Stimulus-Triggered Acquisition of Pluripotency Cells: Embryonic-Like Stem Cells Without Killing Embryos or Genetic Engineering


Embryonic stem cells have been the gold standard for pluripotent stem cells. Pluripotent means capable of differentiating into one of many cell types in the adult body. Ever since James Thomson isolated the first human embryonic stem cell lines in 1998, scientists have dreamed of using embryonic stem cells to treat diseases in human patients.

However, deriving human embryonic stem cell lines requires the destruction or molestation of a human embryo, the smallest, youngest, and most vulnerable member of our community. In 2006, Shinya Yamanaka and his colleges used genetic engineering techniques to make induced pluripotent stem (iPS) cells, which are very similar to embryonic stem cells in many ways. Unfortunately, the derivation of iPSCs introduces mutations into the cells.

Now, researchers from Brigham and Women’s Hospital (BWH), in Boston, in collaboration with the RIKEN Center for Developmental Biology in Japan, have demonstrated that any mature adult cell has the potential to be converted into the equivalent of an embryonic stem cell. Published in the January 30, 2014 issue of the journal Nature, this research team demonstrated in a preclinical model, a novel and unique way to reprogram cells. They called this phenomenon stimulus-triggered acquisition of pluripotency (STAP). Importantly, this process does not require the introduction of new outside DNA, which is required for the reprogramming process that produces iPSCs.

“It may not be necessary to create an embryo to acquire embryonic stem cells. Our research findings demonstrate that creation of an autologous pluripotent stem cell – a stem cell from an individual that has the potential to be used for a therapeutic purpose – without an embryo, is possible. The fate of adult cells can be drastically converted by exposing mature cells to an external stress or injury. This finding has the potential to reduce the need to utilize both embryonic stem cells and DNA-manipulated iPS cells,” said senior author Charles Vacanti, MD, chairman of the Department of Anesthesiology, Perioperative and Pain Medicine and Director of the Laboratory for Tissue Engineering and Regenerative Medicine at BWH and senior author of the study. “This study would not have been possible without the significant international collaboration between BWH and the RIKEN Center,” he added.

The inspiration for this research was an observation in plant cells – the ability of a plant callus, which is made by an injured plant, to grow into a new plant. These relatively dated observations led Vacanti and his collaborators to suggest that any mature adult cell, once differentiated into a specific cell type, could be reprogrammed and de-differentiated through a natural process that does not require inserting genetic material into the cells.

“Could simple injury cause mature, adult cells to turn into stem cells that could in turn develop into any cell type?” hypothesized the Vacanti brothers.

Vacanti and others used cultured, mature adult cells. After stressing the cells almost to the point of death by exposing them to various stressful environments including trauma, a low oxygen and acidic environments, researchers discovered that within a period of only a few days, the cells survived and recovered from the stressful stimulus by naturally reverting into a state that is equivalent to an embryonic stem cell. With the proper culture conditions, those embryonic-like stem cells were propagated and when exposed to external stimuli, they were then able to redifferentiate and mature into any type of cell and grow into any type of tissue.

To examine the growth potential of these STAP cells, Vacanti and his team used mature blood cells from mice that had been genetically engineered to glow green under a specific wavelength of light. They stressed these cells from the blood by exposing them to acid, and found that in the days following the stress, these cells reverted back to an embryonic stem cell-like state. These stem cells then began growing in spherical clusters (like plant callus tissue). The cell clusters were introduced into developing mouse embryos that came from mice that did not glow green. These embryos now contained a mixture of cells (a “chimera”). The implanted clusters were able to differentiate into green-glowing tissues that were distributed in all organs tested, confirming that the implanted cells are pluripotent.

Thus, external stress might activate unknown cellular functions that set mature adult cells free from their current commitment to a particular cell fate and permit them to revert to their naïve cell state.

“Our findings suggest that somehow, through part of a natural repair process, mature cells turn off some of the epigenetic controls that inhibit expression of certain nuclear genes that result in differentiation,” said Vacanti.

Of course, the next step is to explore this process in more sophisticated mammals, and, ultimately in humans.

“If we can work out the mechanisms by which differentiation states are maintained and lost, it could open up a wide range of possibilities for new research and applications using living cells. But for me the most interesting questions will be the ones that let us gain a deeper understanding of the basic principles at work in these phenomena,” said first author Haruko Obokata, PhD.

If human cells can be made into embryonic stem cells by a similar process, then someday, a simple skin biopsy or blood sample might provide the material to generate embryonic stem cells that are specific to each individual, without the need for genetic engineering or killing the smallest among us. This truly creates endless possibilities for therapeutic options.

Injected microparticles shown to greatly reduce heart attack damage


Reducing inflammation in the heart after a heart by injecting biodegradable microparticles.

Leaders in Pharmaceutical Business Intelligence (LPBI) Group

See on Scoop.itCardiovascular and vascular imaging

After a heart attack has occurred, inflammatory cells known as monocytes rush to the damaged tissue. This causes the heart to swell, further damaging the ti…

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Human Stem Cell Gene Therapy Appears Safe and Effective


Two recent studies in the journal Science have reported the outcome of virally-mediated gene correction in hematopoietic stem cells (HSCs) to treat human patients. These studies may usher in a new era of safe and effective gene therapy. These exciting new clinical findings both come from the laboratory of Luigi Naldini at the San Raffaele Scientific Institute, Milan, Italy. The first experiment examined the treatment of metachromatic leukodystrophy (MLD), which is caused by mutations in the arylsulfatase A (ARSA) gene, and the second, investigated treatments for Wiskott-Aldrich syndrome (WAS), which is caused by mutations in the gene that encodes WASP.

MLD is one of several diseases that affects the lysosome; a structure in cells that acts as the garbage disposal of the cell. So called “lysosomal storage diseases” result from the inability of cells to degrade molecules that come to the lysosome for degradation. Without the ability to degrade these molecules, they build up to toxic levels and produce progressive motor and cognitive impairment and death within a few years of the onset of symptoms.

