Neural Stem Cells Relieve Some of the Impairments of Dementia in Mice

Lewy bodies are aggregations of misfolded proteins in nerve cells that can kill them off and cause dementia. When Lewy bodies form in neurons, they can cause “dementia with Lewy bodies” or DLB. After Alzheimer’s disease, DLB is the second-most common type of age-related dementia, and it afflicted the beloved comedian Robin Williams, who took his own life earlier this year.

Scientists at the Sue and Bill Gross Stem Cell Research Center and the Institute for Memory Impairments and Neurological Disorders at UC Irvine have examined the ability of transplanted neural stem cells to ameliorate the symptoms of DLB is an animal model system.

Particular strains of laboratory mice have been genetically engineered to form Lewy bodies in their brains and show some of the symptoms of DLB. Natalie Goldberg and her colleagues used neural stem cells to treat some of these mice in order to determine if these cells could decrease the pathological consequences of DLB.

Transplantation of neural stem cells into the brains of these DLB mice resulted in increases in cognitive and motor function. A battery of tests established this. For example, the Rotarod test places the mouse on a rod that is then rotated at a specific speed. Normal mice can move around the rod and keep from falling off, but mice with motor or balance problems will fall off the rod prematurely. Cognitive tests included Novel Object Recognition (NOR) and Novel Place Recognition (NPR) tests, which are low-stress tasks that quantify the proportion of time spent examining a novel object and provide data on memory. In these tests, the mice that received the neural stem cell transplantation did significantly better than their non-treated siblings.

Goldberg and his team then asked how these cells improved the cognitive and motor function of the DLB mice. It turns out that neutral stem cells secrete respectable amounts of brain-derived neurotrophic factor (BDNF). Goldberg suspected that this growth factor was a major contributor to the healing capabilities of neural stem cells. Therefore, Goldberg’s team engineered neural stem cells that could not make BDNF and injected those directly into the brains of DLB mice. These mutant neural stem cells were incapable of improving the cognitive or motor function of these mice.

To further test her hypothesis, Goldberg then engineered a virus that would infect neurons and overexpress BDNF and used that to treat her DLB mice. Interestingly, the BDNF-expressing virus did a pretty good job at restoring motor functions in these DLB mice, but did not restore the cognitive functions.

Thus, while the secretion of BDNF by neural stem cells is important for their restorative capacities, but it is only part of the means they use to heal affected brains. Goldberg and her coworkers showed that the transplanted neural stem cells did not improve the pathology of the brains, they did preserve neural pathways that use the neurotransmitters dopamine and glutamate.

The neural stem cells used in these experiments were mouse neural stem cells. Before work like this can advance to human clinical trials, human neural stem cells must be tested. Since other neurodegenerative diseases like Parkinson’s disease also result from Lewy body formation in specific cells, neural stem cell treatments might prove beneficial for patients with much diseases.

This work was published in Stem Cell Reports October 2015 DOI: 10.1016/j.stemcr.2015.09.008.

cKit+ cells Do Make Heart Muscle After All

In the heart lies a population of cells that contains a protein called cKit, and are, therefore, called cKit+ cells. cKit+ cells have been the subject of a good deal of attention by researchers, but unfortunately, these have become the focus of a good deal of controversy.

When cKit+ cells were first discovered, there was a good deal of excitement about them, since they seemed to be able to make heart muscle cells and replace damaged heart muscle cells in the heart of living creatures (Beltrami AP, et al., Cell. 2003 Sep 19;114(6):763-76). In 2012, the results of a Phase I clinical trial with cKit+ cells (SCIPIO) were published (Chugh AR et al., Circulation 2012 Sep 11;126(11 Suppl 1):S54-64). This trial seemed to show that patients who received their own cKit+ cells had significant increases in heart function after a heart attack. Follow-up work in pigs even appeared to confirm that infused cKit+ cells could differentiate into heart muscle and integrate into the walls of the heart (Bolli R et al., Circulation 2013 Jul 9;128(2):122-31). So the cells were able to regenerate heart muscle in mice, pigs, and humans. It is not an understatement to say that cKit+ cells were once thought to be the key to cardiac regeneration.

The first trouble in paradise came from mouse experiments. While cKit+ cells could indeed improve the function of damaged hearts, the evidence for engraftment of the cells into the walls of the heart was wanting. Scientists in the laboratory of Jeff Molkentin Cincinnati Children’s Hospital Medical Center reported in a high-profile paper in the journal Nature that cKit+ cells can readily produce cardiac blood vessel cells, they rarely make heart muscle cells (cardiomyocytes). Because Molkentin and his team had carefully marked and traced the cells that they implanted into mice, the result was pretty devastating to the status of cKit+ cells. Molkentin’s results, however, conflicted with data from the laboratory of Bernardo Nadal-Ginard from King’s College London, who showed that heart regeneration in laboratory rodents depends on cKit+ cells and depleting cKit+ populations from the heart abolishes the ability of the heart to repair itself (Ellison GM, et al., Cell. 2013 Aug 15;154(4). Technical differences between the two papers, however, made comparisons between them difficult.

The next issues came with the SCIPIO publication itself. Two of the figures appeared to have some mistakes in them. Piero Anversa from Brigham and Women’s Hospital’s, the senior author of the SCIPIO study, admitted that there might be problems with the figures but insisted that the clinical data of the trial were sound. Other concerns about SCIPIO were expressed as well in print.  Add to that the fact that Anversa had to retract one of his earlier papers, and the whole edifice of SCIPIO and cKit+ cells seemed to totter.  These issues knocked cKit+ cells off their pedestal. At the very least, they put a hold on the SCIPIO trial until other questions had been resolved.

