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Regenerative medicine: Mayo Clinic and collaborators develop new tool for transplanting stem cells

By Dr. Matthew Watson

PUBLIC RELEASE DATE:

16-Dec-2013

Contact: Jennifer Schutz newsbureau@mayo.edu 507-284-5005 Mayo Clinic

ROCHESTER, Minn. -- Mayo Clinic researchers and colleagues in Belgium have developed a specialized catheter for transplanting stem cells into the beating heart. The novel device includes a curved needle and graded openings along the needle shaft, allowing for increased distribution of cells. The result is maximized retention of stem cells to repair the heart. The findings appear in the journal Circulation: Cardiovascular Interventions.

"Although biotherapies are increasingly more sophisticated, the tools for delivering regenerative therapies demonstrate a limited capacity in achieving high cell retention in the heart," says Atta Behfar, M.D., Ph.D., a Mayo Clinic cardiology specialist and lead author of the study. "Retention of cells is, of course, crucial to an effective, practical therapy."

Researchers from the Mayo Clinic Center for Regenerative Medicine in Rochester and Cardio3 Biosciences in Mont-Saint-Guibert, Belgium, collaborated to develop the device, beginning with computer modeling in Belgium. Once refined, the computer-based models were tested in North America for safety and retention efficiency.

What's the significance?

This new catheter is being used in the European CHART-1 clinical trials, now underway. This is the first Phase III trial to regenerate hearts of patients who have suffered heart attack damage. The studies are the outcome of years of basic science research at Mayo Clinic and earlier clinical studies with Cardio3 BioSciences and Cardiovascular Centre in Aalst, Belgium, conducted between 2009 and 2010.

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The development of the catheter and subsequent studies were supported by Cardio3 BioSciences; Walloon Region General Directorate for Economy, Employment & Research; Meijer Lavino Foundation for Cardiac Research Aalst (Belgium); the National Institutes of Health; Grainger Foundation; Florida Heart Research Institute; Marriott Heart Disease Research Program; and the Mayo Clinic Center for Regenerative Medicine.

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Studies: Stem cells reverse heart damage – CNN.com

By Dr. Matthew Watson

A new study says heart damage may be reversible with stem cell therapy without dangerous side effects.

STORY HIGHLIGHTS

(CNN) -- On a June day in 2009, a 39-year-old man named Ken Milles lay on an exam table at Cedars-Sinai Medical Center in Los Angeles. A month earlier, he'd suffered a massive heart attack that destroyed nearly a third of his heart.

"The most difficult part was the uncertainty," he recalls. "Your heart is 30% damaged, and they tell you this could affect you the rest of your life." He was about to receive an infusion of stem cells, grown from cells taken from his own heart a few weeks earlier. No one had ever tried this before.

About three weeks later, in Kentucky, a patient named Mike Jones underwent a similar procedure at the University of Louisville's Jewish Hospital. Jones suffered from advanced heart failure, the result of a heart attack years earlier. Like Milles, he received an infusion of stem cells, grown from his own heart tissue.

"Once you reach this stage of heart disease, you don't get better," says Dr. Robert Bolli, who oversaw Jones' procedure, explaining what doctors have always believed and taught. "You can go down slowly, or go down quickly, but you're going to go down."

Conventional wisdom took a hit Monday, as Bolli's group and a team from Cedars-Sinai each reported that stem cell therapies were able to reverse heart damage, without dangerous side effects, at least in a small group of patients.

In Bolli's study, published in The Lancet, 16 patients with severe heart failure received a purified batch of cardiac stem cells. Within a year, their heart function markedly improved. The heart's pumping ability can be quantified through the "Left Ventricle Ejection Fraction," a measure of how much blood the heart pumps with each contraction. A patient with an LVEF of less than 40% is considered to suffer severe heart failure. When the study began, Bolli's patients had an average LVEF of 30.3%. Four months after receiving stem cells, it was 38.5%. Among seven patients who were followed for a full year, it improved to an astounding 42.5%. A control group of seven patients, given nothing but standard maintenance medications, showed no improvement at all.

"We were surprised by the magnitude of improvement," says Bolli, who says traditional therapies, such as placing a stent to physically widen the patient's artery, typically make a smaller difference. Prior to treatment, Mike Jones couldn't walk to the restroom without stopping for breath, says Bolli. "Now he can drive a tractor on his farm, even play basketball with his grandchildren. His life was transformed."

At Cedars-Sinai, 17 patients, including Milles, were given stem cells approximately six weeks after suffering a moderate to major heart attack. All had lost enough tissue to put them "at big risk" of future heart failure, according to Dr. Eduardo Marban, the director of the Cedars-Sinai Heart Institute, who developed the stem cell procedure used there.

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More Efficient Way to Grow Heart Muscle from Stem Cells Could Yield New Regenerative Therapies

By JoanneRUSSELL25

Durham, NC (PRWEB) December 09, 2013

Generating new cardiac muscle from human embryonic stem cells (hESCs) and/or induced pluripotent stem cells (iPSC) could fulfill the demand for therapeutic applications and drug testing. The production of a similar population of these cells remains a major limitation, but in a study just published in STEM CELLS Translational Medicine, researchers now believe they have found a way to do this.

By combining small molecules and growth factors, the international research team led by investigators at the Cardiovascular Research Center at Icahn School of Medicine at Mount Sinai developed a two-step system that caused stem cells to differentiate into ventricular heart muscle cells from hESCs and iPSCs. The process resulted in high efficiency and reproducibility, in a manner that mimicked the developmental steps of normal cardiovascular development.

These chemically induced, ventricular-like cardiomyocytes (termed ciVCMs) exhibited the expected cardiac electrophysiological and calcium handling properties as well as the appropriate heart rate responses, said lead investigator Ioannis Karakikes, Ph.D., of the Stanford University School Of Medicine, Cardiovascular Institute. Other members of the team included scientists from the Icahn School of Medicine at Mount Sinai, New York, and the Stem Cell & Regenerative Medicine Consortium at the University of Hong Kong.

In addition, using an integrated approach involving computational and experimental systems, the researchers demonstrated that using molecules to modulate the Wnt pathway, which passes signals from cell to cell, plays a key role in whether a cell evolves into an atrial or ventricular muscle cell.

The further clarification of the molecular mechanism(s) that underlie this kind of subtype specification is essential to improving our understanding of cardiovascular development. We may be able to regulate the commitment, proliferation and differentiation of pluripotent stem cells into heart muscle cells and then harness them for therapeutic purposes, Dr. Karakikes said.