To treat MLD, workers in Naldini’s laboratory isolated blood-making stem cells from the bone marrow of three pre-symptomatic MLD patients (MLD01, 02 and 03). These stem cells were infected with genetically engineered viruses that encoded the human ARSA gene. After expanding these stem cells in culture, they were re-introduced into the MLD patients after those same patients had their resident bone marrow wiped out. The expression of the ARSA gene in the reconstituted bone marrow was greater than 10 fold the levels measured in healthy controls and there were no signs of blood cancers or other maladies. One month after the transplant, the implanted cells showed very high-level and stable engraftment. Between 45%-80% of cells isolated and grown from bone marrow samples harbored the fixed ARSA gene. AS expected, the levels of the ARSA protein rose to above-normal levels in therapeutically relevant blood cells and above normal levels of ARSA protein were isolated from hematopoietic cells after one month and cerebrospinal fluid (CSF) one to two years after transfusion. This is remarkable when you consider that one year before, no ARSA was seen. This shows that the implanted cells and their progeny properly homed to the right places in the body. The patient evaluations at time points beyond the expected age of disease onset was even more exciting, since these treat patients showed normal, continuous motor and cognitive development compared to their siblings who had MLD, but were untreated. The sibling of the patient designated “MLD01” was wheelchair-bound and unable to support their head and trunk at 39 months, but excitingly, after treatment, patient MLD01 was able to stand, walk and run at 39 months of age and showed signs of continuous motor and cognitive development. Lastly, and perhaps most importantly, there was no evidence of implanted cells becoming cancerous, even though they underwent self-renewal, like all good stem cells. This is the first report of an MLD patient at 39 months displaying such positive clinical features.

The second study treated WAS, which is an inherited disease that affects the immune system and leads to infections, abnormal platelets, scaly skin (eczema), blood tumors, and autoimmunity. In this second study, blood-making stem cells were collected from three patients infected with genetically engineered viruses that expressed the WASP gene. These stem cells were then reinfused intravenously (~11 million cells ) three days after collection. Blood tests and bone marrow biopsies showed evidence of robust engraftment of gene-corrected cells in bone marrow and peripheral blood up to 30 months later. WASP expression increased with time in most blood cells. Although serious adverse infectious events occurred in two patients, overall clinical improvement resulted in reduced disease severities in all patients. None of the three patients demonstrated signs of blood cancers and the platelet counts rose, but, unfortunately, not to normal levels. Again, no evidence for adverse effects were observed.

Simply put, these authors have presented a strategy for ex vivo gene correction in HSCs for inherited disorders which works and appears safe in comparison to previous strategies. Long-term analyses will undoubtedly need to be intensely scrutinized, but this research surely represents a huge step forward in the safe treatment of these and similar genetic disorders.

Stem Cell-based Baldness Cure One Step Closer


Scientists might be able to offer people with less that optimal amounts of hair new hope when it comes to reversing baldness. Researchers from the University of Pennsylvania say they’ve moved closer to using stem cells to treat thinning hair — at least in mice.

This group said that the use of stem cells to regenerate missing or dying hair follicles is considered a potential way to reverse hair loss. However, the technology did not exist to generate adequate numbers of hair-follicle-generating stem cells.

But new findings indicate that this may now be achievable. “This is the first time anyone has made scalable amounts of epithelial stem cells that are capable of generating the epithelial component of hair follicles,” Dr. Xiaowei Xu, an associate professor of dermatology at Penn’s Perelman School of Medicine, said in a university news release.

According to Xu, those cells have many potential applications that extend to wound healing, cosmetics and hair regeneration.

In their new study, Xu’s team converted induced pluripotent stem cells (iPSCs) – reprogrammed adult stem cells with many of the characteristics of embryonic stem cells – into epithelial stem cells. This is the first time this has been done in either mice or people.

The epithelial stem cells were mixed with certain other cells and implanted into mice. They produced the outermost layers of the skin and hair follicles that are similar to human hair follicles. This study was published in the Jan. 28 edition of the journal Nature Communications.

This suggests that these cells might eventually help regenerate hair in people.

Xu said this achievement with iPSC-derived epithelial stem cells does not mean that a treatment for baldness is around the corner. Hair follicles contain both epithelial cells and a second type of adult cells called dermal papillae.

“When a person loses hair, they lose both types of cells,” Xu said. “We have solved one major problem — the epithelial component of the hair follicle. We need to figure out a way to also make new dermal papillae cells, and no one has figured that part out yet.”

Experts also note that studies conducted in animals often fail when tested in humans.

Vascular Progenitors Made from Induced Pluripotent Stem Cells Repair Blood Vessels in the Eye Regardless of the Site of Injection


Johns Hopkins University medical researchers have reported the derivation of human induced-pluripotent stem cells (iPSCs) that can repair damaged retinal vascular tissue in mice. These stem cells, which were derived from human umbilical cord-blood cells and reprogrammed into an embryonic-like state, were derived without the conventional use of viruses, which can damage genes and initiate cancers. This safer method of growing the cells has drawn increased support among scientists, they say, and paves the way for a stem cell bank of cord-blood derived iPSCs to advance regenerative medical research.

In a report published Jan. 20 in the journal Circulation, Johns Hopkins University stem cell biologist Elias Zambidis and his colleagues described laboratory experiments with these non-viral, human retinal iPSCs, that were created generated using the virus-free method Zambidis first reported in 2011.

“We began with stem cells taken from cord-blood, which have fewer acquired mutations and little, if any, epigenetic memory, which cells accumulate as time goes on,” says Zambidis, associate professor of oncology and pediatrics at the Johns Hopkins Institute for Cell Engineering and the Kimmel Cancer Center. The scientists converted these cells to a status last experienced when they were part of six-day-old embryos.

