the laboratory using healthy pediatric donors recruited via an IRB-approved protocol.

293T cells were plated as 4 105 cells per well in a 12-well plate. After 24 hours, cells were transfected with 1.2 g of pCMV-HA-N vector containing either WT or mutant PAX1 cDNAs, with the Lipofectamine 3000 transfection kit (Thermo Fisher Scientific) following the manufacturers instructions. After 24 hours, cells were collected, lysed, and transferred onto a nitrocellulose membrane. Immunoblotting was performed with rat anti-PAX1/Pax1 monoclonal antibody (mAb) (clone 5A2) (40), followed by staining with horseradish peroxidase (HRP)conjugated goat anti-rat IgG (ab97057; Abcam, Cambridge, MA). After stripping, the membrane was reblotted with rabbit anti-actin mAb (clone 13E5; Cell Signaling Technology, Danvers, MA), followed by Amersham enhanced chemiluminescence anti-rabbit IgG, HRP-linked whole antibody (NA934; GE Healthcare, Helsinki, Finland).

To analyze PAX1 subcellular localization, 293T cells were cultured in polylysine-coated -Slide 8 well (ibidi, Fitchburg, WI) and transfected with 100 ng of pCMV-HA-N vector containing either WT or mutant PAX1 cDNA, with the Lipofectamine 3000 transfection kit (Thermo Fisher Scientific) following the manufacturers instructions. After 24 hours, cells were fixed in 4% paraformaldehyde with phosphate-buffered saline (PBS) for 30 min at room temperature, washed twice in PBS, and then blocked for 1 hour with 10% donkey serum and 0.1% Triton X-100 with PBS at room temperature. Cells were incubated with mouse anti-HA-TRITC mAb (clone H9037; MilliporeSigma, St. Louis) diluted 1:200 in PBS and with 4,6-diamidino-2-phenylindole (DAPI) at room temperature for 1 hour in the dark. Images were obtained with a Leica SP8 (690/730) confocal microscope.

For immunofluorescence analysis of KRT8 expression by TEPs, cells were fixed in 4% paraformaldehyde with PBS for 30 min at room temperature, washed twice in PBS, blocked for 1 hour in 10% donkey serum and 0.1% Triton X-100 with PBS at room temperature, and incubated overnight at 4C with mouse anti-KRT8 antibody (ab2530, C-43) (Abcam, Cambridge, MA) diluted 1:200 in PBS, then for 1 hour at room temperature in the dark with donkey anti-mouse IgG (H+L) Alexa Fluor 488 (ab150105; Abcam) at 1:500 dilution in PBS, and with DAPI (Thermo Fisher Scientific) at 1:1000 dilution in PBS. Images were taken with a Leica SP8 (690/730) confocal microscope.

The promoter region of the mouse Nkx3-2 gene was amplified and cloned into the firefly reporter plasmid pGL4.10 luc2 vector (Promega, Madison, WI), as described (6, 10). To generate expression plasmids containing the mouse Pax1WT, Pax1V138L, Pax1N146del, Pax1G157V, and Pax1C359* coding sequences, the coding sequence of mouse Pax1 (NM_008780.2) was amplified by RT-PCR from isolated adult mouse thymus RNA and cloned into a pCMV-HA-N vector (Addgene, Cambridge, MA) with the In-Fusion HD EcoDry Cloning Kit (Clontech, Mountain View, CA). Pax1 mutant variants were generated by site-directed mutagenesis, and the PCR products were ligated with the Quick Ligation Kit (NEB, Ipswich, MA) and cloned by Turbo competent cells (NEB, Ipswich, MA). The correct sequence of the constructs was confirmed by Sanger sequencing.

The transcriptional activity of WT and mutant PAX1 mouse proteins was assessed in a luciferase reporter assay. 293T cells were cultured in Dulbeccos modified Eagles medium (DMEM) containing 10% fetal bovine serum with antibiotics and plated in 24-well plates 24 hours before transfection. Transient transfections were performed in triplicate with TransIT-293 Transfection Reagent (Mirus, Madison, WI) according to the manufacturers instructions. Cells were cotransfected with 30 ng of either WT or mutant Pax1 expression plasmids, 15 ng of firefly reporter plasmid Nkx3-2-pGL4.10 luc2, and 3 ng of pRL-TK vector (Promega, Madison, WI) for normalization. After 48 hours, cell extracts were collected and frozen in lysis buffer overnight at 20C. After thawing, firefly and renilla luciferase activities were measured using a Dual-Luciferase Reporter Assay Kit (Promega, Madison, WI) and Paradigm Detection platform (Beckman Coulter, Indianapolis, IN). To correct for variations in transfection efficiency, firefly luciferase activity was normalized to renilla luciferase activity. The luciferase activity of pCMV-HA-N vector, which had no Pax1 cDNA, was assumed to have 0% activity, whereas the Pax1WT vector was assumed to have 100% activity.

The three-dimensional complex structures of WT and mutant PAX1 models bound to DNA were generated by homology-based methods (16) using the previously solved structure of the highly homologous protein, PAX6 (PDB: 6PAX) (11). Intermolecular interactions of the PAX1 paired box domain of WT/mutant PAX1 to DNA complex were calculated in the Receptor-Ligand function of Discovery Studio Client 4.0 using the default parameters (BIOVIA, San Diego, CA). The MD simulations were performed as described (16).

Primary skin fibroblasts from P1, P4, and a healthy control (BJ fibroblast line, American Type Culture Collection) were reprogrammed to iPSCs by infection with the nonintegrating CytoTune Sendai viral vector kit (Thermo Fisher Scientific) as described (41).

For differentiation, iPSCs were transferred to plates coated with Corning Matrigel human embryonic stem cell (hESC)qualified Matrix. After four to five passages, the cells were plated on Matrigel-coated 24-well plates at a density of 2.5 105 cells/cm2. For differentiation to DE and TEPs, iPSCs were exposed to various factors and differentiation cues, according to the protocol by Parent et al. (17), with some modifications. In particular, between d1 and d5, iPSC differentiation was carried out in RPMI 1640 medium (Thermo Fisher Scientific, Waltham, MA) supplemented with 1% penicillin/streptomycin, 1% l-glutamine, and increasing concentrations of KSR (0% on d1, 0.2% on d2 and d3, and 2% on d4 and d5). In the period d6 to d14, cells were differentiated in DMEM/F12 with 1% penicillin/streptomycin, 1% l-glutamine, and 0.5% (v/v) B-27 supplement (Thermo Fisher Scientific, Waltham, MA). During this period of time, the following factors were added to the culture: activin A, 100 ng/ml (d1 to d5); Wnt3a, 25 ng/ml (d1) or 50 ng/ml (d8 to d14); all-trans retinoic acid (RA), 0.25 M (d6 to d8) or 0.1 M (d9 to d14); BMP4, 50 ng/ml (d6 to d14); LY364947, 5 mM (d6 to d9); FGF8b, 50 ng/ml (d8 to d14); and KAAD-cyclopamine, 0.5 mM (d8 to d14). Supplements and factors were from Thermo Fisher Scientific, Waltham, MA (B27, KSR); R&D Systems, Minneapolis, MN (activin A, Wnt3a, BMP4, and FGF8b); and MilliporeSigma, St. Louis, MO (RA, KAAD-cyclopamine, LY364947).

Microgram quantities of total RNA were isolated using the RNeasy Kit (QIAGEN, Hilden, Germany) from triplicate samples of control-, P1-, and P4-derived iPSCs, as well as from the corresponding iPSC-derived cells at DE and TEP stages. RNA integrity was tested by microfluidic electrophoresis on a TapeStation system (Agilent, Santa Clara, CA). RNA purity and concentration were assessed using the NanoDrop One UV-Vis Spectrophotometer (Thermo Fischer Scientific, Waltham, MA). Directional, mRNA-seq libraries for experiment 1 were produced using TruSeq Stranded mRNA Library Prep Kit for NeoPrep (catalog no. NP-202-1001) from Illumina (San Diego, CA). Directional, mRNA-seq libraries for experiment 2 were produced using New England Biolabs product NEBNext Poly(A) mRNA Magnetic Isolation Module (catalog no. E7490L), New England Biolabs product NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (catalog no. E7760L), and NEBNext Multiplex Oligos for Illumina (Dual Index Primers Set 1) (catalog no. E7600S) (New England Biolabs, Ipswich MA), with an input of 100 ng of total RNA per sample.

Sequencing was performed on an Illumina NextSeq 500 system, running Illumina NextSeq Control Software System Suite version 2.1.2 and RTA version 2.4.11. The final library pool was sequenced via 1 75base pair (bp) run configuration using the product NextSeq 500/550 High Output v2 sequencing kit, 75 cycles (catalog no. FC-404-2005). Between 15 106 and 20 106 reads were obtained from each sample. RNA-seq FASTQ files were aligned to the reference human genome assembly (GRCh38) with STAR v2.6.0 (42). The transcript annotation (GTF) file was obtained from GENCODE (release 28) (43). The binary alignment files (.bam) were then used to generate a matrix of read counts with the featureCounts program of the package Subread v.1.6.2 (44). Exonic fragments were grouped at the level of genes, based on the GENCODE 28 annotation file. Normalization and differential expression analysis for RNA-seq data were performed with the DESeq2 (45) package in R (46).

Independent pairwise analyses were performed on triplicate samples of cells at each stage of differentiation (iPSC, DE, and TEP). To handle the lower power associated with small numbers of samples, DESeq2 uses an empirical Bayesian procedure to stabilize the log fold change estimates. The Wald test was then applied to the log fold change in each gene, followed by multiple-testing adjustment with the method of Benjamini and Hochberg (47).

For the heatmap of gene expression, t test and hierarchical clustering were computed by Qlucore Omics Explorer 3.3 (Qlucore, Lund, Sweden) for iPSC and TEP stage comparison (Fig. 5A), with cutoff q values of less than 0.01. Analysis of variance (ANOVA) and hierarchical clustering were used for the three-stage (iPSC, DE, and TEP) comparison (fig. S11B). Normalization and differential expression analysis of the RNA-seq data used for GSEA were performed with DESeq2 package in R v.3.5.1. RNA-seq data have been uploaded to the NCBI Gene Expression Omnibus (GEO), under accession no. GSE138784.

GSEA was performed with the GSEA software (48) (http://www.broadinstitute.org/gsea) using a preranked dataset of gene expression differences, 1000 permutations, and the softwares classic enrichment statistic option. Genes were ranked based on the DESeq2 output by taking the signed log10 adjusted P value for differential expression. Gene sets for enrichment analysis correspond to Gene Ontology (GO) Biological Processes and were obtained from the Molecular Signatures Database version 7.0 (GMT file: c5.bp.v7.0.symbols.gmt).

RNA was isolated from control, P1, and P4 cells at iPSC and TEP stages of differentiation, using RNeasy kit (QIAGEN, Hilden, Germany). cDNA was synthesized by a qScript cDNA Synthesis kit (Quantabio, Beverly, MA) according to the manufacturers protocol. qRT-PCR was performed on a 7500 RT-PCR system (Applied Biosystems, Waltham, MA) using PerfeCTa SYBR Green FastMix, Low ROX (Quantabio, Beverly, MA). Gene expression was quantified by normalization to the housekeeping gene TBP for each sample. Primers used for individual genes are reported in the Supplementary Materials.

Statistical analysis was undertaken in GraphPad Prism (v8.0). For luciferase reporter assay, P values were calculated with one-way ANOVA and adjusted by Dunnetts multiple comparisons test. The data are means SEM of six independent experiments (WT, n = 6; Val138Leu, n = 3; Asn146del, n = 5; Cys359*, n = 5; Gly157Val, n = 5; empty, n = 6). For qRT-PCR data, Students t test (paired, two-tailed) was performed. The data are means SEM in Fig. 5D, and means SD in fig. S10. P < 0.05 was considered to be significant. Statistical analysis of RNA-seq data is described above.