"Most cases of heart failure are related to a deficiency of heart muscle cells in the lower chambers of the heart, said said Anthony Atala, MD, editor of STEM CELLS Translational Medicine and director of the Wake Forest Institute for Regenerative Medicine. An efficient, cost-effective and reproducible system for generating ventricular cardiomyocytes would be a valuable resource for cell therapies as well as drug screening.

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The full article, Small Molecule-Mediated Directed Differentiation of Human Embryonic Stem Cells Toward Ventricular Cardiomyocytes, can be accessed at http://www.stemcellstm.com.

About STEM CELLS Translational Medicine: STEM CELLS TRANSLATIONAL MEDICINE (SCTM), published by AlphaMed Press, is a monthly peer-reviewed publication dedicated to significantly advancing the clinical utilization of stem cell molecular and cellular biology. By bridging stem cell research and clinical trials, SCTM will help move applications of these critical investigations closer to accepted best practices.

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More Efficient Way to Grow Heart Muscle from Stem Cells Could Yield New Regenerative Therapies

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Space Station made accessible for stem cell research

By LizaAVILA

Washington, Dec 7 : NASA and the Center for the Advancement of Science in Space (CASIS) are enabling research aboard the International Space Station that could lead to new stem cell-based therapies for medical conditions faced on Earth and in space.

Scientists will take advantage of the space station's microgravity environment to study the properties of non-embryonic stem cells.

NASA is interested in space-based cell research because it is seeking ways to combat the negative health effects astronauts face in microgravity, including bone loss and muscle atrophy.

Mitigation techniques are necessary to allow humans to push the boundaries of space exploration far into the solar system. This knowledge could help people on Earth, particularly the elderly, who are afflicted with similar conditions.

Two stem cell investigations scheduled to fly to the space station next year were highlighted Friday, Dec. 6, at the World Stem Cell Summit in San Diego.

Lee Hood, a member of the CASIS Board of Directors, moderated a panel session in which scientists Mary Kearns-Jonker of Loma Linda University in California and Roland Kaunas of Texas A&M University discussed their planned research, which will gauge the impact of microgravity on fundamental stem cell properties.

Kearns-Jonker's research will study the aging of neonatal and adult cardiac stem cells in microgravity with the ultimate goal of improving cardiac cell therapy.

Kaunas is a part of a team of researchers developing a system for co-culturing and analyzing stem cells mixed with bone tumor cells in microgravity.

This system will allow researchers to identify potential molecular targets for drugs specific to certain types of cancer.

Stem cells are cells that have not yet become specialized in their functions. They display a remarkable ability to give rise to a spectrum of cell types and ensure life-long tissue rejuvenation and regeneration.

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What are the potential uses of human stem cells and the …

By Dr. Matthew Watson

Introduction: What are stem cells, and why are they important? What are the unique properties of all stem cells? What are embryonic stem cells? What are adult stem cells? What are the similarities and differences between embryonic and adult stem cells? What are induced pluripotent stem cells? What are the potential uses of human stem cells and the obstacles that must be overcome before these potential uses will be realized? Where can I get more information? VII. What are the potential uses of human stem cells and the obstacles that must be overcome before these potential uses will be realized?

There are many ways in which human stem cells can be used in research and the clinic. Studies of human embryonic stem cells will yield information about the complex events that occur during human development. A primary goal of this work is to identify how undifferentiated stem cells become the differentiated cells that form the tissues and organs. Scientists know that turning genes on and off is central to this process. Some of the most serious medical conditions, such as cancer and birth defects, are due to abnormal cell division and differentiation. A more complete understanding of the genetic and molecular controls of these processes may yield information about how such diseases arise and suggest new strategies for therapy. Predictably controlling cell proliferation and differentiation requires additional basic research on the molecular and genetic signals that regulate cell division and specialization. While recent developments with iPS cells suggest some of the specific factors that may be involved, techniques must be devised to introduce these factors safely into the cells and control the processes that are induced by these factors.

Human stem cells could also be used to test new drugs. For example, new medications could be tested for safety on differentiated cells generated from human pluripotent cell lines. Other kinds of cell lines are already used in this way. Cancer cell lines, for example, are used to screen potential anti-tumor drugs. The availability of pluripotent stem cells would allow drug testing in a wider range of cell types. However, to screen drugs effectively, the conditions must be identical when comparing different drugs. Therefore, scientists will have to be able to precisely control the differentiation of stem cells into the specific cell type on which drugs will be tested. Current knowledge of the signals controlling differentiation falls short of being able to mimic these conditions precisely to generate pure populations of differentiated cells for each drug being tested.

Perhaps the most important potential application of human stem cells is the generation of cells and tissues that could be used for cell-based therapies. Today, donated organs and tissues are often used to replace ailing or destroyed tissue, but the need for transplantable tissues and organs far outweighs the available supply. Stem cells, directed to differentiate into specific cell types, offer the possibility of a renewable source of replacement cells and tissues to treat diseases including Alzheimer's diseases, spinal cord injury, stroke, burns, heart disease, diabetes, osteoarthritis, and rheumatoid arthritis.

Figure 3. Strategies to repair heart muscle with adult stem cells. Click here for larger image.

2001 Terese Winslow

For example, it may become possible to generate healthy heart muscle cells in the laboratory and then transplant those cells into patients with chronic heart disease. Preliminary research in mice and other animals indicates that bone marrow stromal cells, transplanted into a damaged heart, can have beneficial effects. Whether these cells can generate heart muscle cells or stimulate the growth of new blood vessels that repopulate the heart tissue, or help via some other mechanism is actively under investigation. For example, injected cells may accomplish repair by secreting growth factors, rather than actually incorporating into the heart. Promising results from animal studies have served as the basis for a small number of exploratory studies in humans (for discussion, see call-out box, "Can Stem Cells Mend a Broken Heart?"). Other recent studies in cell culture systems indicate that it may be possible to direct the differentiation of embryonic stem cells or adult bone marrow cells into heart muscle cells (Figure 3).

Cardiovascular disease (CVD), which includes hypertension, coronary heart disease, stroke, and congestive heart failure, has ranked as the number one cause of death in the United States every year since 1900 except 1918, when the nation struggled with an influenza epidemic. Nearly 2600 Americans die of CVD each day, roughly one person every 34 seconds. Given the aging of the population and the relatively dramatic recent increases in the prevalence of cardiovascular risk factors such as obesity and type 2 diabetes, CVD will be a significant health concern well into the 21st century.