Instead of using viruses to deliver a gene package to the cells to turn on processes that convert the cells back to stem cell states, Zambidis and his team used plasmids, which are rings of DNA that replicate briefly inside cells and then are degraded and disappear.

Next, the scientists identified and isolated high-quality, multipotent, vascular stem cells that resulted from the differentiation of these iPSC that can differentiate into the types of blood vessel-rich tissues that can repair retinas and other human tissues as well. They identified these cells by looking for cell surface proteins called CD31 and CD146. Zambidis says that they were able to create twice as many well-functioning vascular stem cells as compared with iPSCs made with other methods, and, “more importantly these cells engrafted and integrated into functioning blood vessels in damaged mouse retina.”

Working with Gerard Lutty, Ph.D., and his team at Johns Hopkins’ Wilmer Eye Institute, Zambidis’ team injected these newly iPSC-derived vascular progenitors into mice with damaged retinas (the light-sensitive part of the eyeball). The cells were injected into the eye, the sinus cavity near the eye or into a tail vein. When Zamdibis and his colleagues took images of the mouse retinas, they found that the iPSC-derived vascular progenitors, regardless of injection location, engrafted and repaired blood vessel structures in the retina.

“The blood vessels enlarged like a balloon in each of the locations where the iPSCs engrafted,” says Zambidis. Their vascular progenitors made from cord blood-derived iPSCs compared very well with the ability of vascular progenitors derived from fibroblast-derived iPSCs to repair retinal damage.

Zambidis says that he has plans to conduct additional experiments in diabetic rats, whose conditions more closely resemble human vascular damage to the retina than the mouse model used for the current study, he says.

With mounting requests from other laboratories, Zambidis says he frequently shares his cord blood-derived iPSC with other scientists. “The popular belief that iPSCs therapies need to be specific to individual patients may not be the case,” says Zambidis. He points to recent success of partially matched bone marrow transplants in humans, shown to be as effective as fully matched transplants.

“Support is growing for building a large bank of iPSCs that scientists around the world can access,” says Zambidis, although large resources and intense quality-control would be needed for such a feat. However, Japanese scientists led by stem-cell pioneer Shinya Yamanaka are doing exactly that, he says, creating a bank of stem cells derived from cord-blood samples from Japanese blood banks.

Synthetic Matrices that Induce Stem Cell-Mediated Bone Formation


Biomimetic matrices resemble living structures even though they are made from synthetic materials. Researchers in the laboratory of Shyni Varghese at the UC San Diego Jacobs School of Engineering have used calcium phosphate to direct mesenchymal stem cells to form bone. In doing so, Varghese and his colleagues have identified a surprising pathway from biomaterials to bone.

Varghese and his colleagues think that their work may point out new targets for treating bone defects, such as major fractures, and bone metabolic disorders such as osteoporosis.

The first goal of this research was to use materials to build something that looked like bone. This way, stem cells harvested from bone marrow (the squishy stuff inside our bones) could sense the presence of bone and differentiate into osteoblasts, the cells in our bodies that build bone.

“We knew for years that calcium phosphate-based materials promote osteogenic differentiation of stem cells, but none of use knew why.” said Varghese. “As engineers, we want to build something that is reproducible and consistent, so we need to know how building factors contribute to this end.”

Varghese and co-workers discovered that phosphate ions dissolved from calcium phosphate-based materials and these stray phosphate ions are taken up by the stem cells and used for the production of adenosine triphosphate or ATP. ATP is the energy currency of the cell, and it is the way cells store energy in a form that is readily usable for powering other reactions.

In stem cells, the generation of ATP eventually increases the intracellular concentration of the ATP breakdown product adenosine, and adenosine signals to stem cells to differentiate into osteoblasts and make bone.

Varghese said that she was surprised that “the biomaterials were connected to metabolic pathways. And we didn’t know how these metabolic pathways could influence stem cells,” and their commitment to bone formation.

These results also explain another clinical observation. Plastic surgeons have been using fat-based stem cells for eyelid lifts, breast augmentation, and other types of reconstructive surgeries. In once case, a plastic surgeon injected a dermal filler that contained calcium hydroxyapatite with the fat-based stem cells into a woman’s eyelid to provide an eye lift. However, the stem cells formed bone, and the poor lady’s lid painfully clicked every time she blinked and she had to have surgery to remove the ectopic bone. These results from Varghese’s laboratory explains why these fat-based stem cells formed bone in this case, and great care should be taken to never use such fillers in fat-based transplantation procedures.

Micro-Grooved Surfaces Influence Stem Cell Differentiation


Martin Knight and his colleagues from the Queen Mary’s School of Engineering and Materials Science and the Institute of Bioengineering in London, UK have shown that growing adult stem cells on micro-grooved surfaces disrupts a particular biochemical pathway that specified the length of a cellular structure called the “primary cilium.” Disruption of the primary cilium ultimately controls the subsequent behavior of these stem cells.

Primary cilia are about one thousand times narrower than a human hair. They are found in most cells and even though they were thought to be irrelevant at one time, this is clearly not the case.

Primary Cilium

The primary cilium acts as a sensory structure that responds to mechanical and chemical stimuli in the environment, and then communicates that external signal to the interior of the cell.  Most of the basic research on this structure was done using a single-celled alga called Chlamydomonas.

Martin Knight and his team, however, are certain that primary cilia in adult stem cells play a definite role in controlling cell differentiation.  Knight said, “Our research shows that they [primary cilia] play a key role in stem cell differentiation.  We found it’s possible to control stem cell specialization by manipulating primary cilia elongation, and that this occurs when stem cells are grown on these special grooved surfaces.”