A new study by Joshua Hare and his group from the University of Miami Miller School of Medicine has stirred up the controversy pot once again. Hare and his team have published a paper in the journal PNAS in which they showed that cKit+ cells can readily form heart muscle cells in culture. However, apparently the cKit+ cells are finicky and only form heart muscle conditions if the conditions are just right. These results from Hare’s group might (and oh what a big might) explain why other groups have not been able to replicate the results of either Anversa or Nadal-Ginard. In Hare’s own words, “It’s not that the [cKit+] cells don’t have the capacity [to form heart muscle], but they’re entering the heart at a time that’s nonpermissive for them to become cardiac myocytes.”

In a nutshell, Hare and his team used mouse induced pluripotent stem cells (iPSCs) and differentiated them into heart muscle cells. They found that if you inhibited bone morphogenetic protein (BMP) signaling in these cells, an integral signaling event in the development of the heart; the iPSCs would express cKit and differentiate into heart muscle cells. The Hare group also used fate-mapping techniques to trace the developmental origin of cKit+ cells in the heart and they discovered that cKit+ cells are derived from the neural crest cells that delaminate from the closing neural tube during the formation of the central nervous system and migrate throughout the body to form a whole host of cell types and contribute to many different tissues.

Unlike Molkentin’s group, Hare and his crew did not observe an increased tendency for cKit+ cells to form heart blood vessel (endothelium) cells. Hare was somewhat unsure why this might be the case, but suggested that the different ways that the two teams labeled their cells for fate mapping purposes might be at least part of the issue.

Despite his success at showing that cKit+ cells can become heart muscle cells, Hare does not think that his work necessarily explains the results of the SCIPIO clinical trial, but he does think that his work might suggest how the regenerative capacities of cKit+ cells might be augmented.

Bernardo Nadal-Ginard found Hare’s work “convincing,” but added that “the paper claims the quandary and the dispute is over. But, unfortunately, it is not.” I think we can say “Amen” to that, since more work almost certainly needs to be done. Nadal-Ginard also brought up a very good point when he added that no one really knows the frequency with which cKit+ cells differentiate into heart muscle cells or other cells types or even the rate with which they replace dead or dying cells. Hare’s paper did not focus on quantitating such events, and since it did not examine the ability of cKit+ cells to repopulate a living heart, these are still questions that must be addressed.

Cornell University’s Michael Kotlikoff also made an excellent point by noting that Hare’s team did not show that cKit+ cells have the same ability to regenerate a living heart in laboratory animals as they do in culture. In an article in The Scientist by Kerry Grens, Kotlikoff said, “They never show the myogenic potential of those cells and don’t show them giving rise to cardiomyogensis” in vivo. Kotlikoff continued: “The expression of [cKit], per se, is not sufficient to identify cells as precursors and the further presumption that signaling processes observed in in vitro differentiation experiments limit such cells from undergoing myogenesis in the adult heart, the stage at which clinical regenerative efforts are focused, is not supported by data,” he added.

Hare almost certainly is either planning or is presently carrying out such experiments with laboratory mice. Presently, however, Hare has founded a company called Vestion, whose goal is to establish off-the-shelf regenerative heart therapies. According the Kerry Grens, Hare is also a part of two planned clinical trials that will administer cKit+ cells to patients with heart failure.

Piero Anversa, who remains a big fan of cKit+ cells despite their knocks, spoke approvingly of Hare’s paper and added, “To say human trials should be stopped because the experiment didn’t work in the mouse is a bit aggressive. The answer is going to be in the trial. If the trial goes well we win, if the trial doesn’t go well, we lose.”

Thyroid Organoids Made from Stem Cells Treat Thyroid-Deficient Mice

Darrell Kotton and his research team from Beth Deaconess Medical Center, in collaboration with researchers from the Boston University School of Medicine have devised a workable protocol for differentiating Human pluripotent stem cells into functional thyroid gland cells.

Every year, many people are diagnosed with an underactive thyroid and many others lose their thyroid as a result of thyroid cancer. Designing treatments that can help replace lost thyroid tissue would certainly be a welcome thing for these patients.

By working with mouse embryonic stem cells, Kotton and his colleagues showed that two growth factors, BMP4 and FGF2, and induce foregut endodermal cells to differentiate into thyroid cells. This simple signaling pathway not only efficiently generates thyroid tissue from endoderm, but this pathway turns out to be commonly used in species as diverse as frogs, mice and humans.

The BMP4/FGF2-treated foregut cells differentiated into small thyroid organics that Kotton and his team were able to transplant into thyroid-deficient mice. These transplantations restored normal thyroid function to these mice.

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While mice cells are a fine model system for human diseases, they are not exactly the same. Can this procedure work with human cells? To answer that question, Kotton and his coworkers used human induced pluripotent stem cells (iPSCs) and subjected them to the same BMP4/FGF2 protocol after they had first differentiated the cells into endoderm. In addition, Kotton and his team made thyroid cells from iPSCs derived from cells taken from patients with a specific type of hypothyroidism (interactive thyroid). These patients lack a gene called NKX2-1, and suffer from congenital hypothyroidism.

The thyroid is responsible for your basal metabolic rate. Hypothyroidism or an interactive thyroid can cause patients to gain weight, feel tired constantly, have trouble concentrating, and have a slow heart rate. Hypothyroidism is usually treated with synthetic thyroid hormones that are taken orally. However, restoring a patient’s own thyroid tissue or even replacing defective thyroid tissue with repaired thyroid tissue would be a huge boon to thyroid patients.