Cardiovascular disease can deprive heart tissue of oxygen, thereby killing cardiac muscle cells (cardiomyocytes). This loss triggers a cascade of detrimental events, including formation of scar tissue, an overload of blood flow and pressure capacity, the overstretching of viable cardiac cells attempting to sustain cardiac output, leading to heart failure, and eventual death. Restoring damaged heart muscle tissue, through repair or regeneration, is therefore a potentially new strategy to treat heart failure.

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Heart Stem Cell Trial: Interview With Researcher Roberto Bolli, MD

By JoanneRUSSELL25

An interview with Roberto Bolli, MD.

University of Louisville cardiologist Roberto Bolli, MD, led the stem cell study that tested using patients' own heart stem cells to help their hearts recover from heart failure. Though that trial was preliminary, the results look promising -- and may one day lead to a cure for heart failure.

Here, Bolli talks about what this work means and when it might become an option for patients.

2012 WebMD, LLC. All rights reserved.

"Realistically, this will not come... for another three or four years, at least," Bolli says. "It may be longer, depending on the results of the next trial, of course."

Larger studies are needed to confirm the procedure's safety and effectiveness. If those succeed, it could be "the biggest advance in cardiovascular medicine in my lifetime," Bolli says.

A total of 20 patients took part in the initial study.

All of them experienced significant improvement in their heart failure and now function better in daily life, according to Bolli. "The patients can do more, there's more ability to exercise, and the quality of life improves markedly," Bolli says.

Bolli's team published its findings on how the patients were doing one year after stem cell treatment in November 2011 in the Lancet, a British medical journal.

Each patient was infused with about 1 million of his or her own cardiac stem cells, which could eventually produce an estimated 4 trillion new cardiac cells, Bolli says. His team plans to follow each patient for two years after their stem cell procedure.

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Howard Leonhardt of Leonhardt Ventures to Present at World …

By Sykes24Tracey

SAN DIEGO, Dec. 5, 2013 /PRNewswire/ -- Howard Leonhardt of Leonhardt Ventures and the Cal-X Stars Innovation and Business Accelerator team will present at the 2013 World Stem Cell Summit on Friday, December 6, 2013 at the Manchester Grand Hyatt in San Diego in two sessions.

2pm Harbor Room AB - Developing Combination Products Cells, Genes, Devices

3pm Harbor Room DE - Startup Considerations for Stem Cell Companies - Getting Funding and Avoiding Pitfalls

Cal-X Stars Business Accelerator, Inc.is an innovation accelerator with an unprecedented portfolio of breakthrough cardiovascular life science and high social good impact innovations that have primarily been majority funded to date by Leonhardt Ventures and its associated angel investor network.

The innovation laboratory and business accelerator has two clearfocusareas:

Management team and board have a proven track record in leading breakthrough innovations in these focused spaces -http://www.calstockexchange.com/team-cal-x/.

Cardiovascular portfolio technologies include...

MyoStim Pacershttp:/www.myostimpacers.com- heart failure pacemaker designed to recruit reparative stem cells to damaged and weakened heart tissue.

Bioheart, Inc.http://www.bioheartinc.com- Phase III leader in applying adult muscle stem cells to treat advanced heart failuresince 1999.Only cell type known to grow new contractile muscle in the depths of heart scar tissue. In the Phase II/III MARVEL randomized, double blinded, placebo controlled study Bioheart's MyoCell achieved 95.7 meters improvement in exercise capacity over placebo (minus 4 meters).

BioPace biological pacemaker made entirely of living cells.

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Priming ‘cocktail’ shows promise as cardiac stem cell grafting …

By NEVAGiles23

PUBLIC RELEASE DATE:

5-Dec-2013

Contact: Jennifer Nachbur jennifer.nachbur@uvm.edu 802-656-7875 University of Vermont

New research by University of Vermont Associate Professor of Medicine Jeffrey Spees, Ph.D., and colleagues has identified a new tool that could help facilitate future stem cell therapy for the more than 700,000 Americans who suffer a heart attack each year. The study appeared online in Stem Cells Express.

Stem cells, which can come from embryos, fetal tissue and adult tissues, have the potential to develop into a variety of cell types in the body, such as muscle cells, brain cells and red blood cells. These cells also possess the ability to repair human tissues. The field of regenerative medicine which explores the viability of using embryonic, fetal and adult stem cells to repair and regenerate tissues and organs has struggled to successfully graft cells from culture back into injured tissue.

"Many grafts simply didn't take; the cells wouldn't stick or would die," explains Spees. So he and his research team set out to develop ways to enhance graft success.

They focused on a type of bone marrow-derived progenitor cell that forms stromal cells. Stromal cells form connective tissue and also support the creation of blood cells. The researchers were aware of that these cells secrete a diverse array of molecules called ligands that protect injured tissue, promote tissue repair and support stem and progenitor cells in culture. Different ligands interact with specific receptors on the surface of a stem or progenitor cell, transmitting signals that can instruct the cell to adhere, to divide, or to differentiate into a mature functional cell.

To confirm whether or not these types of ligands would protect a cardiac progenitor cell and help it graft, the group isolated a conditioned medium from human bone marrow-derived progenitor cells. They found that the medium contained Connective Tissue Growth Factor (CTGF) and the hormone insulin.

"Both CTGF and insulin are protective," says Spees. "Together, they have a synergistic effect."

In the study, Spees and colleagues compared the impact of sending a cardiac stem cell "naked" into a rodent heart with infarction (heart attack) to a cell that instead wore a "backpack" of protective ligands, created by incubating about 125,000 cardiac cells in a "cocktail" of CTGF and insulin on ice for 30 minutes. The team grafted the cells sub-epicardially between the outer layer and the muscle tissue of the heart and found that their priming cocktail resulted in improved graft success.

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Research | Research news | 2013 | The heart’s own stem cells …

By LizaAVILA

The hearts own stem cells play their part in regeneration

Sca1 stem cells replace steadily ageing heart muscle cells

November 28, 2013

Up until a few years ago, the common school of thought held that the mammalian heart had very little regenerative capacity. However, scientists now know that heart muscle cells constantly regenerate, albeit at a very low rate. Researchers at the Max Planck Institute for Heart and Lung Research in Bad Nauheim, have identified a stem cell population responsible for this regeneration. Hopes are growing that it will be possible in future to stimulate the self-healing powers of patients with diseases and disorders of the heart muscle, and thus develop new potential treatments.