When mesenchymal stromal cells were grown on grooved surfaces, the tension inside the cells was altered, and this remodeled the cytoskeleton of the cells.  Cytoskeleton refers to a rigid group of protein inside of cells that act as “rebar.” for the cell.  If you have ever worked with concrete, you will know that structural use of concrete requires the use of reinforcing metal bars to prevent the concrete from crumbling under the force of its own weight.  In the same way, cytoskeletal proteins reinforce the cell, give it shape, help it move, and help it resist shear forces.  Remodeling of the cytoskeleton can greatly change the behavior of the cell.

The primary cilium is important for stem cell differentiation.  Growing mesenchymal stromal cells on micro-grooved surfaces disrupts the primary cilium and prevents stem cell differentiation.  This simple culture technique can help maintain stem cells in an undifferentiated state until they have expanded enough for therapeutic purposes.

Once again we that there are ways to milk adult stem cells for all they are worth.  Destroying embryos is simply not necessary to save the lives of patients.

Frozen Stem Cells Taken from a Cadaver Five Years Ago Vigorously Grow


It is incumbent upon regenerative medicine researchers to discover non-controversial sources of stem cells that are safe and abundant. To that end, harvesting stem cells from deceased donors might represent an innovative and potentially unlimited reservoir of different stem cells.

In this present study, tissues from the blood vessels of cadavers were used as a source of human cadaver mesenchymal stromal/stem cells (hC-MSCs). The scientists in this paper successfully isolated cells from arteries after the death of the patient and subjected them to cryogenic storage in a tissue-banking facility for at least 5 years.

After thawing, the hC-MSCs were re-isolated with high-efficiency (12 × 10[6]) and showed all the usual characteristics of mesenchymal stromal cells. They expressed all the proper markers, were able to differentiate into the right cell types, and showed the same immunosuppressive activity as mesenchymal stromal cells from living persons.

Thus the efficient procurement of stem cells from cadavers demonstrates that such cells can survive harsh conditions, low oxygen tensions, and freezing and dehydration. This paves the way for a scientific revolution where cadaver stromal/stem cells could effectively treat patients who need cell therapies.

See Sabrina Valente, and others, Human cadaver multipotent stromal/stem cells isolated from arteries stored in liquid nitrogen for 5 years.  Stem Cell Research & Therapy 2014, 5:8.

Controlling Transplanted Stem Cells from the Inside Out


Scientists have worked very hard to understand how to control stem cell differentiation.  However, despite how well you direct stem cell behavior in culture, once those stem cells have been transplanted, they will often do as they wish.  Sometimes, transplanted stem cells surprise people.

Several publications describe stem cells that, once transplanted undergo “heterotropic differentiation.” Heterotropic differentiation refers to tissues that form in the wrong place. For example, one lab found that transplantation of mesenchymal stem cells into mouse hearts after a heart attack produced bone (don’t believe me – see Martin Breitbach and others, “Potential risks of bone marrow cell transplantation into infarcted hearts.” Blood 2007 110:1362-1369).  Bone in the heart – that can’t be good. Therefore, new ways to control the differentiation of cells once they have been transplanted are a desirable goal for stem cell research.

From this motivation comes a weird but wonderful paper from Jeffrey Karp and James Ankrum of Brigham and Women’s Hospital and MIT, respectively, that loads stem cells with microparticles that give the transplanted stem cell continuous cues that tell them how to behave over the course of days or weeks as the particles degrade.

“Regardless of where the cell in the body, it’s going to be receiving its cues from the inside,” said Karp. “This is a completely different strategy than the current method of placing cells onto drug-doped microcarriers or scaffolds, which is limiting because the cells need to remain in close proximity to those materials in order to function. Also these types of materials are too large to be infused into the bloodstream.”

Controlling cells in culture is relatively easy. If cells take up the right molecules, they will change their behavior. This level of control, however, is lost after the cell is transplanted. Sometimes implanted cells readily respond to the environment within the body,. but other times, their behavior is erratic and unpredictable. Karp’s strategy, which her called “particle engineering,” corrects this problem by turning cells into pre-programmable units. The internalized particles stably remain inside the transplanted cell and instruct it precisely how to act. It can direct cells to release anti-inflammatory factors, or regenerate lost tissue and heal lesions or wounds.

“Once those particles are internalized into the cells, which can take on the order of 6-24 hours, we can deliver the transplant immediately or even cryopreserve the cells,” said Karp. “When the cells are thawed at the patient’s bedside, they can be administrated and the agents will start to be released inside the cells to control differentiation, immune modulation or matrix production, for example.”

It could take more than a decade for this type of cell therapy to be a common medical practice, but to speed up the pace of this research, Karp published the study to encourage others in the scientific community to apply the technique to their various fields. Karp’s paper also illustrates the range of different cell types that can be controlled by particle engineering, including stem cells, cells of the immune system, and pancreatic cells.

“With this versatile platform, which leveraged Harvard and MIT experts in drug delivery, cell engineering, and biology, we’ve demonstrated the ability to track cells in the body, control stem cell differentiation, and even change the way cells interact with immune cells, said Ankrum, who is a former graduate student in Karp’s laboratory. “We’re excited to see what applications other researchers will imagine using this platform.”

Skin-Based Stem Cells Repair Peripheral Nerves


Italian scientists from Milan have used skin-derived stem cells in combination with a previously developed collagen tube to successfully bridge the gaps in injured nerves in a rat model, On the strength of that animal model system, the Italian group successfully used this procedure to heal the damaged peripheral nerves in the upper arms of a patient whose only other option was limb amputation.

“Peripheral nerve repair with satisfactory functional remains a great surgical challenge, especially for severe nerve injuries resulting in extended nerve defects,” said the corresponding author of this study Dr, Yvan Torrente of the Department of Pathophysiology and Transplantation at the University of Milan. “However, we hypothesized that the combination of autologous (self donated) stem cells placed in collagen tubes to bridge gaps in the damaged nerves would restore the continuity of injured nerves and save from amputation the upper arms of a patient with poly-injury to motor and sensory nerves.”