This work has discovered the regulatory mechanisms that drive the establishment of the thyroid. It also provides a significant step toward cell-based regenerative therapy for hypothyroidism and the replacement of the thyroid after thyroid cancer treatments.

These results were published in the journal Cell Stem Cell, October 2015 DOI:10.1016/j.stem.2015.09.004.

Sleep Deprivation Decreases Stem Cell Activity

We have all been there: You are at your computer, working hard and then a yawn hits you. Alternatively, you are on the phone late at night and you start to nod. We all have our late nights burning the midnight oil, but we need our shut-eye.

Now it turns out that sleep deprivation might wreak havoc with your stem cells. New research in mice might (let me emphasize, might) have profound implications for patients undergoing bone marrow stem cell transplants.

This research was led by Dr. Asya Rolls, who formerly worked as a postdoctoral research fellow at Stanford University, but is now an assistant professor at the Israel Institute of Technology.

With regards to the clinical implications of this work, Dr. Rolls said, “Considering how little attention we typically pay to sleep in the hospital setting, this finding is troubling. We go to all this trouble to find a matching donor, but this research suggests that if the donor is not well-rested it can impact the outcome of the transplantation. However, it’s heartening to think that this is not an insurmountable obstacle; a short period of recovery sleep before transplant can restore the donor’s cells’ ability to function normally.”

Rolls and her colleagues used laboratory mice for this study and broke them into two different groups. One group of mice was physically handled by members of the research team for four hours in order prevent them from going to sleep. The other group of mice were not handled and slept soundly in their cages. Then Rolls and her collaborators isolated bone marrow stem cells from the sleepless and well-rested mice. These bone marrow stem cells were then used to them to help reconstitute the bone marrow of twelve different mice that had been given radiation treatments that wiped out their bone marrow stem cells. It is important to note that these donor mice had bone marrow stem cells that glowed when put under a fluorescent light.

The irradiated mice were then examined eight and 16 weeks after they had received the bone marrow stem cell transplants. By taking blood samples, Roll and others measured the production of blood cells by the transplanted bone marrow stem cells. Mind you, the irradiated mice also received some of their own bone marrow stem cells in combination with the bone marrow stem cells from the donor mice. This was to help determine the percentage of blood cells made by the stem cells from the donor mice. Surveys of the blood cells of the irradiated mice showed that donated stem cells from the mouse donors that had a good night’s sleep gave rise to about 26 percent of the examined blood cells. However, bone marrow stem cells from sleepless donor mice only produced approximately 12 percent of the surveyed blood cells.

Next, the Stanford team investigated the ability of the transplanted stem cells to find their way to the bone marrow of the recipient mice, twelve hours after transplantation. When the bone marrow of the donor mice was subjected to fluorescent light, the 3.3 percent of the bone marrow stem cells were from the well-rested donor mice. However, the same experiment in those recipient mice that had received mice had received bone marrow stem cells from the sleep-deprived mice showed that only 1.7 percent of the stem cells in the bone came from the donor mice. Thus the bone marrow stem cells from those mice that had a good night’s sleep were twice as likely to find their way to the bone marrow of the recipient.

When hematopoietic stem cells from the donor mice were tested in culture, stem cells from the sleepless mice showed a weak response to chemical cues found in bone marrow that activate migration to the bone marrow. Conversely, hematopoietic stem cells from the well-rested mice responded much more robustly to these same chemical cues and migrated appropriately.

Think of it; not sleeping for only four hours can decrease the activity of transplanted bone marrow stem cells by up to half. Remember that bone marrow stem cells contain the coveted hematopoietic stem cell population that produces all the blood cells coursing through our bloodstream. When transplanted into recipient animals (or patients), these stem cells must actively find their way to the bone marrow, take up residence there, and begin to produce all the blood cells necessary for the life and health of the recipient. Therefore even a small reduction in the health or activity of hematopoietic stem cells could drastically affect the success of the bone marrow transplant procedure.

Are the effects of sleeplessness permanent? Not at all, at least in mice. Rolls and her team showed that the decrease in bone marrow stem cell activity could be reversed by allowing the sleep-deprived mice to sleep. In fact, in the hands of Rolls and her co-workers, even letting mice get only two hours of recovery sleep effectively restored the activity of their bone marrow stem cells to properly reconstitute the bone marrow of a recipient in a bone marrow transplant procedure.

“Everyone has these stem cells, and they continuously replenish our blood and immune system,” said Rolls. “We still don’t know how sleep deprivation affects us all, not just bone marrow donors. The fact that recovery sleep is so helpful only emphasizes how important it is to pay attention to sleep.”

Bone marrow transplants are used to treat patients with blood cancers, immune system disorders or others types of conditions. Each year, many thousands of bone marrow transplant procedures are performed. Therefore refining the bone marrow stem cell transplant procedure is essential to helping patients who need such a procedure.

This study was published in Nature Communications, with Asya Rolls as the lead author, who did her work in the laboratory of Irving Weissman, the director of the Stanford Institute of Stem Cell Biology and Regenerative Medicine.

New Gene Therapy Effectively Treats All Muscles in Dogs With Muscular Dystrophy

The X-linked genetic disease, muscular dystrophy, affects the structure and function of skeletal muscles. Muscular dystrophy patients harbor mutations in a gene that encodes a protein known as dystrophin. Dystrophin attaches the internal skeleton of skeletal muscle cells to the cell membrane. In turn, proteins in the skeletal muscle membrane attach to the intracellular matrix that acts as the foundational material upon which muscle cells (and other cells) sit. Therefore, the dystrophin protein serves to attach skeletal muscle cells to the extracellular matrix. The loss of dystrophin causes muscles to separate from the cell matrix and detach from each other. The lack of attachment of muscles to each other causes them to degenerate and die.