Stem cells play a part in heart regeneration. This image of the fluorescence microscope depicts a section of the heart tissue of a mouse. The green colouring of the cells in the middle shows that the cell originated from a so-called Sca1 stem cell.

MPI for Heart and Lung Research

MPI for Heart and Lung Research

Some vertebrates seem to have found the fountain of youth, the source of eternal youth, at least when it comes to their heart. In many amphibians and fish, for example, this important organ has a marked capacity for regeneration and self-healing. Some species in the two animal groups have even perfected this capability and can completely repair damage caused to heart tissue, thus maintaining the organs full functionality.

The situation is different for mammals, whose hearts have a very low regenerative capacity. According to the common school of thought that has prevailed until recently, the reason for this deficit is that the heart muscle cells in mammals cease dividing shortly after birth. It was also assumed that the mammalian heart did not have any stem cells that could be used to form new heart muscle cells. On the contrary: new studies show that aged muscle cells are also replaced in mammalian hearts. Experts estimate, however, that between just one and four percent of heart muscle cells are replaced every year.

Scientists in Thomas Brauns Research Group at the Max Planck Institute for Heart and Lung Research have succeeded in identifying a stem cell population in mice that plays a key role in this regeneration of heart muscle cells. Experiments conducted by the researchers in Bad Nauheim on genetically modified mice show that the Sca1 stem cells in a healthy heart are involved in the ongoing replacement of heart muscle cells. The Sca-1 cells increase their activity if the heart is damaged, with the result that significantly more new heart muscle cells are formed.

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New transformation: Human stem cells into functional lung …

By Sykes24Tracey

In computer-based text processing and digital typesetting, a non-breaking space, no-break space or non-breakable space (NBSP) is a variant of the space character that prevents an automatic line break (line wrap) at its position. In certain formats (such as HTML), it also prevents the collapsing of multiple consecutive whitespace characters into a single space. The non-breaking space is also known as a hard space or fixed space. In Unicode, it is encoded at U+00A0 no-break space (HTML:    ).

Text-processing software typically assumes that an automatic line break may be inserted anywhere a space character occurs; a non-breaking space prevents this from happening (provided the software recognizes the character). For example, if the text 100 km will not quite fit at the end of a line, the software may insert a line break between 100 and km. To avoid this undesirable behaviour, the editor may choose to use a non-breaking space between 100 and km. This guarantees that the text 100km will not be broken: if it does not fit at the end of a line it is moved in its entirety to the next line.

A second common application of non-breaking spaces is in plain text file formats such as SGML, HTML, TeX, and LaTeX, which sometimes treat sequences of whitespace characters (space, newline, tab, form feed, etc.) as if they were a single white-space character. Such collapsing of white-space allows the author to neatly arrange the source text using line breaks, indentation and other forms of spacing without affecting the final typeset result.[1][2]

In contrast, non-breaking spaces are not merged with neighboring whitespace characters, and can therefore be used by an author to insert additional visible space in the formatted text. For example, in HTML, non-breaking spaces may be used in conjunction with a fixed-width font to create tabular alignment (courier new font family used):

Column 1Column 2 ---------------- 1.22.3

(note that the use of the pre tag, the whitespace:pre CSS rule, or a table are alternative, if not necessarily better, ways to achieve the same result in HTML)

If ordinary spaces are used instead then the spaces are collapsed when the HTML is rendered and the layout is broken:

Column 1 Column 2 -------- -------- 1.2 2.3

Non-breaking space can also be used to automatically change formatting in a document. This is useful for things like class plans and recipe files where the description of a cell or line may be different from the actual text or title.

Unicode defines several other non-break space characters[3] that differ from the regular space in width:

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Research | Research news | 2012 | Finished heart switches stem …

By LizaAVILA

Finished heart switches stem cells off

Transcription factor Ajuba regulates stem cell activity in the heart during embryonic development

July 12, 2012

It is not unusual for babies to be born with congenital heart defects. This is because the development of the heart in the embryo is a process which is not only extremely complex, but also error-prone. Scientists from the Max Planck Institute for Heart and Lung Research in Bad Nauheim have now identified a key molecule that plays a central role in regulating the function of stem cells in the heart. As a result, not only could congenital heart defects be avoided in future, but new ways of stimulating the regeneration of damaged hearts in adults may be opened up.

Cardiac development out of control: Absence of the transcription factor Ajuba during cardiac development, as is the case in the right-hand photo due to genetic intervention, disrupts development of the heart in the fish embryo. In addition to an increased number of cardiac muscle cells (green with red-stained nuclei), the heart is additionally deformed during development.

Max Planck Institute for Heart and Lung Research

Max Planck Institute for Heart and Lung Research

It's a long road from a cluster of cells to a finished heart. Cell division transforms what starts out as a collection of only a few cardiac stem cells into an ever-larger structure from which the various parts of the heart, such as ventricles, atria, valves and coronary vessels, develop. This involves the stem and precursor cells undergoing a complex process which, in addition to tightly regulated cell division, also includes cell migration, differentiation and specialisation. Once the heart is complete, the stem cells are finally switched off.

Scientists from the Max Planck Institute for Heart and Lung Research in Bad Nauheim have now discovered how major parts of this development process are regulated. Their search initially focused on finding binding partners for transcription factor Isl1. Isl1 is characteristic of a specific group of cardiac stem cells which are consequently also known as Isl1+ cells. During their search, the researchers came across Ajuba, a transcription factor from the group of LIM proteins. "We then took a closer a look at the interaction between these two molecules and came to the conclusion that Ajuba must be an important switch", says Gergana Dobreva, head of the "Origin of Cardiac Cell Lineages" Research Group at the Bad Nauheim-based Max Planck Institute.

Using an animal model, the scientists then investigated the effects of a defective switch on cardiac development. Embryonic development can be investigated particularly effectively in the zebrafish. The Bad Nauheim-based researchers therefore produced a genetically modified fish that lacked a functioning Ajuba protein. Cardiac development in these fishes was in fact severely disrupted. In addition to deformation of the heart, caused by twisting of the cardiac axis, what particularly struck the researchers was a difference in size in comparison with control animals. "In almost all the investigated fish we observed a dramatic enlargement of the heart. If Ajuba is absent, there is clearly no other switch that finally silences the Isl1-controlled part of cardiac development", says Dobreva.