Acknowledgments: We thank E. Thorland for interpretive assistance with the CNV analysis and B. Bigio for uploading WES data. WES data have been deposited to the NCBI SRA Submission Portal, with the following ID: PRJNA601119. RNA-seq data have been uploaded to the NCBI GEO, under accession no. GSE138784. Funding: This work was supported by the Division of Intramural Research, National Institute of Allergy and Infectious Diseases (NIAID), NIH and by the Angelo Nocivelli Foundation. Y.Y. was supported by JSPS Research Fellowship for Japanese Biomedical and Behavioral Research at the NIH and had travel support from The ITO Foundation for the Promotion of Medical Science. R.U. was supported by NIH/NIDDK R01 DK52913, Advancing a Healthier Wisconsin (AHW) Endowment and the Linda T. and Johm A. Mellowes Endowed Innovation and Discovery Fund. L.M.F. is funded by the Division of Intramural Research of the National Institute of Arthritis, Musculoskeletal and Skin Diseases, at the National Institutes of Health. A.A. is supported by King Abdulaziz City for Science and Technology. Author contributions: Y.Y. performed experiments and wrote the manuscript. R.U. performed structural modeling and MD simulation studies. L.M.F. supervised analysis of RNA-seq and GSEA data. F.O.-C., T.G.M., and S. Ganesan assisted with RNA-seq studies. S. Giliani and S.M. performed Sanger sequencing and Western blot analysis and analyzed WES data. K.Z., A.M.A., H.A., F.Z., C.A.V., and B.B. performed and analyzed WES. A.K.D. generated iPSCs. A.J., R.W.M., A.H.F., C.A., B.K.A.-S., and H.A.-M. provided clinical care and description of the patients. F.F. performed lymph node pathology. M.P.B., M.L.H., and C.M. performed and interpreted imaging studies. J.L.C. and R.S.A. contributed to supervision of the project and to writing of the manuscript. L.D.N. was responsible for the entire research project and wrote the manuscript. Competing interests: The authors declare that they have no competing interests. Data and materials availability: Fibroblast and iPSC lines from P1 and P4 are available upon request but are contingent upon approval of material transfer agreement by the NIAID, NIH. WES data have been uploaded to the NCBI SRA Submission Portal, with the following ID: PRJNA601119. The RNA-seq dataset for this study has been uploaded to the NCBI GEO, under accession no. GSE138784. The GEO accession includes links to the NCBI SRA database, from which the raw data will be accessible in FASTQ format, under accession no. SRP225226.

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Augustinus Bader The Rich Cream ($265)

In the nearly two years since its launch, Augustinus Baderssignature moisturizer, aptly named The Rich Cream, has seen moresuccess than perhaps any beauty product before it. Alexa Chung, Margot Robbie,and Ashley Graham are all fans, and Victoria Beckham loves the brand so muchthat she even collaborated with Bader to create a serum for her eponymousbeauty line. The Rich Cream, developed by Dr. Bader, a German stem cell andbiomedical scientist, is said to promises to reduce signs of aging and skindamage, leaving skin restored, regenerated, and glowing with health. The keyto the moisturizers magic is TFC-8, Dr. Baders proprietary Trigger Factor Complexthat is essentially a blend of more than 40 amino acids, vitamins, andsynthesized molecules. The complex helps activate the skins stem cells,pushing them into healing mode to promote everyday turnover and furthermoresoften the appearance of fine lines and wrinkles, even tone and texture, andhydrate the skin. Starting at $265 for a 50ml bottle, The Rich Cream is farfrom cheap, but its benefits are well worth the large price tag.

For a more affordable alternative, try the Josie Maran Whipped Argan Oil Face Butter($42). Among thelightweight moisturizers selling points is its use of argan oil, areplenishing ingredient also found in Augustinus Baders Rich Cream that helpsprotect skin from free radicals.

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Moisturizers Worth the Money - NewBeauty Magazine

Skin Stem Cells Comments Off on Moisturizers Worth the Money – NewBeauty Magazine | February 29th, 2020

When an animal, such as a dog or cat, is sick or unwell, its first natural instinct is to refuse food. Their body automatically cuts down appetite till they recover. They may drink water, but nothing else. This is bodys intelligence taking over to heal and recover.Intuitive eatingHumans arent any different. At least, intuitively. All of us are products of nature and we have been designed the same. When we call in sick, the first thing that happens is that our appetites shut down too. Our taste of mouth changes and our sensitivity to smell and taste is numbed. This is our bodys intelligence and subtle biofeedback trying to tell us that its time we give our body some rest.

This is fasting in a way and has been practised since the longest time across all religions. Fasting is inbuilt in us and is considered medicinal if you actually go back and read its history. It is our bodys uncomplicated way of cleaning and healing. Unfortunately, we have moved too far from nature and our natural instincts. We are constantly eating and munching even when sick, thereby compromising our own immunity and masking the inbuilt mechanism to heal.Take a break

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Dont feed sickness - Times of India

Skin Stem Cells Comments Off on Dont feed sickness – Times of India | February 29th, 2020

If youve ever felt overwhelmed by the thousands of beauty products on the market, we hear you. Thats why we spent hundreds of hours doing the leg work and finding the best ones for you! We read through scientific studies to understand how ingredients work, did market research to find the best and newest products on the market, and asked our most-trusted experts what products they live and swear by. We take beauty seriously, because we understand the importance a product can play in our lives.

Because sometimes a $5 cheap thrill is as good (or *gasp* better) than its pricey counterpart, we decided to throw in a drugstore and luxe pick for almost every item, and we interviewed experts to get their picks, too. Each product deserves a place on this list for its unique features, and we cant wait to share them with you. Youll also find standout stars that were so great that we had to feature themeven though they didnt fit into any one product category.

The HelloGiggles team poured our hearts, souls, and most importantly, brainpower into this project and we found the best of the best. We stand behind each and every one of these products so that you can find the ones that speak to you. Consider the Beauty Crush Awards your shopping list for the yearmade by your best friends for the people they love.

Put your best face forward with these top-of-the class moisturizers, serums, face masks, and more. We tested hundreds of skincare products to find the crme de la crme. And, because we know skin can mean a hell of a lot of things to a lot of different people, we made sure that every winner blessed the face of women and men with different skin concerns, textures, and tones.

Best Drugstore Cleanser: Honest Beauty Gel-to-Milk Cleanser, $12.99 (orig. $14.99), amazon.com

This cleanser is a godsend for those with sensitive skin because of both the affordable price and the rosewater-infused formula. (Studies show that rose extract has anti-inflammatory benefits, which tackles irritation, making it perfect for those with skin prone to irritation.) The product starts as a gel-like formula and, upon contact with water, turns into a milky liquid that gently and effectively cleanses skin.

Best Luxury Cleanser: SkinCeuticals Glycolic Renewal Cleanser Gel, $38, dermstore.com

If your skin is looking dull or is congested, consider using an exfoliating cleanser two to three times a week to decongest and reveal bright, glowy skin. This cleanser uses glycolic acid, a chemical exfoliant that helps turn over dead skin cells to treat concerns such as uneven skin tone, rough skin, and fine lines.

Best Expert-Recommended Cleanser: Differin Daily Refreshing Cleanser, $8.49, amazon.com

I am a big fan of Differin Gel .1% adapalene, [and] I love this new addition to the line, says Dendy Engelman, M.D., a board-certified dermatologist. When cleansing, you dont want to strip the skin of natural oils. This can cause your skin to go into oil production overdrive. This cleanser uses vitamin B and glycerin to gently cleanse; vitamin B to calm the skin and provide relief to inflammation and redness, and glycerin to prevent water loss so the skins natural oils can nourish and repair.

Best Drugstore Makeup Remover: Almay Micellar Biodegradable Makeup Wipes, $4.99 (orig. $5.99), amazon.com

By now we all know and love micellar water for how effectively it removes makeup and cleanses skin in just one step. For those who need a refresh, micelles work like tiny magnets to attract impurities off the skin. We love these makeup wipes because they allow us to easily remove our makeup (even the stubborn waterproof kind) at the end of the day without irritating even the most sensitive of skin. This product won five extra points during our testing period for its biodegradable material, which is made up of plant fibers and wood pulp, and it will completely compost in just three months.

Best Luxury Makeup Remover: First Aid Beauty Conditioning Eye Makeup Remover, $18, ulta.com

Eye makeup can be a fickle foe when it comes to taking it off at the end of the night. Too often leftover mascara clings to our lashes, eyeshadow lingers in the creases of our lids, and eyeliner finds its way to our inner eye areano thank you. This oil-free formula works to remove even waterproof makeup without leaving a trace or any greasy residue. To use, apply some liquid onto a cotton ball, place over the eye area for five seconds, then gently pull down and away. Bonus pointit uses soothing chamomile and is fragrance-free, so its suitable for sensitive skin, too.

Best Expert-Recommended Makeup Remover: Simple Water Boost Micellar Make-up Remover Eye Pads, $8.49, cvs.com

This budget-friendly product uses micellar technologies to remove even the most longwear makeup without irritating skin. Anyone who knows me can attest to the fact that I go all out with both eyeshadow and mascara every day, so when it comes to taking it off, I need something thats going to really workand requires the least amount of elbow grease on my part, says Lisa DeSantis, Deputy Beauty Director of Real Simple and Health magazines. These rounds are saturated with just enough of the hardworking micellar formula to remove even the most stubborn eye makeup.

Best Drugstore Toner: Burts Bees Micellar Toning Water With Rose Water, $8.99, target.com

If youre looking for a wallet-friendly toner that wont sting your skin (no skincare product ever should) or dry you out, this product is for you. This alcohol-free formula uses calming rose petal extract, hydrating glycerin, and tone-balancing lactic acid.

Best Luxury Toner: belif Aqua BombHydrating Toner, $28, sephora.com

Imagine a facial toner that feels like a cup of water for your skinthats this one. It combines hydrating ingredients like glycerin with soothing ones like oat extract and calendula for a pleasant experience thats both effective in removing makeup and debris and gentle on the skin.

Best Expert-Recommended Toner: Peter Thomas Roth Water Drench Hyaluronic Cloud Hydrating Toner Mist, $28, dermstore.com

This product uses hyaluronic acid to hydrate and instantly reduce the appearance of fine lines and wrinkles, says Deanne Robinson, M.D., a dermatologist based in Connecticut.

Best Drugstore Serum: Honest Beauty Vitamin C Radiance Serum, $27.99, amazon.com

Vitamin C is a powerful antioxidant that neutralizes free radicals, which come from the sun, pollution, stress, and smokingfactors that can ultimately damage the cells DNAand gives your skin a radiant boost. Regularly using a vitamin C skincare product can boost collagen production and reverse cell damage, which in turn helps skin look firmer and looking younger, and even helps fade and prevent brown spots or pigmentation.

Best Luxury Serum: La Mer Regenerating Serum, $365, sephora.com

Environmental factors such as pollution, free radicals, and harmful UV rays can take a real toll on your skin, making it show signs of aging sooner. This super-charged serum uses antioxidants, seaweed extract, and a new proprietary blend that helps boost the skins natural collagen to reduce the appearance of fine lines, pores, and redness. One HG tester says that after using it for a few weeks, shed never seen her skin look healthier: I feel like I can see a whole new layer of skin on my face.

Best Expert-Recommended Serum: La Roche-Posay Hyalu B5 Serum, $29.99, dermstore.com

Kavita Mariwalla, M.D., a board-certified dermatologist based in West Islip, NY, adores this budget-friendly pick: I love this product for a few reasons: 1) Price point. For the quality of ingredients in it, it is priced really well. 2) B5 is a great ingredient for all skin types and madecassoside is a hero ingredient that acts as an antioxidant but also skin soother. Combine that with its hyaluronic acid and youve got a serum that has great workhouse ingredients in it.