The death of skeletal muscles in muscular dystrophy patients leads to the replacement of what was once skeletal muscle with scar tissue, fatty tissue, or even bone. Because muscular dystrophy is caused by mutations in an X-linked gene, the majority of muscular dystrophy patients are boys. The losses of muscle structure, function, and mass cause patients to lose their ability to walk and eventually breath (since the diaphragm is a skeletal muscle) as they age. Thus muscular dystrophy tends to put patients in wheelchairs and condemn them to respirators.

The most common form of muscular dystrophy is called Duchenne Muscular Dystrophy or DMD. Close to 250,000 people in the United States suffer from muscular dystrophy. Treatment options are very limited and usually palliative. However, a research team from the University of Missouri has successfully treated dogs that suffer from DMD. They are optimistic that human clinical trials can be planned in the next few years.

This is a remarkable finding, especially, when you consider that the dystrophin gene is extremely large. In fact, the dystrophin gene is the largest gene in the human genome. This makes gene therapy treatments for DMD problematic.

Dongsheng Duan, who serves as the lead scientist in this study, and is the Margaret Proctor Mulligan Professor in Medical Research at the MU School of Medicine “This is the most common muscle disease in boys, and there is currently no effective therapy. This discovery took our research team more than 10 years, but we believe we are on the cusp of having a treatment for the disease.

Duan continued: “Due to its size, it is impossible to deliver the entire gene with a gene therapy vector, which is the vehicle that carries the therapeutic gene to the correct site in the body,” Duan said. “Through previous research, we were able to develop a miniature version of this gene called a microgene. This minimized dystrophin protected all muscles in the body of diseased mice.”

Duan and his colleagues worked for almost ten years to develop a viable strategy that can safely transfer the micro-dystrophin gene to every muscle in a the body of dogs that have a canine form of DMD. Dogs are an excellent model system for human medicine, since dogs are about the same size as a human boy. Successful treatment of DMA dogs can provide the foundation for human clinical trials.

In this new study, Duan and his team demonstrated that by using a common virus to deliver the micro-dystrophin gene to all the muscles in the body of a diseased dog. Duan and others injected DMA dogs with this genetically engineered virus when they were two-three months old. For dogs, this is about the time when they begin to show some of the DMD-associated signs and symptoms. Now, these dogs are six-seven months old and they are experiencing normal development and muscular activity.

“The virus we are using is one of the most common viruses; it is also a virus that produces no symptoms in the human body, making this a safe way to spread the dystrophin gene throughout the body,” Duan said. “These dogs develop DMD naturally in a similar manner as humans. It’s important to treat DMD early before the disease does a lot of damage as this therapy has the greatest impact at the early stages in life.”

Bone Marrow Stem Cell Injections Restore Fertility In Mice Made Sterile by Chemotherapy

Every year, over 20,000 women of childbearing age are diagnosed with cancer. Cancer treatments often include chemotherapy regimens that damage other tissues and the ovaries and its eggs are particularly sensitive to such treatments. Consequently, many young, female, cancer survivors are infertile as a result of their cancer treatments, and suffer early menopause and ovarian failure.

Now an earth-shaking study by Egyptian and American scientists has shown that stem cell injections into the ovaries can rejuvenate them and restore the fertility of laboratory animals.

“This approach carries high promise to women with chemotherapy-induced and potentially other types of premature ovarian failure,” said Dr Sara Mohamed, lead researcher for this project.

Woman who must undergo chemotherapy are routinely advised to freeze their eggs before they undergo any cancer treatments. However this procedure is labor intensive and takes time, and in urgent cases, there is not enough time to preserve the patient’s eggs. This leaves the woman in the unsavory position of having to decide between her fertility or her life.

A procedure like the one used in this study might give female patients other options that do not force them to choose between the Scylla of their ability to have their own children and the Charybdis of their survival.

To date, this procedure has been successfully performed in laboratory mice. In this experiment, a clutch of eighteen laboratory mice were broke into three groups of six. One group of six female mice was treated with anticancer chemotherapeutic agents, followed by injections of bone marrow stem cells into their ovaries. The second group of six female mice also received chemotherapy, followed by injections of sterile saline into their ovaries. The third group, a control group, received injections of sterile saline into their ovaries without receiving prior treatments with chemotherapy.

One week after receiving their treatments, the stem cell-treated mice showed a significant increase in estrogen production. Since estrogen is a sex steroid hormone that is essential to ovulation, these results suggested that the menstrual cycles of the infertile mice was actually being reconstituted. Then a week later, mice in the stem cell-treated group showed regeneration of their ovarian tissue and increased numbers of ovarian follicles. Ovarian follicles produce the sex steroid hormones estrogen and progesterone and contain a single egg that matures during the follicular stage of the menstrual cycle and is potentially released during ovulation. These same mice, which had experienced ovarian failure as a result of chemotherapy, were able to mate with male mice, and eventually give birth to large litters of healthy mouse pups while those who had saline injections continued to suffer from reduced fertility of even infertility.


These treatments worked so remarkably well, that the members of the researcher team who were involved with this project want to move to human trials as soon as possible.