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[International version] Linda van Laake: "We want to work together to improve stem cell treatment" – Video

By Sykes24Tracey


[International version] Linda van Laake: "We want to work together to improve stem cell treatment"
Dr Linda van Laake is assistant professor and specialist registrar in Cardiology at the University Medical Center Utrecht and Hubrecht Institute. She carries...

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6. Mending a Broken Heart: Stem Cells and Cardiac Repair [Stem …

By Sykes24Tracey

Charles A. Goldthwaite, Jr., Ph.D.

Cardiovascular disease (CVD), which includes hypertension, coronary heart disease (CHD), stroke, and congestive heart failure (CHF), has ranked as the number one cause of death in the United States every year since 1900 except 1918, when the nation struggled with an influenza epidemic.1 In 2002, CVD claimed roughly as many lives as cancer, chronic lower respiratory diseases, accidents, diabetes mellitus, influenza, and pneumonia combined. According to data from the 19992002 National Health and Nutrition Examination Survey (NHANES), CVD caused approximately 1.4 million deaths (38.0 percent of all deaths) in the U.S. in 2002. Nearly 2600 Americans die of CVD each day, roughly one death every 34 seconds. Moreover, within a year of diagnosis, one in five patients with CHF will die. CVD also creates a growing economic burden; the total health care cost of CVD in 2005 was estimated at $393.5 billion dollars.

Given the aging of the U.S. population and the relatively dramatic recent increases in the prevalence of cardiovascular risk factors such as obesity and type 2 diabetes,2,3 CVD will continue to be a significant health concern well into the 21st century. However, improvements in the acute treatment of heart attacks and an increasing arsenal of drugs have facilitated survival. In the U.S. alone, an estimated 7.1 million people have survived a heart attack, while 4.9 million live with CHF.1 These trends suggest an unmet need for therapies to regenerate or repair damaged cardiac tissue.

Ischemic heart failure occurs when cardiac tissue is deprived of oxygen. When the ischemic insult is severe enough to cause the loss of critical amounts of cardiac muscle cells (cardiomyocytes), this loss initiates a cascade of detrimental events, including formation of a non-contractile scar, ventricular wall thinning (see Figure 6.1), an overload of blood flow and pressure, ventricular remodeling (the overstretching of viable cardiac cells to sustain cardiac output), heart failure, and eventual death.4 Restoring damaged heart muscle tissue, through repair or regeneration, therefore represents a fundamental mechanistic strategy to treat heart failure. However, endogenous repair mechanisms, including the proliferation of cardiomyocytes under conditions of severe blood vessel stress or vessel formation and tissue generation via the migration of bone-marrow-derived stem cells to the site of damage, are in themselves insufficient to restore lost heart muscle tissue (myocardium) or cardiac function.5 Current pharmacologic interventions for heart disease, including beta-blockers, diuretics, and angiotensin-converting enzyme (ACE) inhibitors, and surgical treatment options, such as changing the shape of the left ventricle and implanting assistive devices such as pacemakers or defibrillators, do not restore function to damaged tissue. Moreover, while implantation of mechanical ventricular assist devices can provide long-term improvement in heart function, complications such as infection and blood clots remain problematic.6 Although heart transplantation offers a viable option to replace damaged myocardium in selected individuals, organ availability and transplant rejection complications limit the widespread practical use of this approach.

Figure 6.1. Normal vs. Infarcted Heart. The left ventricle has a thick muscular wall, shown in cross-section in A. After a myocardial infarction (heart attack), heart muscle cells in the left ventricle are deprived of oxygen and die (B), eventually causing the ventricular wall to become thinner (C).

2007 Terese Winslow

The difficulty in regenerating damaged myocardial tissue has led researchers to explore the application of embryonic and adult-derived stem cells for cardiac repair. A number of stem cell types, including embryonic stem (ES) cells, cardiac stem cells that naturally reside within the heart, myoblasts (muscle stem cells), adult bone marrow-derived cells, mesenchymal cells (bone marrow-derived cells that give rise to tissues such as muscle, bone, tendons, ligaments, and adipose tissue), endothelial progenitor cells (cells that give rise to the endothelium, the interior lining of blood vessels), and umbilical cord blood cells, have been investigated to varying extents as possible sources for regenerating damaged myocardium. All have been tested in mouse or rat models, and some have been tested in large animal models such as pigs. Preliminary clinical data for many of these cell types have also been gathered in selected patient populations.

However, clinical trials to date using stem cells to repair damaged cardiac tissue vary in terms of the condition being treated, the method of cell delivery, and the primary outcome measured by the study, thus hampering direct comparisons between trials.7 Some patients who have received stem cells for myocardial repair have reduced cardiac blood flow (myocardial ischemia), while others have more pronounced congestive heart failure and still others are recovering from heart attacks. In some cases, the patient's underlying condition influences the way that the stem cells are delivered to his/her heart (see the section, quot;Methods of Cell Deliveryquot; for details). Even among patients undergoing comparable procedures, the clinical study design can affect the reporting of results. Some studies have focused on safety issues and adverse effects of the transplantation procedures; others have assessed improvements in ventricular function or the delivery of arterial blood. Furthermore, no published trial has directly compared two or more stem cell types, and the transplanted cells may be autologous (i.e., derived from the person on whom they are used) or allogeneic (i.e., originating from another person) in origin. Finally, most of these trials use unlabeled cells, making it difficult for investigators to follow the cells' course through the body after transplantation (see the section quot;Considerations for Using These Stem Cells in the Clinical Settingquot; at the end of this article for more details).

Despite the relative infancy of this field, initial results from the application of stem cells to restore cardiac function have been promising. This article will review the research supporting each of the aforementioned cell types as potential source materials for myocardial regeneration and will conclude with a discussion of general issues that relate to their clinical application.

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6. Mending a Broken Heart: Stem Cells and Cardiac Repair [Stem ...

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The heart’s own stem cells play their part in regeneration

By raymumme

Nov. 28, 2013 Up until a few years ago, the common school of thought held that the mammalian heart had very little regenerative capacity. However, scientists now know that heart muscle cells constantly regenerate, albeit at a very low rate. Researchers at the Max Planck Institute for Heart and Lung Research in Bad Nauheim, have identified a stem cell population responsible for this regeneration. Hopes are growing that it will be possible in future to stimulate the self-healing powers of patients with diseases and disorders of the heart muscle, and thus develop new potential treatments.