Although autologous nerve grafting has been the “gold standard” for reconstructive surgeries, these researchers recognized the disadvantages of such a procedure. Graft availability is the first drawback of autologous nerve grafting. Secondly, the condition of the donor site or “donor site morbidity.” If the donor site is in bad shape, taking a nerve from that site will probably make the donor site worse and provide a nerve that does not work as well. Finally, neuropathic pain is also an issue.

Autologous skin-derived stem cells have several advantages over autologous nerve grafts. First, the skin provides an accessible source of stem cells that are rapidly expandable in culture. Secondly, these skin-derived cells are capable of survival and integration within host tissues.

The NeuraGen nerve guide is a tiny collagen tube that connects the two damaged ends of a nerve together to mediate and expedite nerve healing.  NeuraGen tubes guide the transplanted stem cells to the gaps in the damaged nerves.  Torrente and his co-workers developed and tested the NeuraGen tubes in rats, and the US Food and Drug Administration (FDA) has approved NeuraGen for use in human patients.  See this figure from the NeuraGen web site:

NeuraGen_Open

 

Torrente and others successfully used skin-derived stem cells and NeuraGen tubes to heal the severed sciatic nerves in rats.  Therefore, once the FDA approved NeuraGen tubes, Torrente tried NeuraGen tubes in human patients with severe peripheral nerve damage.

A three-year follow-up on one particular patient showed that injured median and ulnar nerves showed extensive healing as ascertained by magnetic resonance imaging.  Functional tests, such as pinch gauge tests, static two-point discrimination and monofilament touch tests established the functional recovery of these peripheral nerves in the patient.

“Our three-year follow-up has witnesses nerve regeneration with suitable functional recovery in the patient and the salvage of upper arms from amputation,” said researchers from Torrente’s group.  “This finding opens an alternative avenue for patients who are at-risk of amputation after the injury to important nerves.”

Pluristem stem cell trial to treat muscle injury meets main goal | Reuters


Pluristem’s placental PLX-PAD cells produced show statistically significant improvement over placebo in the change of the maximal contraction force of the gluteal muscle

Pluristem Therapeutics Inc., a leading developer of placenta-based cell therapies, has announced results from its Phase I/II clinical trial that examines the safety and efficacy of PLacental eXpanded (PLX-PAD) cells in the treatment of muscle injury.  This trial showed patients treated with PLX-PAD had a greater improved change of maximal voluntary muscle contraction force than the placebo group.  These results show that PLX cells may have the ability to improve muscle and tendon healing after orthopedic injuries.

This Phase I/II trial was a randomized, placebo-controlled, double-blinded study conducted at the Orthopedic Clinic of the Charité University Medical School under the oversight of the Paul-Ehrlich-Institute (PEI), Germany’s health authority.  The injured muscle studied was the gluteus medius muscle in the buttock. Total hip replacement surgery via the standard transgluteal approach necessitates injury of the gluteus medius muscle, and post-operative healing is crucial for joint stability and function.

The 20 patients in the study were randomized into three treatment groups. Each patient received an injection in the traumatized gluteal muscle that had been injured during hip replacement surgery.  One group was treated with 150 million PLX-PAD cells per dose (n=7), the second was administered 300 million PLX-PAD cells per dose (n=6), and the third received placebo (n=7).

The primary safety endpoint was clearly met, since patients showed no serious adverse events reported at either dose level.  In this study, PLX-PAD cells were safe and well tolerated.

The primary efficacy endpoint of the study was the change in maximal voluntary isometric contraction force of the gluteal muscle at six months after the surgery.  Efficacy was shown in both PLX-PAD treated patient groups, but the patients in the group who had received the 150 million cell dose displayed a statistically significant 500% improvement over the placebo group in the change of the maximal contraction force of the gluteal muscle (p=0.0067).   Patients treated at the 300 million cell dose showed a 300% improvement over the placebo (p=0.18).

The structure of the gluteal muscle was also evaluated by means of magnetic resonance imaging (MRI).  MRI analysis revealed an increase in muscle volume in those patients treated with PLX-PAD cells versus the placebo group.  This efficacy endpoint was demonstrated in both PLX-PAD treated patient groups, with the group receiving the 150 million cell dose displaying a statistically significant superiority over the placebo group.  Patients treated at the 150 million cell dose showed an approximate 300% improvement over the placebo in the analysis of muscle volume (p=0.004).  Patients treated at the 300 million cell dose showed an approximate 150% improvement over the placebo in the change of muscle volume (p=0.19).  The complete dataset that includes biopsy results and functional assessments and will be presented at a medical conferences in the future.

The study’s Senior Scientist, Dr. Tobias Winkler of the Center for Musculoskeletal Surgery, Julius Wolff Institute Berlin, Charité – Universitaetsmedizin Berlin, Germany, commented, “I am very impressed with the magnitude of the efficacy results seen in this trial. PLX cells demonstrated safety and suggested that the increase in muscle volume could be a mechanism for the improvement of contraction force.”

Zami Aberman Chairman and CEO stated, “This was a very important study not only for Pluristem but for the cell therapy industry in general. The study confirms our pre-clinical findings that PLX-PAD cell therapy can be effective in treating muscle injury. Having a statistically significant result for our primary efficacy endpoint is very encouraging and consistent with our understanding of the mechanism of action associated with cell therapy. Based on these results, we intend to move forward with implementing our strategy towards using PLX cells in orthopedic indications and muscle trauma.”

The Stem Cell Blog

  • cells were safe and well tolerated
  • one group receiving a 150 million cell dose displaying a 500 percent improvement over the placebo group.
  • Patients treated with a 300 million cell dose showed a 300 percent improvement over the placebo.
  • An analysis of the gluteal muscle indicated an increase in muscle volume in those patients treated…versus the placebo group.