Dr Sara Mohamed, of Mansoura Medical School in Egypt, who served as the lead researcher of this project, said she had come up with the idea after meeting a 22-year-old cancer patient who had a high risk of infertility from chemotherapy. Dr. Mohamed said: “It was a very emotional for me so I decided to pursue it and work on it to figure it out. It [is] a very common problem based on statistics of cancer female diagnosis every year. “

Dr. Mohamed continued: “We inject[ed] stem cells in[to] the ovaries of mice which had chemotherapy and were damaged and we got very good ovarian function restoration in form of follicle number, hormonal production, and finally getting pregnant and having new pups, which was our ultimate goal.  We are now working on translating that into clinical trials (for humans).  This approach carries high promise to women with chemotherapy-induced and potentially other types of premature ovarian failure.”

Imperial College gynecologist Stuart Lavery said: “This is very exciting piece of research that adds to our understanding of how cells differentiate to become egg stem cells.” Dr. Lavery served as a consultant on this research. I must add at this point as an aside that it is rather unlikely that the bone marrow stem cells are differentiating into eggs. Instead the bone marrow stem cells are probably augmenting the survival and health of existing eggs in the ovary.

Dr. Lavery continued: “Clearly, there remains an enormous amount of work to see whether these results would be transferable into humans. But it does provide some realistic hope that post-chemotherapy patients who have been made menopausal could one day restore ovarian function and possibly fertility.”

Dr. Mohamed and her colleagues would like to initiate human trials using umbilical cord or even embryonic stem cells. They will need to convince regulatory agencies that the procedures they have designed are safe. For this reason, I find it unlikely in the extreme that the US Food and Drug Administration (FDA) would give approval for an embryonic stem cell-based trial in the ovaries, given the large numbers of regulatory and safety hurdles other recent embryonic stem cell-based trials have had to conquer. Also, it is worth noting that the FDA has not approved other proposed trials that sought to stimulate ovarian-based stem cells. For this reason, getting FDA approval for their trial might prove difficult. Also, one mouse experiment is not going to be enough to persuade the FDA to acquiesce to their proposals. Large experiments will need to be done and large animals studies would also be needed as well.

Women who opt to freeze their eggs can use in vitro fertilization (IVF) to have their own children. Alternatively, if the eggs are fertilized with her mate’s sperm, then the embryos can development to the blastocyst stage after which they are cryopreserved (frozen) before chemotherapy for later family-building purposes.

Such a strategy leads to some problems in countries with nationalized medicine: some provinces have decreased funding for IVF, since IVF is very expensive and the demand is below the cost to maintain such faculties. Likewise, at times, female cancer patients are denied the option of cryopreservation, again because of the costs and the lack of a nearby facility that has the space, means, or funding to keep her embryos on ice for a time. A new regenerative therapy might give such a female patient some solace with regards to her future fertility.

A consultant in Reproductive Medicine and Surgery at Hammersmith Hospital, London, Dr Geoffrey Trew, said of this research: “Fertility-wise, if this works it would be stupendous. Certainly it does appear promising and anything you can do to regenerate and ovary is a good thing. Theoretically if you are regenerating the ovary you should be getting better quality eggs. Clearly we’re not here yet, and it’s good that the researchers are not over-claiming their findings, but it’s a great proof of concept.”

Dr Edgar Mocanu, consultant gynecologist at Rotunda Hospital in Dublin and a board member of the International Federation of Fertility Societies, said: “This could open phenomenal opportunities for women. Millions of women around the world undergo cancer treatment and some of them will become infertile through ovarian failure. While cancer survival rates have increased dramatically, to date there is no effective method of preventing infertility after chemotherapy. It could also open new avenues for the treatment of menopause induced health issues.”

Dr Owen Davis president of the American Society for Reproductive Medicine: “If this experimental treatment can be translated to women who have lost ovarian function from chemotherapy, it will be a great advance. Restoring ovarian hormone production, follicle development and fertility to chemotherapy patients is a potential new application for bone marrow donation that could help many women.”

Anti-Aging Protein GDF11: Does it Work?

The protein is called GDF11 and some scientists claim that is can rejuvenate older laboratory animals and make them healthier. Sounds like science fiction, but could it be true?

Several decades ago, in the 1950s, some creative and enterprising scientists connected the circulatory systems of two inbred mice, one of which was old and the second of which was young. The blood from the young mouse seemed to rejuvenate the older mouse. That led to a question: “If blood from younger mouse rejuvenated the older mouse, what was it in the blood that did it?” Further work has landed on GDF11 as the rejuvenating protein, but the experimental path to this protein has been fraught with false starts, bumps, and wrong turns. New work by a team of Harvard University scientists hopes to set the record straight on GDF11.

Work by Harvard stem cell biologist Amy Wagers, cardiologist Richard Lee and the members of their laboratories and their collaborators have discovered that the blood concentrations of GDF11 drop in mice as they age. Such a finding is a correlation, which might be suggestive, but it falls short of proving that GDF11 is an anti-aging protein. However, Wagers and Lee and their colleagues also showed that when older mice are injected with GDF11, the protein partially reverses the thickening of the heart that comes with age. Wagers and her team also showed in two papers that were published in the journal Science that administration of GDF11 can rejuvenate the muscles and brains of older mice.

Wagers’ findings, however, received some push-back in May, 2015. According to Jocelyn Kaiser, writing at the Science web site, David Glass, who works at the Novartis Institutes for Biomedical Research in Cambridge, Massachusetts, and his colleagues have made use of an antibody that specifically binds to GDF11 to detect the protein and measure its concentration in the blood and tissues. Experiments with the anti-GDF11 antibody revealed that blood levels of GDF11 increase as rats and people get older. Also, in the hands of Glass and his team, injected GDF11 protein inhibited muscle regeneration in young mice. Furthermore, work from Steven Houser’s group at Temple University in Philadelphia, Pennsylvania, has shown that injections of GDF11 do not decrease the age-related thickening of the hearts of older mice. Now we have a genuine scientific controversy: so who’s right?