Best Drugstore Moisturizer: Olay Regenerist Whip Face Moisturizer SPF 40, $38.99, ulta.com

To protect your skin from the sun, opt for a face moisturizer with SPF to streamline your routine. Testers loved that this Olay pick looked smooth on skin and acted as the perfect primer for makeup.

Best Luxury Moisturizer: Drunk Elephant Lala RetroWhipped Cream Moisturizer, $60, sephora.com

Testers liked that this cream instantly melted into their skin and made it feel healthy, hydrated, and glowy. It uses a mix of oils, ceramides, and antioxidants, which together provide moisture and help skin retain that moisture. Its also formulated at an ideal pH level of 5.2, so it wont throw your skin off. Oh, and did we mention it has a fun pump? The pump dispenses the perfect-sized amount of moisturizer onto your skin without any of the mess.

Best Expert-Recommended Moisturizer: Cetaphil Ultra Healing Lotion with Ceramides, $19.97, amazon.com

Although this moisturizing lotion is technically formulated for the body, Tiffany Libby, M.D., a board-certified dermatologist in Rhode Island, recommends it for the face as well. I use this on my body and face when I need to double down on my moisture, especially in the winter months, and I love that it is formulated with ceramides to help enhance my skin barrier and keep my skin smooth and hydrated, she says.

Best Drugstore Face Oil: e.l.f. All The Feels Facial Oil, $11.99, target.com

Squalane, hemp seed oil, and rosehip seed oil make this lightweight facial oil the perfect last step in your skincare routine. Simply apply a few drops onto your skin after your moisturizer, then pat it in to encourage absorption and help lock in your skincare.

Best Luxury Face Oil: Snow Fox Herbal Youth Oil, $40, saksfifthavenue.com

This silky-feeling serum combines hydrating and anti-aging properties into one product. It uses blueberry seed oil, which studies show contains high levels of antioxidant properties that protect skin from free radicals, and an exotic herb called Brazilian Paracressextract, which is a muscle relaxant that helps relieve micro-tension. HG testers raved that it gave their skin a healthy glow, one of them even saying it made her skin look like there was a ring light shining on her face at all times. Natural-looking radiance with no filterwere here for it.

Best Expert-Recommended Face Oil: Mary Kay Naturally Nourishing Oil, $48, marykay.com

[This blend] uses a delicious combination of almond, olive, and sesame oils which leave your skin super hydrated, says Ursula Carranza, Beauty and Fashion Director of People en Espaol. You can even use it on your cuticles and on the ends of your hair!

Best Drugstore Eye Cream: Inkey List Caffeine, $9.99, sephora.com

Consider this concentrate a cup of coffee for your under-eye area. Studies show that when applied topically, caffeine can temporarily plump the skin, so a jolt of java-infused product to your weary eyes will make you look instantly more awake.

Best Luxury Eye Cream: Cl de Peau Enhancing Eye Contour Cream Supreme, $270, nordstrom.com

It takes six years to extract the iris extract in this luxe cream (three to cultivate and three to dry), the key result being plumper, firmer, revived skin. Using the cold steel applicator is kind of like jade rolling, and I visibly notice it de-puffing my eyes, mentioned one HG tester, adding that the difference between eyes after applying on just one side was very drastic.

Best Expert-Recommended Eye Cream: Care Skincare Eye + Lip Nourishing Cream, $30, careskincare.com

Formulated to treat and repair the delicate area around the eyes and around the lips, this dense cream with a light diffusing finish softens the look of fine lines and dark circles, says Dr. Robinson. It absorbs quickly and wont drift into eyes or interfere with makeup.

Best Drugstore Exfoliator: Yes To Grapefruit AHA + BHA Exfoliating Tonic, $14.99, amazon.com

AHAs and beta hydroxy acids (BHAs) are known chemical exfoliants that help remove dead skin cells and excess sebum inside the pores. This product is packed with 10% AHA and BHA, making it a powerful exfoliator that more sensitive skin types should probably avoid. However, if your skin can tolerate it, using it two to three times a week will help you achieve a more even skin texture and a newfound radiance.

Best Luxury Exfoliator: Pestle & Mortar Exfoliate Glycolic Acid Toner, $38, neimanmarcus.com

Brighten, tone, and reveal a more radiant face with the help of this glycolic-based exfoliator that will gently slough away dead skin cells. This non-irritating, vegan, and fragrance-free formula is suitable for all skin types.

Best Expert-Recommended Exfoliator: SkinBetter AlphaRet Peel Pads, $95, skinbetter.com

This new peel is an exfoliation powerhouse. Using lactic, glycolic, and salicylic acids, it exfoliates using AHAs and BHAs, targeting discoloration and age spots, boosting collagen and elastin production, and penetrating deep into the cell to dissolve dead skin buildup, explains Dr. Engelman. It contains a patented retinoid, AlphaRet and it also helps treat acne.

Best Drugstore Face Mask: AveenoOat Face Mask with Pomegranate Seed Extract, $8.38, amazon.com

This brightening face mask is a great option for people with dry skinit uses colloidal oatmeal to maintain the skins natural moisture barrier while treating and calming, dried skin. Studies show that pomegranate, the products key ingredient promotes skin cell turnover, which will help users achieve a more radiant appearance. The texture is rich and velvety, and youll get the best results if used two to three times a week.

Best Luxury Face Mask: Glow Recipe Avocado Melt Retinol Sleeping Mask, $49, sephora.com

Face masks dont have to be rinse-and-go. This Glow Recipe mask can be used overnight for added benefits or can work in as little as 20 minutes as a quick skin-boost if thats your preference. Its a good introduction to retinol, the gold-standard of anti-aging ingredients, since its gentle enough that even sensitive skin types can use it without irritation. One HG tester commented that her skin looked less congested the day after using it, and adding that she didnt notice any dryness sometimes associated with retinol products. Additionally, our testers were impressed by how cooling it felt on skin, how soft it felt after washing off, and how it didnt transfer onto their pillows when using overnight.

Best Expert-Recommended Face Mask: SkinBetter Refresh Detoxifying Scrub Mask, $55, skinbetter.com

The clay minerals [in this mask] absorb unwanted oil and remove toxins, while biodegradable spherical beads to lightly resurface the skin, explains Dr. Robinson of her top pick. It can even be used as a spot treatment on areas of focus!

Best Drugstore Sunscreen: La Roche-Posay Anthelios SPF 50 Mineral Sunscreen, $22.99, amazon.com

This mineral sunscreen is oil-free (so non-comedogenic), non-greasy, and easily blends into all skin shades without leaving a white cast. If youre a minimalist who prefers multi-purpose products, this SPF 50 made for face and body should be your go-to. Its TSA-approved too, so you can throw it into your carry-on next time you travel to the beach.

Best Luxury Sunscreen: Coola Mineral Sun Silk Crme SPF 30, $42, amazon.com

The silky feel this mineral sunscreen leaves on the skin will make you feel like a million bucks. And while yes, it makes you feel like you have a Chinchilla-soft face, what we love most about this sunscreen is its formulabecause science. It has UVA and UVB protection, which means that it will help protect your skin from both the suns harmful rays and also from the blue-light emitted from your smart-phone and laptop.

Best Expert-Recommended Sunscreen: Paulas Choice On-The-Go Shielding Powder SPF 30, $29, dermstore.com

Dermatologists recommend reapplying your sunscreen every two hours, but realistically, not too many of us are down for applying a lotion over our makeup throughout the day. Thats why this powder formula is so greatit comes in what looks like a makeup brush, and all you have to do is swipe it across your face to achieve your recommended SPF dose. Plus, it gets rid of excess shine, as Carranza points out.

Best Drugstore Lip Treatment: Awake BeautyMoisture Balm Daytime Lip Mask, $14, ulta.com

This hydrating lip mask smells a little like a watermelon Jelly Rancher and works like a charm thanks to its concentrated blend of vitamin E, olive, argan, rosehip, and raspberry seed oil. Testers unanimously gave it a high score for seeing noticeably healthier lips the day after using it. For best results, use overnight since it is on the thicker side.

Best Luxury Lip Treatment: Biossance Squalane + Rose Vegan Lip Balm, $18, sephora.com

A recent poll showed that 3 percent of the US population say they are vegan, which may not seem like a lot, but that amounts to nearly 10 million people (or nearly the entire population of the state of Michigan). For those 10 million, it may be important to pick a vegan lip treatment that aligns with their dietand this is the best one. It uses plant-derived squalane, rose oil, ceramides, and algae to nourish and plump skin. Apply at night and wake up to smoother and visibly healthier lips.

Best Expert-Recommended Lip Treatment: Hourglass No 28 Tinted Lip Treatment Oil, $49, sephora.com

Treat your mouth to a luxurious anti-aging treatment that rivals even the best high-shine lip glosses on the market. It uses top-notch ingredients that work to hydrate and plump the lips, comes in four colors, and has Carranzas stamp of approval: Its the perfect lip oilits deeply hydrating without being sticky, gives your lips an amazing shine, and the gold-plated applicator is to die for!

Best Self-Tanning Face Towelettes: Haute Bronze Face Towel, $35, haute-bronze.com

For those hoping to get a sun-kissed glow without having to spend time in the sun, these bronzing face towels are a holy grail, and theyre so easy to use! Just unfold the towel, and using circular motions apply it to dry, clean skinthats it! Plus, since it is a towel, its carry-on approved.

Best Self-Tanning Face Serum: Tan-Luxe Super Glow Hyaluronic Self-Tan Serum, $49, sephora.com

My goal: maintaining a year-round healthy glow without the harmful sun exposure. The answer: this magic fluid, shares DeSantis. Its an anti-aging serum and tanning formula rolled into one, so it helps me cut down on my skincare steps every morning and night, and Im left with a believable bronze that scores me tons of compliments.

Best Face Roller: MDNA Skincare The Beauty Roller, $150 (orig. $200), nordstrom.com

This tool is great for de-puffing and encouraging lymphatic drainage, since the grooves and rolling action stimulates circulation and relieves tension, says Dr. Engelman. It can also help drive your serum deep into the skin which allows your products to work better for younot to mention, it feels great! Regular lymphatic drainage can help reduce puffiness and can temporarily help sculpt the face for a more lifted look.

Best Gua Sha: Wildling Empress Stone, $65, wildling.com

If youre more of a Gua Sha person, clean beauty expert and HelloGiggles columnist Jessica Yarbrough recommends this product, saying its at the top of her list. I have an extensive collection of what I like to call non-skincare skincare products, things that support my skin without technically absorbing into my skin, she explains. With this in hand, [giving myself a] mindful facial massage has become a daily morning ritual, and my skin is healthier and glowier for it.

Best Device: Finishing Touch Flawless Dermaplane Glo, $19.88, amazon.com

Dermaplaning is a treatment primarily found at a dermatologist or estheticians office, wherein a sharp blade sloughs away the peach fuzz and dead skin cells from the uppermost layer of the skin, leaving your face radiant and baby-soft. This at-home device isnt as sharp as a professionals would be, for obvious safety reasons, but its effective in removing fuzz and excessive dead skin cell buildup. It also comes with a built-in light that improves visibility and in turn, helps you get better results.

Best Tweezers: Tweezerman Rose Gold Point Tweezer, $20.32, amazon.com

Designed for precision, this pointed-tip tweezer is the answer to effectively removing stubble, pesky ingrown hairs, and super-fine facial strands.