Pluristem stem cell trial to treat muscle injury meets main goal

TEL AVIV Tue Jan 21, 2014 3:07am EST

(Reuters) – Pluristem Therapeutics Inc said results from its early/mid-stage clinical trial indicated its placenta-derived stem cells for the treatment of muscle injury were safe and provided evidence the cells might be effective in treating orthopedic injuries.

\”Patients treated with PLX-PAD had a greater improved change of maximal voluntary muscle contraction force than the placebo group,\” Israel-based Pluristem said in a statement on Tuesday.

The trial was conducted at the Orthopedic Clinic of…

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Treating Age-Related Blindness with a Stem Cell Replacement Method


A collaboration between German and American scientists in New York City has resulted in the invention of a new method for transplanting stem cells into the eyes of patients who suffer from age-related macular degeneration, which is the most frequent cause of blindness. In an animal test, the implanted stem cells survived in the eyes of rabbits for several weeks.

Approximately 4.5 millino people in Germany suffer from age-related macular degeneration (AMD), which causes gradual loss of visual acuity and affects the ability to read, drive a car or do fine work. The center of the vision field becomes blurry as though covered by a veil. This vision loss is a consequence of the death of cells in the retinal pigment epithelium or RPE, which lies are the back of the eye, underneath the neural retina.

Inflammation within the RPE causes AMD. Increased inflammation prevents efficient recycling of metabolic waste products, and the build-up of toxic wastes causes RPE die off. Without the RPE, the photoreceptors in front of the RPE cells that also depend on the RPE to repair the damage suffered from continuous light exposure, begin to die off too.

RPE

Retinal Pigmented Epithelium

Presently no cure exists for AMD, but scientists at Bonn University, in the Department of Ophthalmology and New York City have tested a new procedure that replaces damaged RPE cells.

In the present experiment, RPE cells made from human stem cells were successfully implanted into the retinas of rabbits.

Boris V. Stanzel, the lead author of this work, said, “These cells have now been used for the first time in research for transplantation purposes.”

The adult RPE stem cells were characterized by Timothy Blenkinsop and his colleagues at the Neural Stem Cell Institute in New York City. Blenkinsop designed methods to isolate and grow these cells. He also flew to Germany to assist Dr. Stanzel with the transplantation experiments.  Blenkinsop obtained his RPE cells from human cadavers, and he grew them on polyester matrices.

These experiments demonstrate that RPE cells obtained from adult stem cells can replace cells destroyed by AMD. This newly developed transplantation method makes it possible to test which stem cells lines are most suitable for transplantation into the eye.

Stem Cell-Based Gene Therapy Restores Normal Skin Function


Michele De Luca from the University of Modena, Italy and his collaborator Reggio Emilia have used a stem cell-based gene therapy to treat an inherited skin disorder.

Epidermolysis bullosa is a painful skin disorder that causes the skin to be very fragile and blister easily. These blisters can lead to life-threatening infections. Unfortunately, no cure exists for this condition and most treatments try to alleviate the symptoms and infections.

Stem cell-based therapy seems to be one of the best ways to treat this disease, there are no clinical studies that have examined the long-term outcomes of such a treatment.

However, De Luca and his colleagues have examined a particular patients with epidermolysis bullosa who was treated with a stem cell-based gene therapy nearly seven years ago as part of a clinical trial.

The treatment of this patient has established that transplantation of a small quantity of stem cells into the skin on this patient’s legs restored normal skin function without causing any adverse side effects.

“These findings pave the way for the future safe use of epidermal stem cells for combined cell and gene therapy of epidermolysis bullosa and other genetic skin diseases,” said Michele De Luca.

De Luca and his research team found that their treatment of their patient, named Claudio, caused the skin covering his upper legs to looker normal and show no signs of blisters. To treat Claudio, De Luca and his colleague extracted skin cells from Claudio’s palm, used genetic engineering techniques to correct the genetic defect in the cells, and then transplanted these cells back into the skin of his upper legs. This was part of a clinical trial conducted at the University of Modena.

Claudio’s legs also showed no signs of tumors and the small number of transplanted cells sufficiently repaired Claudio’s skin long-term. Keep in mind that Claudio’s skin cells had undergone approximately 80 cycles of cell division and still had many of the features of palm skin cells, they show proper elasticity and strength and did not blister.

“This finding suggests that adult stem cell primarily regenerate the tissue in which they normally reside, with little plasticity to regenerate other tissues,” De Luca said. “This calls into question the supposed plasticity of adult stem cells and highlights the need to carefully chose the right type of stem cell for therapeutic tissue regeneration.”

I think De Luca slightly overstates his case here. Certainly choosing the right stem cells is crucial for successful stem cell treatments, but to take stem cells from skin, which are dedicated to making skin and expect them to form other tissues is unreasonable. However, several experiments have shown that stem cells from hair follicles and form neural tissues and several other cell types as well (see Jaks V, Kasper M, Toftgård R. The hair follicle-a stem cell zoo. Exp Cell Res. 2010 May 1;316(8):1422-8).

Adult stem cells have limited plasticity to be sure, but their plasticity is far greater than originally thought and a wealth of experiments have established that.

Despite these quibbles, this is a remarkable experiment that illustrates the feasibility and safety of such a treatment.  A larger problem is that large quantities of cells will be required to treat a person.  It is doubtful that small skin biopsies around the body can provide enough cells to treat the whole person.  Therefore, this might a case for induced pluripotent skin cells, which seriously complicates this treatment strategy.

Stem Cell Therapy Following Meniscus Knee Surgery Reduces Pain and Regenerates Meniscus


According to a new study published in the January issue of the Journal of Bone and Joint Surgery (JBJS), a single stem cell injection after meniscus knee surgery can provide pain relief and aid in meniscus regrowth.

In the US alone, over one million knee arthroscopy procedures are performed each year. These surgeries are usually prescribed to treat tears to the wedge-shaped piece of cartilage on either side of the knee called the “meniscus.” The meniscus acts as an important shock absorber between the thighbone (femur) and the shinbone (tibia) at the knee-joint.