Wagers and Lee have concluded that the specific assay Novartis used to detect GDF11 and a related protein (GDF8 or myostatin) did not work properly. In their own experiments, the combined efforts of the Wagers and Lee teams showed that the main protein detected by the antibody test designed and used by the Glass group is immunoglobulin (antibodies). The levels of antibody proteins in the blood are known to rise in the blood as people get older. As a control, when the Wagers and Lee group used the Novartis-designed test to measure the proteins levels of laboratory mice that do not possess the gene that encodes antibodies, the blood of those mice tested negative. According to Jocelyn Kaiser, these data were published in a paper that appeared in the journal Circulation Research.

Wagers summarized the results of her and Lee’s laboratories, “They actually had very consistent findings to ours with respect to the blood levels of GDF11/8 with the antibody we all used.” However, according to Wagers, “their interpretation was confused by this case of mistaken identity.” To corroborate her point, Wagers cited a recently published study by scientists from the University of California, San Francisco, who found that GDF11/8 blood levels decline as people age, and are low in heart disease patients. These results support the hypothesis that GDF11 has antiaging activity.

The Harvard team’s paper also examined the results from the Houser laboratory. According to Wagers, Houser and his colleagues utilized commercially purchased GDF11, and this source of protein can vary in activity and levels. Wagers noted that it “wasn’t something that affected us early on, but we figured out it was an issue. The variability of commercially purchased GDF11 might explain why Houser and his colleagues were unable to see any results from injected GDF11. Houser and his team were quite careful to make sure that they injected the same dose of GDF11 as the Wagers and Lee. However, Wagers pointed out that if only a fraction of the protein was as active as the protein used by Wagers and Lee, then it is likely that Houser and his group actually used a lower effective dose than the Harvard group. Lee has also noted that he and his group have data that suggests that the GDF11 dose they used was actually higher than they initially thought.

Wagers and others also showed that daily injections of GDF11 can shrink heart muscle in both old and new mice, and, incredibly, the mice also lost weight. “We don’t have much insight into that right now, but we’re looking into it,” Wagers says. Wagers suspects that GDF11 only works within a particular therapeutic concentration, outside of which is will not work and above which it might cause side effects that are harmful.

What does the competition think? Houser thinks that Wager and Lee are probably correct that at least one of the assays used by the Novartis team to measure GDF11 detected immunoglobulin. However, both Houser David Glass have pointed out that the Novartis team used a different GDF11 detection assay whose accuracy was not challenged by the work in this new paper.

Houser remains sanguine about finding molecules that can delay aging.  “I’m going to be 65 in a couple months. I’d love to have something that improves my heart, brain, and muscle function,” said Houser. “I think the field is going to figure this out and this is another piece of the puzzle.”

The jury is still out when it comes to GDF11, but Wagers and Lee have made a positive contribution to a robust and thrillingly interesting scientific discussion.

Protein Regulates Heart Muscle Development

Scientists from the Center for Genomic. Regulation in Barcelona, Spain have discovered a genetic regulatory network that revolves around a protein called Mel18. This regulatory network acts as a genetic switch during the differentiation of embryonic stem cells into heart muscle cells.

Mel18 acts in combination with a vitally important set of proteins called the “Polycomb Regulatory Complexes” or PRCs. PRCs are probably one of the major repressors of genes in adult and embryonic stem cells, and in this paper, Luciano De Croce and his colleagues showed that Mel18 acts with the PRCs to suppress gene expression.

Beyond that, however, once differentiation occurs, Mel18 combines with other proteins to continue to shut off the expression of unnecessary genes, but during early cardiac development, Mel18 completely shifts and becomes a driver of gene expression. It shifts its function by forming new complexes with other proteins that regulate gene expression in various ways.

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Thus Mel18 acts as a genetic switch that guides stem cells into the cardiac fate and eventually into the heart muscle cell lineage.

This fascinating work, which was published in the journal Cell Stem Cell, can help stem cell scientists grow better heart muscle from induced pluripotent stem cells in the laboratory. It could also elucidate the underlying causes of heart defects in congenital heart disease. They may also lead to new ways of controlling stem cells in the laboratory to grow cellular repair kits and patches for patients with damaged or sick hearts.

New Gene Therapy for Retinitis Pigmentosa Treats Early and Late Stages of the Disease in Dogs

Collaboration between scientists from the University of Pennsylvania and the University of Florida, Gainesville has hit pay dirt when it comes to treating an inherited eye disease. This study used gene therapy to treat the disease and the results of this research project make a definitive contribution to the development of gene therapies for people with the blinding eye disorders for which there is presently no cure.

The disease in question is called retinitis pigmentosa, which is a group of rare, genetic disorders characterized by the degradation and subsequent loss of photoreceptors in the retina. People who suffer from retinitis pigmentosa have difficulty seeing at night and experience a loss of peripheral vision.

As mentioned, retinitis pigmentosa is an inherited disorder that results from mutations in any one of more than 50 different genes. These genes encode proteins that are required for retinal photoreceptors, and mutations in these genes compromises photoreceptor survival and function.

In human patients, retinitis pigmentosa is the most common inherited disease that results in degeneration of the photoreceptors of the retina. Approximately 1 in 4,000 people are affected with retinitis pigmentosa and 10 to 20 percent have a particularly severe form called X-linked retinitis pigmentosa. This disease predominately affects males, who experience night blindness by age 10 and progressive loss of the visual field by age 45. 70 percent of people with the X-linked retinitis pigmentosa harbor loss-of-function mutations in the retinitis pigmentosa GTPase Regulator (RPGR) gene. RPGR encodes a protein that maintains the health and survival of retinal photoreceptors. There are two types of photoreceptors; rods that give us the ability to see in dim light, and cones that allow us to see fine detail and color in bright light. Loss of the RPGR protein damages both types of photoreceptors.