Some vertebrates seem to have found the fountain of youth, the source of eternal youth, at least when it comes to their heart. In many amphibians and fish, for example, this important organ has a marked capacity for regeneration and self-healing. Some species in the two animal groups have even perfected this capability and can completely repair damage caused to heart tissue, thus maintaining the organ's full functionality.

The situation is different for mammals, whose hearts have a very low regenerative capacity. According to the common school of thought that has prevailed until recently, the reason for this deficit is that the heart muscle cells in mammals cease dividing shortly after birth. It was also assumed that the mammalian heart did not have any stem cells that could be used to form new heart muscle cells. On the contrary: new studies show that aged muscle cells are also replaced in mammalian hearts. Experts estimate, however, that between just one and four percent of heart muscle cells are replaced every year.

Scientists in Thomas Braun's Research Group at the Max Planck Institute for Heart and Lung Research have succeeded in identifying a stem cell population in mice that plays a key role in this regeneration of heart muscle cells. Experiments conducted by the researchers in Bad Nauheim on genetically modified mice show that the Sca1 stem cells in a healthy heart are involved in the ongoing replacement of heart muscle cells. The Sca-1 cells increase their activity if the heart is damaged, with the result that significantly more new heart muscle cells are formed.

Since, in comparison to the large amount of heart muscle cells, Sca-1 stem cells account for just a tiny proportion of the cells in the heart muscle, searching for them is like searching for a needle in a haystack. "We also faced the problem that Sca-1 is no longer available in the cells as a marker protein for stem cells after they have been changed into heart muscle cells. To prove this, we had to be inventive," says project leader Shizuka Uchida. The Max Planck researchers genetically modified the stem cells to such an extent that, in addition to the Sca-1, they produced another visible marker. Even if Sca-1 was subsequently no longer visible, the marker could still be detected permanently.

"In this way, we were able to establish that the proportion of heart muscle cells originating from Sca-1 stem cells increased continuously in healthy mice. Around five percent of the heart muscle cells regenerated themselves within 18 months," says Uchida. Moreover, mice suffering from heart disease triggered by the experiment had up to three times more of these newly formed heart muscle cells.

"The data shows that, in principle, the mammalian heart is able to trigger regeneration and renewal processes. Under normal circumstances, however, these processes are not enough to ultimately repair cardiac damage," says Braun. The aim is to find ways in which the formation of new heart muscle cells from heart stem cells can be improved and thereby strengthen the heart's self-healing powers.

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Cardiac regeneration: current therapies—future concepts

By JoanneRUSSELL25

Abstract

Cardiovascular disease (CVD) continues to be one of the main causes of death in the western world. A high burden of disease and the high costs for the healthcare systems claim for novel therapeutic strategies besides current conventional medical care. One decade ago first clinical trials addressed stem cell based therapies as a potential alternative therapeutic strategy for myocardial regeneration and repair. Besides bone marrow derived stem cells (BMCs), adult stem cells from adipose or cardiac tissue have been used in current clinical studies with inconsistent results. Although outcomes in terms of safety and feasibility are generally encouraging, functional improvements were mostly disappointingly low and have failed to reach expectations. In the future, new concepts for myocardial regeneration, especially concerning recovery of cardiomyocyte loss, have to be developed. Transplantation of novel stem or progenitor cell populations with true regenerative potential, direct reprogramming of scar tissue into functional myocardium, tissue engineering or stimulation of endogenous cardiac repair by pharmacological agents are conceivable. This review summarizes current evidence of stem cell based regenerative therapies and discusses future strategies to improve functional outcomes.

KEYWORDS : Myocardial infarction, regenerative medicine, stem cells, tissue engineering, reprogramming

In 2009 cardiovascular disease (CVD) still accounted for 32.3% of all deaths in the United States and therefore continues to be one of the main causes of death (1). From 1999 to 2009, the rate of death due to CVD has declined, but nevertheless the burden of disease remains high. Although improved medical care and acute management of myocardial infarction have led to a considerable reduction of early mortality rate survivors are susceptible to an increased prevalence of chronic heart failure as they develop scarring followed by ventricular remodeling despite optimum medical care (2,3).

Interestingly, cardiovascular operations and interventional procedures increased by 28% from 2000 to 2010 implicating an enormous cost factor for the healthcare system (1). For 2009, it was estimated that the direct and indirect costs of CVD and stroke add up to about $312.6 billion in the United States, which was more than for any other diagnostic group (1).

The main issue of current pharmacological, interventional or operative therapies is their disability to compensate the irreversible loss of functional cardiomyocytes (4). Hence, the future challenge of cardiovascular therapies will be the functional regeneration of myocardial contractility by novel concepts, like cell based therapy, tissue engineering or reprogramming of scar fibroblasts (5,6).

After promising preclinical results using adult stem and precursor cells for cardiac regeneration a rapid clinical translation using autologous bone marrow cells (BMCs) in patients was initiated (7,8). In the last few years numerous clinical trials addressing the transplantation of various adult stem cell populations for cardiac regeneration have been performed. Essential characteristics for the selected adult stem cell populations are the potential to proliferate, migrate and the ability to transdifferentiate into various mature cell types (9). Today, different adult stem cell sources like BMCs, myocardium or adipose tissue derived cells were already used in clinical trials. Beside direct intracoronary or intramyocardial transplantation of adult stem cells into the heart mobilization of autologous progenitor cells by administration of different cytokines [i.e., erythropoietin (EPO) or granulocyte colony stimulating factor (G-CSF)] were also evaluated in first clinical trials (summarized in and ,).

Regenerative therapies and cell sources currently administered in clinical trials. Current clinical trials use BMCs, ADRCs or CPCs to regenerate impaired myocardium after ischemic events. Alternatively cytokines like EPO or G-CSF are employed to mobilize ...

Transplantation of adult stem cells-clinical trials mentioned in the text.

Mobilization of adult stem cells-clinical trials mentioned in the text.

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Endogenous cardiac stem cell – Wikipedia, the free encyclopedia

By Dr. Matthew Watson

Endogenous cardiac stem cells (eCSCs) are tissue-specific stem progenitor cells harboured within the adult mammalian heart.