Skincare products arent, and shouldnt, be reserved for only the face. Advancements in skincare technology have allowed for the creation of body care products that blew our minds this year. From those that protect our skin from environmental aggressors and reverse signs of aging by altering our genetic code to innovative fuss-free packaging and bath products that are vagina-friendly, here are the body products were currently crushing on.

Best Drugstore Body Wash: Dove Body Wash Mousse with Rose Oil, $5.99, target.com

Body wash, but in a mousse. This gentle body wash has a subtle rose scent that you can actually scratch-and-sniff on the bottle to test out before purchasing. A little goes a long way for head-to-toe moisture.

Best Luxury Body Wash: LOccitane Almond Shower Shake, $25, amazon.com

Because we clearly love a fun body wash, its no wonder we loved this shower shake. Studies show that when applied topically, almond oil improves skin tone and prevents UV damage, and its the key ingredient in this product. To use, shake the bottle to mix the ingredients into a milky formula and apply onto wet skin. It will leave your body feeling hydrated and soft.. Now, Alexa, play OutKasts Hey Ya!.

Best Expert-Recommended Body Wash: Olay Moisture Ribbons Plus Shea Butter + Blue Lotus Body Wash, $6.99, walgreens.com

Dr. Mariwalla recommends this moisturizing body wash for its shea butterwhich studies show is anti-inflammatory, rich in antioxidants, and is a rich emollientperfect for sensitive skin types.

Best Drugstore Body Lotion: Curl Dry Skin Therapy Itch Defense, $10.94, amazon.com

Tackle dry, itchy skin with this fast-absorbing body lotion that almost instantly soothes and calms skin. This moisturizer, has approval from the National Psoriasis Foundation and holds a seal from the National Eczema Association, is packed with ceramides, vitamin E, and oatmeal extractand its moisturizing effects last all day.

Best Luxury Body Lotion: Augustinus Bader The Body Cream, $165, neimanmarcus.com

The advanced skincare technology in this cream makes this splurge worth the while. It uses epigenetic technology, which essentially tells the skin cells to be healthy overall and can over time alter your genetic code, and helping treat stretch marks, bumpy skin, and cellulite.

Best Expert-Recommended Body Lotion: CetaphilRestoring Lotion with Antioxidants, $19.97, amazon.com

Cetaphil took their classic lotion and supercharged it, says Dr. Engelman. This product is safe for sensitive skin and the added antioxidants and niacinamides will neutralize free radical damage and repair the skin barrier. Niacinamides have a similar effect to retinol, strengthening the skin barrier. However, unlike retinol, it fortifies from the get-go without sensitivity or irritation.

Best Drugstore Hand Cream: eos Shea Better Coconut Hand Cream, $2.98, amazon.com

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The best beauty products from the 2020 Beauty Crush Awards - Yahoo Lifestyle

Skin Stem Cells Comments Off on The best beauty products from the 2020 Beauty Crush Awards – Yahoo Lifestyle | February 29th, 2020

Numerous revelations about the world have been made in various forms throughout history. Researchers and scientists have continuously managed to discover new ways we can understand the world around us.

Major scientific breakthroughs have been made that helped improve our way of life and will make it easier for us to achieve even more amazing innovation eventually. The 21st century was especially fruitful for the advancements in technology and science. Many of the essential revelations in history were made during the last 100 years. This article will attempt to name the most important ones.

In 2015, the National Aeronautics and Space Administration managed to find evidence that proves that there is liquid water on Mars. Scientists were aided by the first spectrometer provided by NASA, called the Mars Reconnaissance Orbiter.

With its help, they were able to detect hydrated salts all over the distant planet. The hydrated salts are more prevalent during the warmer seasons, which means that water is a crucial ingredient in their development.

Many scientists believe this to be the most crucial discovery of the 21st century. Since Albert Einsteins theory of relativity was published, the thought of time travel has excited scientific minds all over the world.

The LIGO (Laser Interferometer Gravitational-Wave Observatory) project in the United States is responsible for the detection of gravitational waves, which would imply that with enough research time travel could be possible. Journey to the earliest parts of our universe does not seem as impossible as before since this discovery.

Proof of the existence of Dark Matter was found in 2006 by a team of researchers, led by Maxim Markevitch of the Harvard-Smithsonian Center for Astrophysics in Cambridge. They proved its existence by measuring the location of mass that gets created when galaxies collide. Specific clusters of mass get disconnected, and a large amount of visible matter is what makes up dark matter. While this sounds overly complex, the important thing to remember is that this proves that dark matter makes up for 68% of the universe.

Through stem cell research, we can provide better access to organs for patients, meaning that patients no longer need to wait for donors, making it easier to cure certain conditions. Stem cells make it possible to grow an indefinite number of cells of the same type, but other types of cells also arise from that process. This means that it is possible to regenerate organs using skin cells. In the future, the discovery could make it so that the organs needed for treatment are created in a laboratory.

The process of face transplantation uses tissues of a dead person to replace another persons face. It is a complicated process that was first successfully performed in France in 2005. This transplant was only partial, but the first full-face transplant happened only five years later in Spain.

Since then, people with significant congenital disabilities or facial disfigures caused by diseases or burns have had this procedure performed on them in multiple countries. What seemed like science fiction in the 1990s is a reality now.

HIV is considered to be one of the deadliest viruses in the history of humankind. While we still have not found a cure for AIDS, the disease that is a result of an HIV that has advanced too much, we did manage to improve the treatment of HIV.

With the new methods, some of which were developed in Germany, patients with HIV can live longer, almost being able to lead healthy lives. With these exciting advancements, millions of lives have been saved, and it is only a matter of time before we find a complete cure.

Scientists atthe Massachusetts Institute of Technology (MIT)managed to find a way to create false memories and plant them into the brains of mice. While this enormous scientific breakthrough can help us better understand the concept of mind and help us with dealing with mental health issues, it can still be problematic.

It does seem like a scene out of an old science fiction movie, but with this advancement, it could be possible to manipulate a persons memory, and who knows what consequences that could have.

Scientists at the University of Twente have developed robotic body parts using biomechanics. They managed to create robotic arms that can make life easier for people with severe muscular dystrophy. They also implemented previous research on prosthetics in creating these limbs. These robotic body parts could prove to be extremely helpful for wounded soldiers, people suffering from disabilities or older people.

A photon was teleported into space with the help of a laser beam by Canadian scientists. This process is called quantum teleportation, and it can be used to transport information about something, not its physical state. However, it is a step in the right direction of possibly making teleportation of physical matter possible someday.

The process itself was incredibly hard and required extreme precision. Photons are tiny, meaning that it will be a while until we can successfully teleport larger items.

Although the World Wide Web originated much earlier, it was not until the 21st century that we saw everything the internet is capable of. And it is a fitting way to end this list because it impacted more lives than anything else. Thanks to the internet, we can do things we could only imagine 20 years ago, basically from anywhere in the world. We can access any information, watch every movie in existence, or talk to people halfway around the world. It is the discovery of all the possibilities of the internet that made this century what it is, at least for now.

Where and when was the first face transplant procedure performed?

The first face transplant procedure was performed in France in 2005.

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10 Of The Biggest World Revelations In The 21st Century - World Atlas

Skin Stem Cells Comments Off on 10 Of The Biggest World Revelations In The 21st Century – World Atlas | February 29th, 2020

The bag that Laurie Olesen gripped as she walked through the airport looked like any other carry-on. But the bright blue canvas tote would carry more than her cellphone, e-reader and toiletries. It would hold the last, best hope of survival for a desperately ill patient.

Bound for the East Coast, Olesen was on a mission to pick up blood stem cells or bone marrow provided by a donor, then fly with it to another city where it would be transfused into a recipient.

The product travels so the patient or the donor doesnt have to, said Olesen, 66, of St. Paul.

Olesen is a volunteer courier for Be The Match. Based in Minneapolis, the nonprofit registry serves people diagnosed with a variety of life-threatening blood, bone marrow or immune system disorders.

Shes one of a cadre of 400 of specially trained volunteers that form a crucial, reliable and affordable link between donors and patients.

These couriers are prepared to get a call, race to the airport and reach across time zones with a perishable product that comes with a true deadline. The consequences of a delay can be devastating even lethal for a patient waiting for the unique match.

Our volunteer couriers have to work on a tight time frame. They manage the paperwork and fill out a chain of custody form to document exactly where the cooler has been. We want it in a volunteers line of sight at all times, said Rut Kessel, volunteer specialist with Be The Match. They protect it with their life because it is a life.

Only about 30% of patients who need a bone marrow or blood stem cell transplant can find a donor within their family. Be The Match provides an international database of more than 20 million to locate an unrelated donor. When such a wide net is cast, the recipient and the anonymous donor rarely live in the same city. Theyre usually in different states, regions or even countries.

Thats where couriers come in.

Many volunteers are retirees who have time and flexibility. The gig also attracts firefighters, health care workers and airline employees whose shift work creates consistent open days in their schedules. While costs for their flights, hotels and other travel expenses are covered, couriers arent paid for their time.

They also never meet or even learn the names of donors or recipients. They typically pick up a numbered product at one lab and deliver it to another.

We have strict rules about confidentiality, explained Kessel. The courier experience is detached from the people involved. Something has gone terribly wrong if a courier ever meets or even sees a donor or recipient.

But many couriers have a personal connection to Be The Match.

Someone did this for me, said Lisa Maxson, 37, an Ohioan who was diagnosed with acute myeloid leukemia in 2011. Hospitalized for months while she had chemotherapy and radiation to kill her own diseased bone marrow, the mother of three underwent a transplant to replace it with healthy donor cells.

While I was sick, I decided I would give back to the organization that saved my life, she said.

Her family now sponsors a 5K race to benefit Be The Match. And this winter, she traveled to the registrys headquarters in the North Loop to take the two-day training for volunteer couriers.

Im so excited to be a courier for my transplant brothers and sisters, she said. I know what it feels like to be afraid youre going to die.

The gift of life

Every day of the year, volunteer couriers are in the air, crisscrossing the globe with the lifesaving cargo in temperature-controlled, medical coolers tucked under their seats. Last year, couriers living in 15 America cities made more than 2,600 trips, about a quarter of them to international destinations.

Five Minneapolis-based travel agents book their flights and manage their itineraries. Like the couriers themselves, the travel agents have to be nimble and act quickly when the unexpected occurs. Theyre also on call round-the-clock to rebook trips if mechanical difficulties cancel a flight or blizzards or hurricanes snag the travel grid, said Bonnie Bagley, who supervises the agents.

Two years ago, Bagley became a volunteer courier and now uses vacation time to make deliveries.

That closed the loop for me. Now I literally see how the system that Ive had a glimpse of works for patients, she said. I understand the passion our volunteers have. Theres an adrenaline rush when youre carrying the product.

For her part, Olesen likely holds the record for the most trips, which she estimates at a hundred deliveries to every region in the United States as well as a number of foreign countries.

Couriers have to be assertive, but must also remain calm, she said. You cant get rattled when things dont go as planned.

Shes also learned how best to deal with foreign customs agents.

When you bring a product from another country into the U.S., you have to declare the product to customs. You carry a special letter but, to tell the truth, some agents dont know what theyre supposed to do. Thats where the diplomacy comes in, said Olesen. You have to help them do their job without alienating or provoking them. You learn to kill them with kindness.

In 1986, Olesen was the first employee for the organization that became Be The Match. A registered nurse, she was working with blood collection at the American Red Cross in St. Paul when it was among a consortium of blood banks that received a grant to develop the nations first bone marrow registry. She was hired to identify donors for specific patients.

She joined the few lab technicians and transplant center employees who flew donations from donor to recipient. As the registry expanded and number of patients and donors increased, Be The Match added volunteer couriers in 2004. Olesen set up volunteer and education programs, managed search operations and kept up her courier duties.