Because there are no treatments for retinitis pigmentosa, gene therapy might be the best option to treat this disease. Fortunately, some varieties of dogs have a naturally occurring, late-stage retinitis pigmentosa that closely resembles the human disease. In previous experiments, gene therapies were used in diseased dogs, but such studies showed that benefits from gene therapy were only observed when it was used in the earliest stages of the disease.

“The study shows that a corrective gene can stop the loss of photoreceptors in the retina, and provides good proof of concept for gene therapy at the intermediate stage of the disease, thus widening the therapeutic window,” said Neeraj Agarwal, Ph.D., a program director at National Eye Institute, a part of the National Institutes of Health, who funded this research.

The dogs used in this study all suffered from a naturally occurring canine form of RPGR X-linked retinitis pigmentosa that is observed in some mixed breeds. These animals provided an excellent model system for their gene therapy tests, since affected dogs with early to late stages of the disease could be treated with the experimental therapy in one eye while the other untreated eye could be evaluated in parallel as a control.

To treat these blind dogs, the team utilized adeno-associated virus (AAV). They engineered AAV particles that possessed the entire RPGR gene. Then they devised a way to deliver these viruses to the retinal cells so that the viruses could infect the retinal cells and produce normal copies of the RPGR protein.

When the eyes treated with the AAV vectors were subjected to detailed imaging, it was clear that the gene therapy protocol arrested the thinning of the retinal layer. This shows that the treatment halted the degeneration of the photoreceptors in the affected dogs. When the treated eyes were compared with the untreated eye, the structure of the rod and cone photoreceptors was obviously improved and better preserved in the treated eye in comparison to the untreated eye. When the neural physiology of the retinas from the treated and untreated eyes was compared, once again, the retinas from the eyes treated with the gene therapy were more normal than the untreated eyes. In fact, the gene therapy halted the photoreceptor cell death associated with retinitis pigmentosa for two and a half years, which was the length of the study.

The team also treated dogs who suffered from later-stage disease in the hope that the gene therapy could not only improve the condition of dogs in the early stages of the disease, but also those with later stages of the disease. Interestingly, the gene therapy also froze the loss of retinal thickness and preserved the structure of surviving photoreceptors, but the retinas in the untreated eyes continued to thin and their photoreceptor function deteriorated as well. When the dogs were sent through an obstacle course and a maze under dim light, the animals did significantly better when they used their eye that had been treated with the gene therapy compared with their performance when they used the untreated eye. This shows that this gene therapy also works in dogs suffering from the late-stages of retinitis pigmentosa.

Can such a therapy be used in people in human clinical trials? Not yet. More safety testing must be done in order to properly determine if it is safe over long periods of time, according to this study’s co-leaders, Gustavo Aguirre, V.M.D., Ph.D., and William Beltran, D.V.M., Ph.D., of the University of Pennsylvania. Other collaborators, University of Pennsylvania scientists Artur Cideciyan, Ph.D., and Samuel Jacobson, M.D., Ph.D. are presently screening potential patients who have RPGR mutations as a prolegomena for a future clinical trial.

Their results are published in Proceedings of the National Academy of Sciences.

Pre-treatment of MSCs Can Reduce Their Regenerative Properties

Mesenchymal stem cells (MSCs) are excellent suppressors of unwanted inflammation.  This anti-inflammatory activity has been established for systemic inflammatory diseases in animal experiments (Klinker MW, Wei CH. World J Stem Cells. 2015 Apr 26;7(3):556-67), and in clinical trials with human patients (Dulamea A. J Med Life. 2015 Jan-Mar;8(1):24-7; Simonson OE et al., Stem Cells Transl Med. 2015 Oct;4(10):1199-213. doi: 10.5966/sctm.2015-0021).  Stem cell researchers have also shown that MSCs can suppress inflammation in the bowel (see Swenson E and Theise N. Clinical and Experimental Gastroenterology 2010;3:1-10; Chen Z, et al., Biochem Biophys Res Commun. 2014 Aug 8;450(4):1402-8).

After being introduced into the body of a patient, MSCs to move to the site where they are needed (a phenomenon known as “homing”) and promote tissue repair and healing.  Sometime MSC homing works quite well, but other times, it is so-so.  Therefore, several inventive scientists have devised ways to beef up homing to specific sites in order to improve MSC-based tissue healing.  Also, investigators are equally interested in increasing the ability of MSCs to stick to tissues once they arrive there to ensure that the homed MSCs stay where they are needed (see Kavanagh DP, Robinson J, and Kalia N. Stem Cell Rev 2014;10:587-599).  Unfortunately, at the moment, the whole homing process is a bit of a black box and while artificially increasing homing might help in the laboratory, whether or not it increases the therapeutic benefit of MSCs is even less well understood.

A new report from the laboratory of Neena Kalia, who works at the University of Birmingham, UK, has examined the effect of artificial enhancement on the therapeutic capacity of MSCs to treat inflammation in the bowel.  This is an important study because pre-treatment strategies have been suggested as ways to boost MSC homing and retention to various tissues.  The Kalia study suggests such pre-treatment strategies should be viewed with a degree of skepticism.