They were first discovered in 2003 by Bernardo Nadal-Ginard, Piero Anversa and colleagues [1][2] in the adult rat heart and since then have been identified and isolated from mouse, dog, porcine and human hearts.[3][4]

The adult heart was previously thought to be a post mitotic organ without any regenerative capability. The identification of eCSCs has provided an explanation for the hitherto unexplained existence of a subpopulation of immature cycling myocytes in the adult myocardium. Indeed, recent evidence from a genetic fate-mapping study established that stem cells replenish adult mammalian cardiomyocytes lost by cardiac wear and tear and injury throughout the adult life.[5] Moreover, it is now accepted that myocyte death and myocyte renewal are the two sides of the proverbial coin of cardiac homeostasis in which the eCSCs play a central role.[6] These findings produced a paradigm shift in cardiac biology and opened new opportunities and approaches for future treatment of cardiac diseases by placing the heart squarely amongst other organs with regenerative potential such as the liver, skin, muscle, CNS. However, they have not changed the well-established fact that the working myocardium is mainly constituted of terminally differentiated contractile myocytes. This fact does not exclude, but is it fully compatible with the heart being endowed with a robust intrinsic regenerative capacity which resides in the presence of the eCSCs throughout the individual lifespan.

Briefly, eCSCs have been first identified through the expression of c-kit, the receptor of the stem cell factor and the absence of common hematopoietic markers, like CD45. Afterwards, different membrane markers (Sca-1, Abcg-2, Flk-1) and transcription factors (Isl-1, Nkx2.5, GATA4) have been employed to identify and characterize these cells in the embryonic and adult life.[7] eCSCs are clonogenic, self renewing and multipotent in vitro and in vivo,[8] capable of generating the 3 major cell types of the myocardium: myocytes, smooth muscle and endothelial vascular cells.[9] They express several markers of stemness (i.e. Oct3/4, Bmi-1, Nanog) and have significant regenerative potential in vivo.[10] When cloned in suspension they form cardiospheres,[11] which when cultured in a myogenic differentiation medium, attach and differentiate into beating cardiomyocytes.

In 2012, it was proposed that Isl-1 is not a marker for endogenous cardiac stem cells.[12] That same year, a different group demonstrated that Isl-1 is not restricted to second heart field progenitors in the developing heart, but also labels cardiac neural crest.[13] It has also been reported that Flk-1 is not a specific marker for endogenous and mouse ESC-derived Isl1+ CPCs. While some eCSC discoveries have been brought into question, there has been success with other membrane markers. For instance, it was demonstrated that the combination of Flt1+/Flt4+ membrane markers identifies an Isl1+/Nkx2.5+ cell population in the developing heart. It was also shown that endogenous Flt1+/Flt4+ cells could be expanded in vitro and displayed trilineage differentiation potential. Flt1+/Flt4+ CPCs derived from iPSCs were shown to engraft into the adult myocardium and robustly differentiate into cardiomyocytes with phenotypic and electrophysiologic characteristics of adult cardiomyocytes.[14]

With the myocardium now recognized as a tissue with limited regenerating potential,[15] harbouring eCSCs that can be isolated and amplified in vitro [16] for regenerative protocols of cell transplantation or stimulated to replicate and differentiate in situ in response to growth factors,[17] it has become reasonable to exploit this endogenous regenerative potential to replace lost/damaged cardiac muscle with autologous functional myocardium.

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Human neural stem cells could meet the clinical problem of …

By Dr. Matthew Watson

New research has shown human neural stem cells could improve blood flow in critical limb ischemia through the growth of new vessels. Critical limb ischemia (CLI) is a disease that severely obstructs arteries and reduces the blood flow to legs and feet. CLI remains an unmet clinical problem and with an ageing population and the rise in type II diabetes, the incidence of CLI is expected to increase.

The study, led by academics in the University of Bristol's School of Clinical Sciences, is published online in the American Heart Association journal Arteriosclerosis, Thrombosis, and Vascular Biology.

Current stem cell therapy trials for the treatment of CLI have revitalised new hope for improving symptoms and prolonging life expectancy. However, there are limitations on the use of autologous cell therapy. The patient's own stem cells are generally invasively harvested from bone marrow or require purification from peripheral blood after cytokine stimulation. Other sources contain so few stem cells that ex vivo expansion through lengthy bespoke Good Manufacturing Practice processes is required. Ultimately, these approaches lead to cells of variable quality and potency that are affected by the patient's age and disease status and lead to inconsistent therapeutic outcomes.

In order to circumvent the problem a team, led by Professor Paolo Madeddu in the Bristol Heart Institute at the University of Bristol, has used a conditionally immortalised clonal human neural stem cell (hNSC) line to treat animal models with limb ischaemia and superimposed diabetes. The CTX cell line, established by stem cell company ReNeuron, is genetically modified to produce genetically and phenotypically stable cell banks.

Results of the new study have shown that CTX treatment effectively improves the recovery from ischaemia through the promotion of the growth of new vessels. The safety of CTX cell treatment is currently being assessed in disabled patients with stroke [PISCES trial, NCT01151124]. As a result, the same cell product is immediately available for starting dose ranging safety and efficacy studies in CLI patients.

Professor Paolo Madeddu, Chair of Experimental Cardiovascular Medicine and Head of Regenerative Medicine Section in the Bristol Heart Institute at the University of Bristol, said: "Currently, there are no effective drug interventions to treat CLI. The consequences are a very poor quality of life, possible major amputation and a life expectancy of less than one year from diagnosis in 50 per cent of all CLI patients.

"Our findings have shown a remarkable advancement towards more effective treatments for CLI and we have also demonstrated the importance of collaborations between universities and industry that can have a social and medical impact."

Dr John Sinden, Chief Scientific Officer of ReNeuron, added: "The novel idea of using neural stem cells to treat vascular disease arose from a chance discussion with Professor Madeddu. The discussion led to a short pilot study with our cells producing very clear data, which then developed into a further eight experiments exploring different variants of the disease model, the product formulation and dose variation.

"The study also explored the cascade of molecular events that produced vascular and muscle recovery. It is a great example of industry and academia working successfully towards the key goal, clinical translation."

Explore further: UH Case Medical Center launches novel clinical trial using stem cells to prevent amputation

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Cardiac Stem Cells (CSCs) | University of Maryland Medical Center

By JoanneRUSSELL25

For immediate release: September 10, 2012

Baltimore, MD --Researchers at the University of Maryland School of Medicine, who are exploring novel ways to treat serious heart problems in children, have conducted the first direct comparison of the regenerative abilities of neonatal and adult-derived human cardiac stem cells. Among their findings: cardiac stem cells (CSCs) from newborns have a three-fold ability to restore heart function to nearly normal levels compared with adult CSCs. Further, in animal models of heart attack, hearts treated with neonatal stem cells pumped stronger than those given adult cells. The study is published in the September 11, 2012, issue of Circulation.