While most of her deliveries have gone off without a hitch, shes had a few near-misses, including the time when a drop-off spot in Barcelona turned out to be a dead end. She and a cabdriver bridged their language gap to figure out the correct spot and hustle across the city.

We have guidelines for tipping, but that was one time I gave a little extra out of my own pocket, Olesen recalled. He went above and beyond.

And then there was the time when she had completed her pickup in London, only to arrive at the airport as a snowstorm shut it down.

I went to the gate agent and told them I was carrying bone marrow. They declared the flight a life flight and our plane was prioritized. We took off when the first runway opened, she said. When you do this, the courier gods are always on your side.

Although Olesen retired from Be The Match two years ago, she keeps her bag packed and her passport ready so she can continue to fly as a volunteer.

Being a courier reminds me of what were about. It affirms why we do what we do every day, she said. I know that within 24 hours after I get the product to its destination, it will be transfused into the recipient. That can give a person their life back.

Kevyn Burger is a Minneapolis based freelance broadcaster and writer.

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Carry-on item for Be the Match volunteers: Organ transplants - Minneapolis Star Tribune

Bone Marrow Stem Cells Comments Off on Carry-on item for Be the Match volunteers: Organ transplants – Minneapolis Star Tribune | February 28th, 2020

TEL AVIV, Israel, Feb. 27, 2020 /PRNewswire/ -- BioLineRx Ltd. (NASDAQ: BLRX), (TASE: BLRX), a late clinical-stage biopharmaceutical company focused on oncology, announced today that a Notice of Allowance has been issued by the United States Patent and Trademark Office (USPTO) for a patent application claiming the use of motixafortide (BL-8040), a novel immunotherapy compound, combined with any PD-1 inhibitor, for the treatment of any type of cancer.

The PD-1 antagonist can be any agent that prevents and/or inhibits the biological function and/or expression of PD-1, such as pembrolizumab (KEYTRUDA). The targeted cancer can be solid, non-solid, and/or a cancer metastasis.

This patent, whenmedi issued, will be valid until July 2036 with a possibility of up to five years patent term extension. Additional corresponding patent applications are pending in Europe, Japan, China, Canada, Australia, India, Korea, Mexico, Brazil and Israel.

"We are extremely pleased to receive this valuable notice of allowance from the USPTO, which entitles us to long-term, highly enforceable and broad patent protection for our lead product, motixafortide, in combination with any PD-1 inhibitor, and more importantly, for all cancer indications, including, of course, any solid tumor," stated Philip Serlin, Chief Executive Officer of BioLineRx. "This important patent allowance also supports our ongoing Phase 2a COMBAT/KEYNOTE-202, for which we have recently completed patient recruitment in the triple combination arm investigating the safety, tolerability and efficacy of motixafortide, KEYTRUDA and chemotherapy. Following promising initial results demonstrating robust and durable responses to the triple combination treatment, we look forward to the progression-free and overall survival data from the triple combination arm expected in mid-2020."

The COMBAT/KEYNOTE-202 Study

The Phase 2a COMBAT/KEYNOTE-202 study was originally designed as an open-label, multicenter, single-arm trial to evaluate the safety and efficacy of the dual combination of motixafortideand KEYTRUDA (pembrolizumab), an anti-PD-1 therapy marketed by Merck & Co., Inc., Kenilworth, N.J., USA (known as MSD outside the United States and Canada), in over 30 subjects with metastatic pancreatic adenocarcinoma. The study was primarily designed to evaluate the clinical response, safety and tolerability of the combination of these therapies, and was carried out in the US, Israel and additional territories. The study is being conducted by BioLineRx under a collaboration agreement signed in 2016 between BioLineRx and MSD, through a subsidiary.

In July 2018, the Company announced the expansion of its immuno-oncology collaboration with MSD to include the triple combination arm investigating the safety, tolerability and efficacy of motixafortide, KEYTRUDA and chemotherapy as part of the Phase 2a COMBAT/KEYNOTE-202 study. In January 2020, the Company announced completion of recruitment of the 40 patients planned for the triple combination arm of the study.

About Motixafortide in Cancer Immunotherapy

Motixafortideis targeting CXCR4, a chemokine receptor and a well validated therapeutic target that is over-expressed in many human cancers including PDAC. CXCR4 plays a key role in tumor growth, invasion, angiogenesis, metastasis and therapeutic resistance, and CXCR4 overexpression has been shown to be correlated with poor prognosis.

Motixafortideis a short synthetic peptide used as a platform for cancer immunotherapy with unique features allowing it to function as a best-in-class antagonist of CXCR4. It shows high-affinity, long receptor occupancy and acts as an inverse agonist.

In a number of clinical and preclinical studies, motixafortidehas been shown to affect multiple modes of action in "cold" tumors, including immune cell trafficking, tumor infiltration by immune effector T cells, and reduction in immunosuppressive cells (such as MDSCs) within the tumor niche, turning "cold" tumors, such as pancreatic cancer, "hot" (i.e., sensitizing them to immune checkpoint inhibitors and chemotherapy).

About BioLineRx

BioLineRx Ltd. (NASDAQ: BLRX), (TASE: BLRX) is a clinical-stage biopharmaceutical company focused on oncology. The Company's business model is to in-license novel compounds, develop them through clinical stages, and then partner with pharmaceutical companies for further clinical development and/or commercialization.

The Company's lead program, motixafortide, is a cancer therapy platform currently being evaluated in a Phase 2a study for the treatment of pancreatic cancer in combination with KEYTRUDA and chemotherapy under a collaboration agreement with MSD. Motixafortideis also being evaluated in a Phase 2b study in consolidation AML and a Phase 3 study in stem cell mobilization for autologous bone-marrow transplantation.

BioLineRx is developing a second oncology program, AGI-134, an immunotherapy treatment for multiple solid tumors that is currently being investigated in a Phase 1/2a study.

For additional information on BioLineRx, please visit the Company's website at http://www.biolinerx.com, where you can review the Company's SEC filings, press releases, announcements and events. BioLineRx industry updates are also regularly updated on Facebook,Twitter, and LinkedIn.

Various statements in this release concerning BioLineRx's future expectations constitute "forward-looking statements" within the meaning of the Private Securities Litigation Reform Act of 1995. These statements include words such as "may," "expects," "anticipates," "believes," and "intends," and describe opinions about future events. These forward-looking statements involve known and unknown risks and uncertainties that may cause the actual results, performance or achievements of BioLineRx to be materially different from any future results, performance or achievements expressed or implied by such forward-looking statements. Some of these risks are: changes in relationships with collaborators; the impact of competitive products and technological changes; risks relating to the development of new products; and the ability to implement technological improvements. These and other factors are more fully discussed in the "Risk Factors" section of BioLineRx's most recent annual report on Form 20-F filed with the Securities and Exchange Commission on March 28, 2019. In addition, any forward-looking statements represent BioLineRx's views only as of the date of this release and should not be relied upon as representing its views as of any subsequent date. BioLineRx does not assume any obligation to update any forward-looking statements unless required by law.

Contact:

Tim McCarthyLifeSci Advisors, LLC+1-212-915-2564tim@lifesciadvisors.com

or

Tsipi HaitovskyPublic Relations+972-52-598-9892tsipihai5@gmail.com

View original content:http://www.prnewswire.com/news-releases/biolinerx-announces-notice-of-allowance-from-uspto-for-patent-covering-motixafortide-bl-8040-in-combination-with-anti-pd-1-for-the-treatment-of-any-type-of-cancer-301012614.html

SOURCE BioLineRx Ltd.

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BioLineRx Announces Notice of Allowance from USPTO for Patent Covering Motixafortide (BL-8040) in Combination With Anti-PD-1 for the Treatment of Any...

Bone Marrow Stem Cells Comments Off on BioLineRx Announces Notice of Allowance from USPTO for Patent Covering Motixafortide (BL-8040) in Combination With Anti-PD-1 for the Treatment of Any… | February 28th, 2020

Creative Medical Technology Holdings Inc (OTCMKTS: CELZD) is the new temporary ticker symbol for Creative Medical since the Company affected a 1 for 150 reverse stock split. According to the Company the reverse split of our stock was a decision that did not come lightly. In order to secure more competitive financing terms and to reduce existing convertible debt, the company needed to return to the OTCQB. As the fundamentals of the company continue to expand and commercialization is ramped up, this was determined to be necessary for the long-term benefit to the company and its shareholders. The reverse affected ALL shareholders, including founding shareholders, Officers and Directors, who have substantial holdings in CELZ stock as well said Timothy Warbington CEO.

Microcapdaily has been reporting on CELZ for years; on November 18, 2018 we stated: CELZ is an exciting stock that has attracted legions of shareholders who see big things happening here. CELZ flagship CaverStem has the only procedure to treat Erectile Dysfunction with adult stem cells in the US. CELZ has runner in its blood and a long history of huge moves skyrocketing from $0.002 in March of last year to highs topping $0.07 per share in August, CELZ loves to run and is a volume leader regularly among the top most traded on the OTCBB.

Creative Medical Technology Holdings Inc (OTCMKTS: CELZ)is a commercial-stage biotechnology company focused on Urology and Neurology using stem cell treatments. The companys team consists of leading international researchers in regenerative medicine with a science-first approach to treatments ensuring that all of its treatments are proven to be both safe and effective. CELZ is engaged in stem cell research and applications for use to treat male and female sexual dysfunction, infertility and related issues. It holds a patent for its erectile dysfunction (ED) treatment and was granted a license by Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, a non-profit biomedical research and education institute (LABIOMED), for the infertility treatment. It has also filed a patent application focused on physical manifestations of female sexual arousal disorder, as an extension of the work with stem cell therapies for ED.

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Creative Medical is currently in the commercial stage of its bone marrow based stem cell treatment for Erectile Dysfunction known as CAVERSTEM, which is being marketed both nationally, and internationally. Earlier this year the Company formed the subsidiary CaverStem International LLC for the purpose of commercializing its erectile dysfunction technology to international physicians.

In a recent update on StemSpine Thomas Ichim PhD of CELZ stated StemSpine will surely be welcomed by the over 50 million Americans suffering from CLBP in the United States as a drug free alternative. Currently, there are minimal treatment options for patients that suffer from this debilitating pain, with roughly 50% of patients progressing to opioids and surgery. I have been thrilled with the positive reception of StemSpine across all fronts as we quietly progressed the program forward over the last few months, said Timothy Warbington, President and CEO of Creative Medical Technology Holdings, Inc. I am especially energized by the positive reception from potential healthcare providers who have overwhelmingly confirmed this is a necessary and highly desirable alternative to current treatment options. We look forward to partnering with these providers and bringing this therapy to the forefront in 2020 for the benefit of the many patients that stand to benefit from it and for our shareholders as we think it will drive tremendous value for the organization.

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Currently on the move since affecting a 1 for 150 reverse CELZ is fully reporting OTCBB, has minimal cash in the treasury, $3.9 million in current liabilities and some revenues reporting $169k in sales in 2019. we stated in 2016: CELZ is an exciting Company developing in small caps; CELZ flagship CaverStem has the only procedure to treat Erectile Dysfunction with adult stem cells in the US. The erectile dysfunction market is booming! According to a recent report from ResearchAndMarkets.com, the size of the global erectile dysfunction market is expected to reach $4.25 billion by 2023. CELZ loves to run and is a volume leader regularly among the top most traded on the OTCBB.We will be updating on CELZ when more details emerge so make sure you are subscribed to Microcapdaily so you know whats going on with CELZ.

Disclosure: we hold no position in CELZ either long or short and we have not been compensated for this article.