Knee-Ligament-Pain-and-Strains-Meniscus-Tear-and-Pain

This novel study, “Adult Human Mesenchymal Stem Cells (MSC) Delivered via Intra-Articular Injection to the Knee, Following Partial Medial Meniscectomy,” examined 55 patients who had undergone a surgical removal or all or part of a torn meniscus (known as a partial medial meniscectomy). Each patient was randomly assigned to one of three treatment groups: Groups A, B and C. The 18 patients in group A received a “low-dose” injection of 50 million stem cells within seven to 10 days after their meniscus surgery. Another 18 patients in group B received a higher dose of 150 million stem cells seven to ten days after their knee surgery. The controls group consisted of 19 patients who received injections of sodium hyaluronate only (no stem cells). All patients were evaluated to determine the safety of the procedure, the degree of meniscus regeneration (i.e. with MRI and X-ray images), the overall condition of the knee-joint, and the clinical outcomes through two years. Most of the patients enrolled in this study had some arthritis, but patients with severe (level three or four) arthritis, were excluded from the study.

Most of the patients who had received stem cell treatments reported a significant reduction in pain. 24 percent of the patients in one MSC group and 6 percent of the other showed at least a 15 percent increase in meniscal volume at one year. Unfortunately, there was no additional increase in meniscal volume at year two.

“The results demonstrated that high doses of mesenchymal stem cells can be safely delivered in a concentrated manner to a knee-joint without abnormal tissue formation,” said lead study author C. Thomas Vangsness, Jr., MD. “No one has ever done that before.” In addition, “the patients with arthritis got strong improvement in pain” and some experienced meniscal regrowth.

The key findings of this study are that there no abnormal (ectopic) tissue formation or “clinically important” safety issues identified. Also, 24 percent of the patients in the low-dose injection group (A) and six percent of the high-dose injection group (B) at one year showed “significantly increased meniscal volume,” as determined by an MRI, and this increase did not continue into the second year, but remained stable (should future studies try a second injection of MSCs?). Third, none of the patients in the control group (non-MSC group) showed significant meniscus regrowth. Finally, patients with osteoarthritis experienced a reduction in pain in the stem cell treatment groups, but there was no reduction in pain in the control (non-MSC group).

“The results of this study suggest that mesenchymal stem cells have the potential to improve the overall condition of the knee joint,” said Dr. Vangsness. “I am very excited and encouraged” by the results. With the success of a single injection, “it begs the question: What if we give a series of injections?”

Stem-Cell Gene Therapy for Sickle Cell Disease


Donald Kohn, a professor of pediatrics and microbiology, immunology and molecular genetics in the UCLA College of Letters and Science, and his colleagues, have successfully established the means to cure sickle-cell disease. This strategy uses hematopoietic (blood-producing) stem cells from the bone marrow of patients with sickle-cell disease in order to treat the disease itself.

This approach provides a revolutionary alternative to current treatments, since it creates self-renewing, normal blood cells by inserting a gene that abrogates the sickling properties into hematopoietic stem cells. With this technique, there is no need to identify a matched donor, and therefore, patients avoid the risk of their bodies rejecting donor cells.

During the clinical trial, the anti-sickling hematopoietic stem cells will be transplanted back into patients’ bone marrow to increase the population of “corrected” cells that make red blood cells that don’t sickle. Kohn will hopefully begin enrolling patients in the trial within three months. The first subject will be enrolled and observed for safety for six months. The second subject will then be enrolled and observed for safety for three months. If evaluations show that no problems have arisen, the study will continue with two more subjects and another evaluation, until a total of six subjects have been enrolled.

Sickle cell disease, which affects more than 90,000 individuals in the U.S., is seen primarily in people of sub-Saharan African descent. It is caused by an inherited mutation in the beta-globin gene that transforms normal-shaped red blood cells, which are round and pliable, into rigid, sickle-shaped cells. Normal red blood cells are able to pass easily through the tiniest blood vessels (capillaries) and carry oxygen to organs like the lungs, liver and kidneys. However, sickled cells get stuck in the capillaries, depriving the organs of oxygen, which can lead to organ dysfunction and failure.

Current treatments include transplanting patients with hematopoietic stem cells from a donor. This is a potential cure for the disease, but due to the serious risks of rejection, only a small number of patients have undergone this procedure, and it is usually restricted to children with severe symptoms.

“Patients with sickle-cell disease have had few therapeutic options,” Kohn said. “With this award, we will initiate a clinical trial that we hope will become a treatment for patients with this devastating disease.”

Finding for this work comes from new grants to researchers at UCLA’s Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, which total nearly $21 million.  These grants were announced Dec. 12 at a meeting of the California Institute of Regenerative Medicine (CIRM) Citizen’s Oversight Committee.  They are apart of the state agency’s Disease Team Therapy Development III initiative.

Transplanted Liver Cells do Better When Co-Cultured with Mesenchymal Stem Cells


Implanting frozen liver cells is a relatively new procedure that has, reportedly, been used to treat very young patients with liver problems. Thawing frozen liver cells, however, tends to cause a fraction of the cells to die off and other damaged cells show poor function.

To ameliorate this problem, researchers at Kings College Hospital, London have used mesenchymal stem cells from fat or umbilical cord to improve the viability and function of frozen liver cells.

Emer Fitzpatrick and her colleagues at Kings College Hospital reasoned that mesenchymal stem cells and the multitudes of healing molecules that these cells secrete should be able to “lend proregenerative characteristics to liver cells.”

Thus by co-culturing thawed liver cells with mesenchymal stem cells from fat or umbilical cord, Fitzpatrick and others demonstrated that the rate of cell survival of the liver cells and their functionality increased in comparison with liver cells grown on their own.