The surprising finding is that the cells from neonates are extremely regenerative and perform better than adult stem cells, says the study's senor author, Sunjay Kaushal, M.D., Ph.D., associate professor of surgery at the University of Maryland School of Medicine and director, pediatric cardiac surgery at the University of Maryland Medical Center. We are extremely excited and hopeful that this new cell-based therapy can play an important role in the treatment of children with congenital heart disease, many of whom don't have other options.

Dr. Kaushal envisions cellular therapy as either a stand-alone therapy for children with heart failure or an adjunct to medical and surgical treatments. While surgery can provide structural relief for some patients with congenital heart disease and medicine can boost heart function up to two percent, he says cellular therapy may improve heart function even more dramatically. We're looking at this type of therapy to improve heart function in children by 10, 12, or 15 percent. This will be a quantum leap in heart function improvement.

Heart failure in children, as in adults, has been on the rise in the past decade and the prognosis for patients hospitalized with heart failure remains poor. In contrast to adults, Dr. Kaushal says heart failure in children is typically the result of a constellation of problems: reduced cardiac blood flow; weakening and enlargement of the heart; and various congenital malformations. Recent research has shown that several types of cardiac stem cells can help the heart repair itself, essentially reversing the theory that a broken heart cannot be mended.

Stem cells are unspecialized cells that can become tissue- or organ-specific cells with a particular function. In a process called differentiation, cardiac stem cells may develop into rhythmically contracting muscle cells, smooth muscle cells or endothelial cells. Stem cells in the heart may also secrete growth factors conducive to forming heart muscle and keeping the muscle from dying.

To conduct the study, researchers obtained a small amount of heart tissue during normal cardiac surgery from 43 neonates and 13 adults. The cells were expanded in a growth medium yielding millions of cells. The researchers developed a consistent way to isolate and grow neonatal stem cells from as little as 20 milligrams of heart tissue. Adult and neonate stem cell activity was observed both in the laboratory and in animal models. In addition, the animal models were compared to controls that were not given the stem cells.

Dr. Kaushal says it is not clear why the neonatal stem cells performed so well. One explanation hinges on sheer numbers: there are many more stem cells in a baby's heart than in the adult heart. Another explanation: neonate-derived cells release more growth factors that trigger blood vessel development and/or preservation than adult cells.

This research provides an important link in our quest to understand how stem cells function and how they can best be applied to cure disease and correct medical deficiencies, says E. Albert Reece, M.D., Ph.D., M.B.A., vice president for medical affairs, University of Maryland; the John Z. and Akiko K. Bowers Distinguished Professor; and dean, University of Maryland School of Medicine. Sometimes simple science is the best science. In this case, a basic, comparative study has revealed in stark terms the powerful regenerative qualities of neonatal cardiac stem cells, heretofore unknown.

Insights gained through this research may provide new treatment options for a life-threatening congenital heart syndrome called hypoplastic left heart syndrome (HLHS). Dr. Kaushal and his team will soon begin the first clinical trial in the United States to determine whether the damage to hearts of babies with HLHS can be reversed with stem cell therapy. HLHS limits the heart's ability to pump blood from the left side of the heart to the body. Current treatment options include either a heart transplant or a series of reconstructive surgical procedures. Nevertheless, only 50-60 percent of children who have had those procedures survive to age five.

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Stem cell therapy: When will it help the heart? | The Why Files

By JoanneRUSSELL25

Stem cells: When will they heal the heart?

Its been 15 years since a University of Wisconsin-Madison researcher isolated embryonic stem cells the do-anything cells that appear in early development. Its been six years since adult human cells were transformed into the related induced pluripotent stem cells.

ENLARGE

Some day, stem cell therapy could restore cells, save hearts, and avoid the need for some heart transplants, such as this one. This heart is ready for its new home.

And yet the early hope to grow spare parts turning stem cells into specialized cells for repairing a failing brain, pancreas or heart, remains mostly promise rather than reality.

Researchers have since found how to transform stem cells into a wide variety of body cells, including heart muscle cells, or cardiomyocytes. But the holy Grail tissue supplementation or replacement remains tantalizingly out of reach.

Last week, Why Files attended a symposium on treating cardiovascular disease with stem cells, at the BioPharmaceutical Technology Center Institute near Madison, Wis. We found the picture unexpectedly complicated: as multiple kinds of stem cells are grown and delivered in a bewildering variety of ways to treat a catalog of conditions.

So far, stem cells have not been approved to treat any heart disease in the United States.

Still, the need remains clear. Disorders of the heart and blood vessels, which deliver oxygen and nutrients to the body, continue to kill. Today, one of every 2.6 Americans will die of some cause related to their heart, writes Columbia University Medical Center.

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Stem cell therapy: When will it help the heart? | The Why Files

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UCLA doctors test stem-cell therapy to improve blood flow in …

By daniellenierenberg

Marty Greenfield with UCLA doctors

Marty Greenfield lives with crushing pain every day due to angina, a condition that is caused by an inadequate supply of blood to the heart. He has suffered a heart attack, and a coronary bypass procedure and angioplasty have provided little relief. His doctor referred him to UCLA to be considered for a heart transplant.

Dr. Jonathan Tobis, a UCLA clinical professor of cardiology, performed an angiogram and angioplasty on Greenfield, 64, but found that the patient was not a candidate for a heart transplant because his heart muscle function was still good.

Instead, Tobis suggested that Greenfield consider participating in a Phase 3 clinical trial that uses a patient's own blood-derived stem cells to try to restore circulation to the heart. The procedure uses the latest technology to map the heart in 3-D and guides the doctor to deliver the stem-cell injections to targeted sites in the heart muscle.

On Oct. 17, Greenfield became the first patient at UCLA to participate in the multicenter clinical trial. He said he jumped at the chance to help, even though the study is double blind, which means that neither the patients nor the researchers know who is receiving stem-cell injections and who is receiving placebos.

"This just isn't about me," said Greenfield, a married father of two sons who lives near Las Vegas. "If I can help move this research forward so that it helps just one person, it will be worth it."

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