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Creative Medical Technology Holdings Inc (OTCMKTS: CELZD) Gets the Extra D - MicroCap Daily

Bone Marrow Stem Cells Comments Off on Creative Medical Technology Holdings Inc (OTCMKTS: CELZD) Gets the Extra D – MicroCap Daily | February 28th, 2020

People sustain approximately 11,000 spinal cord injuries each year. Approximately 38 percent of these injuries are from motor vehicle accidents, with falls, gunshot wounds and similar forms of violence, sports activities, and medical or surgical complications being the other common causes. Regardless of the cause, any type of injury to the spinal cord can damage nerve and tissue cells and result in a loss of sensation or paralysis. One possible option for treatment for spinal cord injuries thats been gaining traction among Beverly Hills spine surgeons is the use of stem cells.

Stem cells are undifferentiated cells. They are useful because they can become specialized cells in the area where they are injected. These cells have the ability to become other types of cells and encourage the production of new, healthy cells. There are two common types of stem cells:

Normally, treatment for a spinal cord injury involves extensive physical therapy and rehab. However, recovery is usually limited because an injured spine cant heal due to the formation of scar tissue triggered by an inflammatory response that keeps healthy cells from reaching the damaged area.

Adult stem cells that come from either bone marrow or a donated umbilical cord from a healthy pregnancy are usually used for spinal injury treatments. With umbilical cord tissue, there is a more rigorous screening process to look for viruses and bacteria. Regardless of how they are collected, stem cells may help with spinal cord injuries by:

Along with a local anesthetic, stem cells used to treat spinal injuries are injected directly into the affected area, and they are placed into the spinal fluid to allow the undefined cells to reach the injured part of the spine. Patients usually receive multiple injections over the course of several weeks. Treatment is often coupled with:

Stem cells wont completely repair an injured spinal cord. However, there are several promising studies that suggest some patients do see noticeable improvements, such as the ability to feel light touch below the injured area. At one facility in India, eight out of ten patients with no motor or sensory function below the waist were able to walk for about an hour with the assistance of a walker eight months after receiving transplanted stem cells. Stem cell therapy is still in its infancy, but it does offer some hope for patients with spinal injuries who are looking for an alternative to minimally invasive spinal surgery. Beverly Hills residents should contact The Spine Institute at 310-828-7757 for more information.

According to an article on Beckers Spine Review, The Spine Institutes Dr. Hyun Bae has spent a significant amount of time researching stem cell repair for degenerative disc disease as well as how growth factors can treat spinal cord injuries. Dr. Bae was among the first spinal surgeons to utilize growth factor tissue engineering for intervertebral discs, and in 2010 he also chaired a course for the North American Spine Society that dealt with navigating research in spinal biologics.

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Stem Cells: A New Way to Potentially Treat Spinal Cord ...

Spinal Cord Stem Cells Comments Off on Stem Cells: A New Way to Potentially Treat Spinal Cord … | February 28th, 2020

Last summer, U.S. Army Lieutenant Colonel Jason Barnhill traveled to an undisclosed desert location in Africa with a ruggedized 3D printer and other basic supplies that could be used to biofabricate for field medical care, such as human mesenchymal stem/stromal cells (hMSCs). The aim was to discover how a 3D bioprinter could expedite healing and even replace damaged tissue for troops injured in combat.

Jason Barnhill with a 3D bioprinter that could replace damaged tissues for troops injured on the battlefield. (Image: Military Health System/West Point)

Barnhill, who is the life science program director of the United States Military Academy West Point Department of Chemistry and Life Sciences, is leading a project with a team of cadets working on experiments to advance bioprinting research in the field with an ultimate goal to develop technology for creating wound-healing biologics, bandages, and more for soldiers on-site or near the point-of-care. According to U.S. Army news, 26 first-class cadets at the United States Military Academy at West Point, in New York, are doing bioprinting research across seven different projects: two teams are working on biobandages for burn and field care; other two teams are working on how to bioengineer blood vessels to enable other bioprinted items that require a blood source, such as organs, to be viable; while one team is working on printing a viable meniscus, and another team is looking to print a liver.

A lot of this has to do with the bioink that we want to use, exactly what material were using as our printer ink, if you will, explained Class of 2020 cadet Allen Gong, a life science major conducting research for the meniscus project. Once we have that 3D model where we want it, then its just a matter of being able to stack the ink on top of each other properly.

Gong, along with his teammates, are researching how to use bioinks to create a meniscus that could be implanted into a soldiers injured knee, while other cadets are seeking to print a liver that could be used to test medicine and maybe one day eliminate the shortage of transplantable organs. This is not the first time we hear the U.S. Army is using bioprinting for regenerative medicine, after all, they often suffer from trauma, resulting in loss of limbs, injuries to the face and severe burns. Deployed soldiers confront the risks of battle on a daily basis. However, being able to have immediate access to specialized bioprinters created to solve catastrophic medical injuries could be the dream-scenario solution many have been waiting for.

In 2014, scientists at the Armed Forces Institute of Regenerative Medicine (AFIRM), established by the Department of Defense, were using 3D bioprinters extensively for skin repair research; but the Army is also actively developing artificial 3D printed hearts, blood vessels, and other organs in a quest to develop customizable and 3D printed medicine. Barnhills pilot program in 2019, conducted by the Uniformed Services University of the Health Sciences (USU) in collaboration with the U.S. Military Academy at West Point, has shown that a 3D printer capable of biofabrication could potentially change the way deployed warfighters receive care also. Under his direction, the 3D printer successfully fabricated a number of products, including a scalpel capable of immediate use and a hemostat (a surgical tool used to control bleeding during surgery and capable of gripping objects) while locking them into place to hold a tissue or other medical implements. The tools were made of a material that could be sterilized on-site, reducing the chance of infection during practical use.

Common combat injuries include second and third-degree burns, broken bones, shrapnel wounds, brain injuries, spinal cord injuries, nerve damage, paralysis, loss of sight and hearing, post-traumatic stress disorder (PTSD), and limb loss. Many of these injuries could be tackled with customizable, on-site bioprinting machines, but for now, the cadets on each of the teams are in the beginning stages of their research before starting the actual printing process. This stage includes reading the research already available in their area of focus and learning how to use the printers, and after spring break, they will have their first chance to start printing with cells. The teams focusing on biobandage, meniscus, and liver will try to print a tangible product by the end of the semester as part of the initial research.

Another cadet and life science major working on the meniscus project, Thatcher Shepard, described in the U.S. Army article that there are definitely some leaps before we can get to that point [of actually implanting what they print]. We have to make sure the body doesnt reject the new bioprinted meniscus and also the emplacement. There can be difficulties with that. Right now, were trying to just make a viable meniscus, then, well look into further research to be able to work on methods of actually placing it into the body.

They claim that the meniscus team is starting with magnetic resonance images (MRI) of knees and working to build a 3D model of a meniscus, which they will eventually be able to print. A great deal of the teams research will be figuring out how and when to implant those cells into the complex cellular structure they are printing.

Cadets at West Point Department of Chemistry and Life Sciences (Image: West Point)

According to Michael Deegan, another life science major and cadet working on one of the blood vessel projects, for now, it will involve a lot of research into what has already been done in the field and the questions that still need to be answered. He described the experience as kind of like putting the cart before the horse. Saying that youve printed it, great, but whats the point of printing it if its not going to survive inside your body? Being able to work on that fundamental step thats actually going to make these organs viable is what drew me and my teammates to be able to do this. Deegan and his colleagues will eventually decide on the scope and direction of their projects, knowing that their research will be key to allowing other areas of the field to move forward, since organs, such as livers and pancreases, have been printed, but so far, they can only be produced at the micro level because they have no blood flow.

While generating organs and blood vessels will be one of the great benefits of customized medicine in the future, the work behind the biobandage teams could have a direct use in the field during combat. The U.S. Army suggests that the goal is to be able to take cells from an injured soldier, specifically one who suffered burns and print a bandage with built-in biomaterial on it to jumpstart the healing process. Medical personnel could potentially be deployed with a 3D printer in their Forward Operating Base or it could be sent along in a column with a Humvee to enable bandages to be printed on-site.

Were researching how the body actually heals from burns, said Channah Mills, a life science major working on one of the biobandage projects. So, what are some things we can do to speed along that process? Introducing a bandage could kickstart that healing process. The faster you start healing, the less scarring and the more likely youre going to recover.

Being on the forefront of it and just seeing the potential in bioengineering, its pretty astounding, Gong said. But it has also been sobering just to see how much more complicated it is to 3D print biomaterials than plastic.

At the moment, the projects are building on existing research on printing sterile bandages and then adding a bioengineering element. The bandages would be printed with specialized skin and stem cells necessary for the healing process.

More than half of the cadets working on the bioprinting projects plan to continue on to medical school following their graduation from West Point. This research, which will be presented during the academys annual Projects Day on April 30, is a great starting point for the future army doctors, as they begin to understand and work on some of the more complex technologies that could become their allies in the future, helping them heal soldiers in the field.

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West Point: Bioprinting for Soldiers in the Battlefield - 3DPrint.com

Spinal Cord Stem Cells Comments Off on West Point: Bioprinting for Soldiers in the Battlefield – 3DPrint.com | February 28th, 2020

There is new hope for patients with a rare autoimmune disorder. In mild cases, scleroderma causes areas of hardened skin. But in severe cases, it can also cause deadly hardening of internal organs like the lungs.

A transplant typically used to treat cancer is having remarkable results for patients who had little hope of surviving.

A year ago, Chuck Beschta couldn't walk more than a few minutes without stopping to rest.

"Just going out and doing normal activities outside raking the lawn, mowing the grass, shoveling the driveway, whatever, snow blowing those became impossible," he said.

After months of testing, he was diagnosed with severe scleroderma, which was hardening his skin. But even worse, it was hardening his lungs, making it hard to breathe.

"He was getting worse despite the best therapy we had to offer," University of Wisconsin rheumatologist Dr. Kevin McKown said.

McKown recommended a stem cell transplant newly approved for scleroderma to reboot Beschta's immune system.

"There's a process by which they try to remove the autoreactive immune cells, the cells that are caught in the immune process, and then they infuse that back in and hope that the body will basically take up and graft that immune system," McKown said.

Beschta saw almost immediate results. His skin was softer and his breathing improved. He hopes his scleroderma has been cured.

"I think we can be optimistic, and so far the people who have been followed out as far as 10 years out don't seem to be getting it back," McKown said.

Without a transplant, less than half the patients who have diffuse scleroderma and severe lung disease live 10 years past diagnosis.

Stem cell transplants are commonly used to treat leukemia and lymphoma, cancers that affect the blood and lymphatic system.