Fitzpatrick hopes that such a co-culture technique might improve the clinical usefulness of frozen liver cells for transplantation.

Cancer Stem Cell Research Leads to Clinical Trials


Dennis Slamon and Zev Wainberg from the UCLA Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research have been awarded a Disease Team Therapy Development award to begin clinical trials in human patients early in 2014.

In this clinical trial, Slamon and Wainberg will test a new drug that targets cancer stem cells. This drug was developed by research and development over the last decade on the cancer stem cell hypothesis. The cancer stem cell hypothesis predicts that proliferating stem cells are the main drivers of tumor growth and are also resistant to standard cancer treatments.  This new drug, CFI-400945, has prevented cancer growth in an extensive series of laboratory animal tests.

An important extension of the cancer stem cell hypothesis is that cancer stem cells inhabit a particular niche that prevents anticancer drugs from reaching them. Alternatively, tumors become resistant to cancer drugs by a process called “cell fate decision,” in which some cancer stem cells are killed by chemotherapy, but other cells replace them and repopulate the tumor. This tumor repopulation is the main reason for cancer recurrence.

The new anticancer drug to be tested in this clinical trial targets the “polo-like kinase 4.” Inhibition of this enzyme effectively blocks cell fate decisions that cause cancer stem cell renewal and tumor cell growth. Thus inhibition of this enzyme effectively stops tumor growth.

This clinical trial will test this novel chemotherapeutic agent in patients to establish the safety of the drug. After these initial safety tests, the trial will quickly proceed to further clinical tests. “We are excited to continue to test this drug in humans for the first time,” said Wainberg. Slamon, Wainberg and others will also look for biological markers to determine how well their drug is working in each patient.

The US Food and Drug Administration approved the Investigational New Drug (IND) for this drug trial. Also, Health Canada, the Canadian government’s therapeutic regulatory agency, also approved this trial. These approvals are part of an international effort to bring leading-edge stem cell science to patients.

New Tool for Stem Cell Transplantation into the Heart


Researchers from the famed Mayo Clinic, in collaboration with scientists at a biopharmaceutical biotechnology company in Belgium have invented a specialized catheter for transplanting stem cells into a beating heart.

This new device contains a curved needle with graded openings along the shaft of the needle. The cells are released into the needle and out through the openings in the side of the needle shaft. This results in maximum retention of implanted stem cells to repair the heart.

“Although biotherapies are increasingly more sophisticated, the tools for delivering regenerative therapies demonstrate a limited capacity in achieving high cell retention in the heart,” said Atta Behfar, the lead author of this study and a cardiologist. “Retention of cells is, of course, crucial to an effective, practical therapy.”

Researchers from the Mayo Clinic Center for Regenerative Medicine in Rochester, MN and Cardio3 Biosciences in Mont-Saint-Guibert, Belgium, collaborated to develop the device. Development of this technology began by modeling the dynamic motions of the heart in a computer model. Once the Belgium group had refined this computer model, the model was tested in North America for safety and retention efficiency.

These experiments showed that the new, curved design of the catheter eliminates backflow and minimizes cell loss. The graded holes that go from small to large diameters decrease the pressures in the heart and this helps properly target the cells. This new design works well in healthy and damaged hearts.

Clinical trials are already testing this new catheter. In Europe, the CHART-1 clinical trial is presently underway, and this is the first phase 3 trial to examine the regeneration of heart muscle in heart attack patients.

These particular studies are the culmination of years of basic science research at Mayo Clinic and earlier clinical studies with Cardio3 BioSciences and Cardiovascular Centre in Aalst, Belgium, which were conducted between 2009 and 2010.  This study, the C-CURE or Cardiopoietic stem Cell therapy in heart failURE study examined 47 patients, (15 control and 32 experimental) who received injections of bone marrow-derived mesenchymal stem cells from their own bone marrow into their heart muscle.  Control patients only received standard care.  After six months, those patients who received the stem cell treatment showed an increase in heart function and the distance they could walk in six minutes.   No adverse effects were observed in the stem cell recipients.

This study established the efficacy of mesenchymal stem cell treatments in heart attack patients.  However, other animal and computer studies established the efficacy of this new catheter for injecting heart muscle with stem cells.  Hopefully, the results of the CHART-1 study will be available soon.

Postscript:  The CHART-2 clinical trial is also starting.  See this video about it.

Induced Pluripotent Stem Cells Self-Repair a Chromosome Abnormality


Japanese and American scientists have made a fantastic discovery. An incurable type of chromosomal abnormality, in which one end of the chromosome has fused with the other, is known as a “ring chromosome.” The presence of ring chromosomes often correlates with neurological abnormalities. The collaborative work by these two researcher groups has shown that if induced pluripotent stem cells are derived from abnormal adult cells that contain ring chromosomes will spontaneously repair themselves.

This research team, which included researchers from Kyoto University professor and iPS cell pioneer Shinya Yamanaka, also included Yohei Hayashi and other researchers from the U.S.-based Gladstone Institutes.

“I was very surprised at the results,” said Hayashi. “There still remains a risk, but the findings may lead to the development of breakthrough treatment for chromosomal abnormalities.”

Normal chromosomes pairs consist of two rod-shaped chromosomes, but in the case of a ring chromosome, the arms of one of the two chromosomes are fused to form a ring.

Ring chromosomes tend to be associated with mental disabilities and growth retardation, and there are no therapeutic strategies for ring chromosomes.

Hayashi and his colleagues developed iPS cells from skin cells of patients with ring chromosome disorder to study the effects of this chromosomal defect in stem cells. However, the ring chromosomes could not be observed in the engineered iPS cells made from this patient’s cells.

For reasons still unknown, only normal chromosomes survived, according to the researchers. Typically, a cell that contains ring chromosomes divides into two abnormal cells in the normal mitotic process.

These findings were published in the online edition of the British science journal Nature on Jan. 13, 2014.