MEDICAL BREAKTHROUGHSRESEARCH SUMMARYTOPIC: NEW THERAPY FOR SCLERODERMAREPORT: MB #4698

BACKGROUND: Scleroderma is an autoimmune rheumatic disease where an overproduction of collagen produced in the body tissues causes the skin and internal organs to harden. The symptoms and effects range by person, but some common symptoms include hardened patches of skin (locations on the body vary,) painful and numb-feeling fingers and toes, and sharp internal pain in the esophagus, intestines, heart, lungs, or kidneys. Women are four times as likely to have scleroderma and the onset is between 30 and 50 years of age. However, anyone from infants to the elderly can have scleroderma. Possible risk factors include having certain gene variations as other family members, ethnic groups, exposure to certain medications or drugs, and already having another autoimmune disease, like rheumatoid arthritis, lupus or Sjogren's syndrome. (Source: https://www.scleroderma.org/site/SPageNavigator/patients_whatis.html;jsessionid=00000000.app30132b?NONCE_TOKEN=9B76519DF6B5819859319F0B63B805C9#.XheCGVVKhaQ , https://www.mayoclinic.org/diseases-conditions/scleroderma/symptoms-causes/syc-20351952 )

DIAGNOSING: A physical exam will be conducted as well as a blood test to check for elevated levels of antibodies the immune system produced. The doctor will also take a sample of skin to be tested in the lab. If there are complaints about internal pain, the doctor may run other tests, including imaging, organ function, and other blood tests. (Source: https://www.mayoclinic.org/diseases-conditions/scleroderma/diagnosis-treatment/drc-20351957 )

NEW TECHNOLOGY: A new stem cell transplant that's commonly known to treat cancer is improving the quality and quantity of life for those with scleroderma. Rheumatologists at University of Wisconsin Health tested the treatment since they have already been conducting bone marrow transplants for decades. Surgeons take out a sample of the patient's bone marrow, isolate the stem cells, and use radiation and chemotherapy to clean out their immune system. The same stem cells are later injected back into the patient's immune system with the hope that new cells will grow and the system is rid of the bad ones. The process is dangerous when the cells are taken out because the patient's immune system is more vulnerable, making infections more likely to occur. However, after four and a half years, 79% of patients that underwent the treatment were alive without serious complications compared to 50% that were treated with the original drugs. (Source: https://madison.com/wsj/news/local/health-med-fit/man-with-severe-autoimmune-disease-gets-stem-cell-transplant-at/article_7e8e17a5-21da-52f8-b728-fe584dab2b77.html)

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New technique developed to treat hardening of internal organs - WNDU-TV

Skin Stem Cells Comments Off on New technique developed to treat hardening of internal organs – WNDU-TV | February 28th, 2020

Parkinsons disease researchers have used gene-editing tools to introduce the disorders most common genetic mutation into marmoset monkey stem cells and to successfully tamp down cellular chemistry that often goes awry in Parkinsons patients.

The edited cells are a step toward studying the degenerative neurological disorder in a primate model, which has proven elusive. Parkinsons, which affects more than 10 million people worldwide, progressively degrades the nervous system, causing characteristic tremors, dangerous loss of muscle control, cardiac and gastrointestinal dysfunction and other issues.

Marina Emborg

We know now how to insert a single mutation, a point mutation, into the marmoset stem cell, says Marina Emborg, professor of medical physics and leader of University of WisconsinMadison scientists who published their findings Feb. 26 in the journal Scientific Reports. This is an exquisite model of Parkinsons. For testing therapies, this is the perfect platform.

The researchers used a version of the gene-editing technology CRISPR to change a single nucleotide one molecule among more than 2.8 billion pairs of them found in a common marmosets DNA in the cells genetic code and give them a mutation called G2019S.

In human Parkinsons patients, the mutation causes abnormal over-activity of an enzyme, a kinase called LRRK2, involved in a cells metabolism. Other gene-editing studies have employed methods in which the cells produced both normal and mutated enzymes at the same time. The new study is the first to result in cells that make only enzymes with the G2019S mutation, which makes it easier to study what role this mutation plays in the disease.

The metabolism inside our stem cells with the mutation was not as efficient as a normal cell, just as we see in Parkinsons, says Emborg, whose work is supported by the National Institutes of Health. Our cells had a shorter life in a dish. And when they were exposed to oxidative stress, they were less resilient to that.

The mutated cells shared another shortcoming of Parkinsons: lackluster connections to other cells. Stem cells are an especially powerful research tool because they can develop into many different types of cells found throughout the body. When the researchers spurred their mutated stem cells to differentiate into neurons, they developed fewer branches to connect and communicate with neighboring neurons.

We can see the impact of these mutations on the cells in the dish, and that gives us a glimpse of what we could see if we used the same genetic principles to introduce the mutation into a marmoset, says Jenna Kropp Schmidt, a Wisconsin National Primate Research Center scientist and co-author of the study. A precisely genetically-modified monkey would allow us to monitor disease progression and test new therapeutics to affect the course of the disease.

The concept has applications in research beyond Parkinsons.

We can use some of the same genetic techniques and apply it to create other primate models of human diseases, Schmidt says.

The researchers also used marmoset stem cells to test a genetic treatment for Parkinsons. They shortened part of a gene to block LRRK2 production, which made positive changes in cellular metabolism.

We found no differences in viability between the cells with the truncated kinase and normal cells, which is a big thing. And when we made neurons from these cells, we actually found an increased number of branches, Emborg says. This kinase gene target is a good candidate to explore as a potential Parkinsons therapy.

This research was supported by grants from the National Institutes of Health (R24OD019803, P51OD011106 and UL1TR000427).

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Cells carrying Parkinson's mutation could lead to new model for studying disease - University of Wisconsin-Madison

Cardiac Stem Cells Comments Off on Cells carrying Parkinson’s mutation could lead to new model for studying disease – University of Wisconsin-Madison | February 28th, 2020

Feb 25, 2020 12:00 AM

Every ten minutes, someone passes away from a blood disorder. Thats 148 people a day. There is a way to prevent many of these deathsa bone marrow transplant. DNA matching has the power to help thousands of people waiting for a life-saving bone marrow donation, but this special donor list depends entirely upon the willingness of individuals to sign up. Could your unique DNA hold the match that helps one person live to see tomorrow? Heres how you can find out.

Be The Match is a global hub for bone marrow donor registry working with hundreds of partners to support the transplant community. Signing up is easy online. You provide registration information, receive a kit in the mail, use the DNA swab as directed, and send it back for DNA typing. Your potentially life-saving information is secure and becomes available to specialized doctors around the world.

Even if you arent a match right away, the fact that every three minutes a person is diagnosed with a blood disorder means you could be called at any time to be a hero in someones time of need. Paloma Cariello, MD, MPH, says, Its absolutely a life-saving procedure. Its a new life that people getwe call it a new birthday, and at many hospitals they give it as a new birthday date in their chart. We sing Happy Birthday. Its a big event.

To find a close enough match to help fortify a patients immune system, doctors have to be precise. They first reach out to family, but even then, only 30% of patients find a good match. The odds of finding a match in an unrelated donor can be as low as 18%, especially with minorities.

The need for more individuals of every background cannot be overstated, says University of Utah Health Hematologist Sagar Patel, MD. He emphasizes the need for ethnic minorities to register. Every ethnicity is represented in the pool of patients, so the donor pool likewise needs to be diversified to improve the availability of similar DNA typing.

If a doctor finds you to be a suitable match, they select the ideal method for their patient and prepare you for donation. There are two donation methods: peripheral blood stem cell (PBSC) donation and bone marrow donation. Because every donor is carefully screened and prepared, and because a small amount of fluid is ultimately needed, neither procedure method impacts the performance of your own immune system, says Cariello.

With PBSC retrieval, you receive a stimulant for five days to increase the presence of blood-forming cells in your blood stream. Then a refined process of extraction occurs: Your blood is drawn, a machine collects just the cells the patient needs, and your remaining fluids are safely returned to you. This process can usually be done in one eight-hour session. Most donors report a full recovery within a week to 10 days, but you will be followed-up with until your full recovery.

If the doctor determines that the patient needs bone marrow, your procedure is a bit different. Marrow needs to be drawn from your pelvic bones. It happens in a hospital and under anesthesia, and you will feel no pain as the donation is collected. You can go back to routine activity the same day, and your system fully replenishes within four to six weeks.

Even with thousands of people in need, only about one in 430 donors in the Be The Match system are called in as a match. And the simple processes and expert professional care you receive minimize potential risk. A common side effect is bruising at the procedure sites, and some donors occasionally experience mild pain, fatigue, or dizziness. Reactions related to the use of anesthesia might also occur.

With such little risk, it shouldnt be a question as to whether you sign up, but when. And today is a perfect day. The low odds of finding a cure that these patients face are as extreme as the high rewards that await themand youwhen you make the choice to become a donor. Visit BeTheMatch.org to learn more and to become the one who initiates the miraculous call: We found a match.

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Be a Bone Marrow Hero - University of Utah Health Care

Bone Marrow Stem Cells Comments Off on Be a Bone Marrow Hero – University of Utah Health Care | February 26th, 2020

CHICAGO About 54 million Americans suffer from the aches and pains of arthritis.

Treatments range from pain medications to injections to surgery.

None of it seemed to work for 77-year-old Marty Ciesielczyk.

And it jeopardized something he loved: jogging.

"For me, it's just enjoyable, and if you're not a runner, then you would have no idea what I'm talking about."

But Marty's active lifestyle was in jeopardy when knee pain took over.

"When you got to lay on the floor to get dressed, it's tough."

It happens when there's a loss of cartilage in the joint.

"It's like a tire, and as you slowly lose rubber on the tire, it wears away," explained Dr. Adam Yanke, a surgeon with Midwest Orthopaedics at Rush University.

"You might need to have the tire replaced at some point."

Marty's arthritis was too advanced for a scope procedure but not bad enough for a joint replacement.

So he enrolled in a study testing whether amniotic fluid, which surrounds a growing baby in the uterus, could help his pain.

"Amniotic products come from patients that are having healthy, elective C-sections, and they choose to donate these products at the time of the delivery," said Dr. Yanke.

It's thought to increase tissue healing and lower inflammation.

Doctor-diagnosed arthritisis more common in womenthan in men. Arthritis and other joint disorders are among the five most costly conditions among adults 18 and older.

Your bone marrow makes mesenchymal stem cells, or MSCs. They are known to grow into new tissues, including cartilage.

By gathering these cells and injecting them into the knee joint, the hope is that they will give growth to new cartilage and reduce inflammation.

Marty received a placebo during the study, but then chose to have the amniotic fluid when the study ended.

"I mean I didn't care if it was Pixie dust, as long as my knee was going to feel better."

He went from not being able to get dressed to jogging about a week after having the injection.

"This morning, I ran three, three miles, and I had no problem at all."

Amniotic fluid is also being used to treat ulcers in the eye.

Rush University will be enrolling patients for a larger follow-up study on amniotic fluid for joint pain in the future.

Clinical trialsare still going on and most studies are still early.

A review published in 2016 in BMC Musculoskeletal Disorders concluded that MSC-based therapies offer an "exciting possibility" for treatment, but further studies need to be done on how they can best be used and how well they work.

They are also known to be very expensive.

If this story has impacted your life or prompted you or someone you know to seek or change treatments, please let us know by contacting Jim Mertens atjim.mertens@wqad.comor Marjorie Bekaert Thomas atmthomas@ivanhoe.com.

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YOUR HEALTH: Knee relief can be found in the womb - WQAD.com

Bone Marrow Stem Cells Comments Off on YOUR HEALTH: Knee relief can be found in the womb – WQAD.com | February 26th, 2020

A mum has shared the emotional experience of meeting the stranger who saved her son's life.

Alfie Commons, now aged four, was just seven months old when he was diagnosed with leukaemia in 2016.

After three rounds of chemotherapy failed, Alfie received a life-saving bone marrow donation from a school teacher in Germany, who recently made the trip to the UK to meet him.

Alfie's mum, Lorna Commons, 40, of Toton has spoken about the experience in the hope it will encourage more people to sign up to become potential donors.

Looking back to the day of diagnosis, she said Alfie had been to his GP for a third time in February 2016 after suffering a cold since Christmas.

She said: "The GP told us to go to A&E for further tests as he was a little concerned.

"We got to Queen's Medical Centre in the morning and by early evening, we had the diagnosis; Alfie had infant acute lymphoblastic leukaemia (ALL)."

Ms Commons, who works in HR, added: "Even now, four years down the line, I still feel the emotions of that day. Nothing can prepare you."

The plan was to treat Alfie with chemotherapy, but after the first round failed, Ms Commons was told his only chance of survival was to get a bone marrow transplant.

The family was told Alfie was unlikely to leave hospital for the next six months.

She added: "Worse was to follow, his second course also failed and on the same day, we were told that Alfies nine-year-old brother, Billy, wasnt a bloodstemcellmatch for him either.

"The fear of losing Alfie was overwhelming, I felt helpless but I had to carry on for Alfies sake.

The transplant could not go ahead without the cancer being near enough eradicated and even when the good news came that a donor had been located, Alfie still had a mountain to climb.

After a third failed round of chemotherapy, Alfie was put on a trial immunotherapy drug as a '"last ditch attempt". Against all the odds, it worked.

"I think at that point all the doctors and nurses were preparing us for the worst. Your head has to go there," Ms Commons said.

"But then the cancer went, and it was enough to give us the bridge to getting the transplant done."

While the transplant was a success, Alfie suffered for months with Graft versus Host Disease (GvHD) on his skin and in his gut, which is the body's reaction to the new stem cells.

However, doctor's were encouraged the body was gradually accepting the cells and beginning to produce cells of their own.

On February 19, Alfie and his mum were able to meet the woman who saved his life after she made the 600-mile trip.

Christin Bouvier, 34, from Schwerin in Germany, was matched with Alfie after she registered in 2010 with DKMS, a charity dedicated to the fight against blood cancer.

The school teacher had been on the bloodstemcellregister for a number of years before she was contacted and tested as a match for Alfie.

Ms Bouvier said: When they told me that the recipient was a baby I just cried.

"Its a moment that is always with me and whenever I feel a bit down, I think back to it as it always brings me so much happiness!"

Ms Commons said she had been able to contact Ms Bouvier anonymously, as per UK law, but they were permitted to meet two years after the transplant.

Ms Bouvier added: It was always a dream to meet Lorna and Alfie and I never thought it would happen I was so delighted when Lorna invited me. I was very nervous but also very excited to meet them both in person.

"I knew the meeting would be one of those very special moments in my life."

Ms Commons feels the meeting has meant a new chapter has begun in both hers and Alfie's life and she is now focussed on the positives.

She added: "For something so small, there really is no greater gift than being a donor - I get to see my child grow up. To meet Christin, I was able to say 'this is what you've done'.

"We will be in each other's lives forever now - Alfie has her DNA in his blood. But Christin and I also share a special bond, we're just so similar and some people say we even look like sisters.

Alfie is such a special little boy and I truly believe that this story can make a real difference and save more lives.

"There is a match out there for everyone with blood cancer, people just need to come forward and register."

Anyone aged between 17-55 and in general good health can go on standby as a potential lifesaver.

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Mum meets the stem cell donor who saved her four-year-old son's life - Nottinghamshire Live

Skin Stem Cells Comments Off on Mum meets the stem cell donor who saved her four-year-old son’s life – Nottinghamshire Live | February 26th, 2020

Scientists believe that humans will soon be able to recover from injuries such as broken spines, as treatment looks to boost the body's ability to heal itself.

A new study in the journal Regenerative Medicine describes how scientists were able to stimulate the self-repair response in rats.

Rats in the study were given two drugs, one of which is usually given to bone marrow transplant patients, and another which is used for bladder control.

This cocktail caused the rats' bone marrow to produce a greater number of mesenchymal stem cells, the cells which can develop into bone tissue.

As a result, enhanced calcium binding was seen at the site of the rats' spinal injuries, speeding up the production of new bone as well as healing wounds.

The study's authors hope that one day, such treatments will work on humans.

"We know that when bones break they will heal, and this requires the activation of stem cells in the bone," study co-author Sara Rankin from the National Heart and Lung Institute at Imperial College London, said in a statement.

"However, when the damage is severe, there are limits to what the body can do of its own accord.

"We hope that by using these existing medications to mobilise stem cells, as we were able to do in rats in our new study, we could potentially call up extra numbers of these stem cells, in order to boost our bodies' own ability to mend itself and accelerate the repair process."

Both drugs tested on rats are already widely used, so researchers are hopeful human trails can begin soon.

If these trials produce the same results as those seen in rats, then it's hoped the treatment could help to not only repair spinal injuries but also speed up the rate at which broken bones heal and mend damaged tissues in other organs.

Dr Tariq Fellous, first author of the research, said: "We first need to see if these medications release the stem cells in healthy volunteers before we can test them in patients with fractures.

"We have the drugs and know they are safe to use in humans we just need the funding for the human trials."

Dr Andia Redpath, who also co-authored the paper, added that repurposing existing medicines - so-called Regenerative Pharmacology - could have major potential as an efficient and cheaper way of treating diseases.

"Rather than devising new stem cell treatments from scratch that involve lengthy and expensive trials, our approach harnesses the power of the body's own stem cells, using existing drugs.

"We already know the treatments in our study are safe, it's now just a matter of exploring further if they help our bodies heal."

Stem cells are providing incredible new medical breakthroughs all the time.

Earlier this month, scientists trialled 3D-printed skin containing stem cells to treat burns victims .

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Humans soon able to regrow spines as body given 'new power to heal itself' - Daily Star

Skin Stem Cells Comments Off on Humans soon able to regrow spines as body given ‘new power to heal itself’ – Daily Star | February 26th, 2020

For years, scientists have predicted that 3-D printingwhich has been used it to make toys, homes, scientific tools and even a plastic bunny that contained a DNA code for its own replicationcould one day be harnessed to print live, human body parts to mitigate a shortage of donor organs. So far, researchers also used 3-D printing in medicine and dentistry to create dental implants, prosthetics, and models for surgeons to practice on before they make cuts on a patient. But many researchers have moved beyond printing with plastics and metalsprinting with cells that then form living human tissues.

No one has printed fully functional, transplantable human organs just yet, but scientists are getting closer, making pieces of tissue that can be used to test drugs and designing methods to overcome the challenges of recreating the bodys complex biology.

A confocal microscopy image showing 3-Dprinted stem cells differentiating into bone cells

The first 3-D printer was developed in the late 1980s. It could print small objects designed using computer-aided design (CAD) software. A design would be virtually sliced into layers only three-thousandths of a millimeter thick. Then, the printer would piece that design into the complete product.

There were two main strategies a printer might use to lay down the pattern: it could extrude a paste through a very fine tip, printing the design starting with the bottom layer and working upward with each layer being supported by the previous layers. Alternatively, it could start with a container filled with resin and use a pointed laser to solidify portions of that resin to create a solid object from the top down, which would be lifted and removed from the surrounding resin.

When it comes to printing cells and biomaterials to make replicas of body parts and organs, these same two strategies apply, but the ability to work with biological materials in this way has required input from cell biologists, engineers, developmental biologists, materials scientists, and others.

So far, scientists have printed mini organoids and microfluidics models of tissues, also known as organs on chips. Both have yielded practical and theoretical insights into the function of the human body. Some of these models are used by pharmaceutical companies to test drugs before moving on to animal studies and eventually clinical trials. One group, for example, printed cardiac cells on a chip and connected it to a bioreactor before using it to test the cardiac toxicity of a well-known cancer drug, doxorubicin. The team showed that the cells beating rate decreased dramatically after exposure to the drug.

However, scientists have yet to construct organs that truly replicate the myriad structural characteristics and functions of human tissues. There are a number of companies who are attempting to do things like 3-D print ears, and researchers have already reported transplanting 3-D printed ears onto children who had birth defects that left their ears underdeveloped, notes Robby Bowles, a bioengineer at the University of Utah. The ear transplants are, he says, kind of the first proof of concept of 3-D printing for medicine.

THE SCIENTIST STAFF

Bowles adds that researchers are still a ways away from printing more-complex tissues and organs that can be transplanted into living organisms. But, for many scientists, thats precisely the goal. As of February 2020, more than 112,000 people in the US are waiting for an organ transplant, according to the United Network for Organ Sharing. About 20 of them die each day.

For many years, biological engineers have tried to build 3-D scaffolds that they could seed with stem cells that would eventually differentiate and grow into the shapes of organs, but to a large extent those techniques dont allow you to introduce kind of the organization of gradients and the patterning that is in the tissue, says Bowles. There is no control over where the cells go in that tissue. By contrast, 3-D printing enables researchers with to very precisely direct the placement of cellsa feat that could lead to better control over organ development.

Ideally, 3-D printed organs would be built from cells that a patients immune system could recognize as its own, to avoid immune rejection and the need for patients to take immunosuppressive drugs. Such organs could potentially be built from patient-specific induced pluripotent stem cells, but one challenge is getting the cells to differentiate into the subtype of mature cell thats needed to build a particular organ. The difficulty is kind of coming together and producing complex patternings of cells and biomaterials together to produce different functions of the different tissues and organs, says Bowles.

To imitate the patterns seen in vivo, scientists print cells into hydrogels or other environments with molecular signals and gradients designed to coax the cells into organizing themselves into lifelike organs. Scientists can use 3-D printing to build these hydrogels as well. With other techniques, the patterns achieved have typically been two-dimensional, Eben Alsberg, a bioengineer at the University of Illinois, tells The Scientist in an email. Three-dimensional bioprinting permits much more control over signal presentation in 3D.

So far, researchers have created patches of tissue that mimic portions of certain organs but havent managed to replicate the complexity or cell density of a full organ. But its possible that in some patients, even a patch would be an effective treatment. At the end of 2016, a company called Organovo announced the start of a program to develop 3-D printed liver tissue for human transplants after a study showed that transplanted patches of 3-D printed liver cells successfully engrafted in a mouse model of a genetic liver disease and boosted several biomarkers that suggested an improvement in liver function.

Only in the past few years have researchers started to make headway with one of the biggest challenges in printing 3-D organs: creating vasculature. After the patches were engrafted into the mouses liver in the Organovo study, blood was delivered to it by the surrounding liver tissue, but an entire organ would need to come prepared for blood flow.

For any cells to stay alive, [the organ] needs that blood supply, so it cant just be this huge chunk of tissue, says Courtney Gegg, a senior director of tissue engineering at Prellis Biologics, which makes and sells scaffolds to support 3-D printed tissue. Thats been recognized as one of the key issues.

Mark Skylar-Scott, a bioengineer at the Wyss Institute, says that the problem has held back tissue engineering for decades. But in 2018, Sbastian Uzel, Skylar-Scott, and a team at the Wyss Institute managed to 3-D print a tiny, beating heart ventricle complete with blood vessels. A few days after printing the tissue, Uzel says he came into the lab to find a piece of twitching tissue, which was both very terrifying and exciting.

For any cells to stay alive, [the organ] needs that blood supply, so it cant just be this huge chunk of tissue.

Courtney Gegg, Prellis Biologics

Instead of printing the veins in layers, the team used embedded printinga technique in which, instead of building from the bottom of a slide upwards, material is extruded directly into a bath, or matrix. This strategy, which allows the researchers to print free form in 3-D, says Skylar-Scott, rather having to print each layer one on top of the other to support the structure, is a more efficient way to print a vascular tree. The matrix in this case was the cellular material that made up the heart ventricle. A gelatin-like ink pushed these cells gently out of the way to create a network of channels. Once printing was finished, the combination was warmed up. This heat caused the cellular matrix to solidify, but the gelatin to liquify so it could then be rinsed out, leaving space for blood to flow through.

But that doesnt mean the problem is completely solved. The Wyss Institute teams ventricle had blood vessels, but not nearly as many as a full-sized heart. Gegg points out that to truly imitate human biology, an individual cell will have to be within 200 microns of your nearest blood supply. . . . Everything has to be very, very close. Thats far more intricate than what researchers have printed so far.

Due to hurdles with adding vasculature and many other challenges that still face 3-Dprinted tissues, laboratory-built organs wont be available for transplant anytime soon. In the meantime, 3-D printing portions of tissue is helping accelerate both basic and clinical research about the human body.

Emma Yasinski is a Florida-based freelance reporter. Follow her on Twitter@EmmaYas24.

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On the Road to 3-D Printed Organs - The Scientist

Cardiac Stem Cells Comments Off on On the Road to 3-D Printed Organs – The Scientist | February 26th, 2020