In this study, Kalia her coworkers induced inflammation in the gastrointestinal tracts of mice by clamping off the blood supply to the this tissue for a time and then releasing the clamps and letting the blood flow anew.  This type of damage, known as ischemia/reperfusion (IR) injury deprives cells of vital oxygen and nutrients for a short period of time, which causes some cells to die.  When the blood is allowed to flow into the tissue, inflammation is induced in the damaged tissue.  Therefore, this technique can efficiently induce  inflammation in tissues in the gastrointestinal tract.

Two groups of mice were treated with bone marrow-derived MSCs.  One group had experiences IR injury to their gastrointestinal tracts, and the other group did not.  In these experiments, administered MSCs showed similar levels of and cell adhesion in both injured and non-injured guts.  In general, cell adhesion levels were nothing to write home about:  as reported in the paper, “limited cell adhesion observed.”  Despite these initial observations, those MSCs that found their way to the gut were able to help heal the tissues to some degree.  There were fewer white blood cells in the middle part of the small intestine (jejunum), and the degree of blood flow seemed to have improved.  Unfortunately, the lower part of the small intestine (ileum) was not helped to the same degree, and the paper reports that a fair number of MSCs got stuck in small blood vessels, which suggests that these vessels got stuck on their way to the intestine.

If these results seem underwhelming, it might be because they are.  Undaunted, Kalia and her crew tried to boost the regenerative abilities of their isolated MSCs by pretreating them.  Kalia’s laboratory and other laboratories as well have used a variety of chemical agents to augment the healing abilities of MSCs.  These agents include things like tumor necrosis factor (TNF)-α, CXCL12 (also known as stromal cell-derived factor 1 or SDF1, which strongly activates white blood cells), interferon (IFN)-γ, or hydrogen peroxide.  When these pre-treated MSCs were administered to mice whose guts were damaged by means of IR injury, the pretreatment not only did not enhance their intestinal recruitment, but actually decreased the healing capacities of MSCs.  Pretreatment of MSCs with tumor necrosis factor (TNF)-α, CXCL12, interferon (IFN)-γ, or hydrogen peroxide did not enhance their intestinal recruitment.  Pretreatment with TNFα and IFNγ abrogated ability of transplanted MSCs to reduce white blood cells infiltration and improve blood flow in the jejunum.

Kalia and her colleagues utilized a technique called “intravital” microscopy for this study.  Intravital microscopy can track individual cells in a living animals (Kavanagh DP, Yemm AI, Zhao Y, et al. PLoS One 2013;8:e59150). With this technique, they were able to efficiently monitor adhesion in the tinyu blood vessels in the injured intestinal tissue.  They documented poor MSC adhesion to the gut lining and that pre-treatment with various factors hopes failed to enhance adhesion of MSCs to the gut.

This study successfully demonstrated that MSCs can rapidly limit white blood cells recruitment to the inflamed gut, and improve tissue perfusion if they are administered after intestinal IR injury. However, Kalia’s study also shows that strategies to improve MSC therapeutic efficacy by means of pretreatment of MSCs may not be all it’s cracked up to be.  They suggest that in the future, cytokine or chemical pretreatments designed to enhance MSC recruitment and function will require more than just successful experiments in a cell culture system.  Instead, pretreatment strategies will need to be carefully validated in living organisms in order the confirm that such protocols help rather than hinder the therapeutic function of implanted stem cells.

This paper was published in the journal Stem Cells – Kavanagh DP, Suresh S, Newsome PN, et al. Stem Cells 2015;33:2785-2797

A New Target for Treating Stroke: The Spleen

If the blood vessels of the brain become plugged as a result of a clot or some other obstructive event, then the brain suffers a trans-ischemic attack (TIA), which is more commonly known as a stroke. The initial stroke starves brain cells of oxygen, which causes cell death by suffocation. However, dying brain cells  often spill enormous amounts of lethal material into the surrounding area, which kills off even more brain cells. Worse still, these dead or dying called can induce inflammation in the brain, which continues to kill off brain cells.

New work, however, from the laboratory of César Borlongan at the University of Southern Florida in Tampa, indicates that the spleen may be a target for treating the stroke-induced chronic inflammation that continues to kill brain cells after the initial stroke.

At the University of Florida Center of Excellence for Aging and Brain Repair, a study found that the intravenous administration of human bone marrow stem cells to post-stroke rats reduced the inflammatory-plagued secondary cell death associated with stroke progression in the brain. The intravenously administered cells preferentially migrated to the spleen where they reduced this post-stroke inflammation.

This study answers some of the perplexing questions surrounding animal experiments that used stem cells to treat stroke. Typically, stem cell administration to animals that suffered an artificially-induced stroke causes some functional recovery, but when their brains are examined for the stem cells that were implanted into them, very few surviving cells are observed.

“Our findings suggest that even if stem cells do not enter the brain or survive there, as long as the transplanted cells survive in the spleen the anti-inflammatory effect they promote may be sufficient enough to therapeutically benefit the stroke brain,” said César Borlongan, principal investigator of this study.

Stroke is the leading cause of death and the number one cause of chronic disability in the United States, yet treatment options are limited.

Stem cell therapy has emerged as a potential treatment for ischemic stroke, but most pre-clinical studies have examined the effects of stem cells transplanted during acute stroke (one hour to three hours aster the onset of the stroke).

In the wake of an acute stroke, an initial brain lesion forms from the lack of blood flow to the brain. The blood-brain barrier is also breached and this allows the infiltration of inflammatory molecules that trigger secondary brain cell death in the weeks and months that follow. This expanded inflammation is the hallmark of chronic stroke.

In this study, Borlongan and his colleagues intravenously administered human bone marrow stem cells 60 days after the onset of a stroke. Thus these animals were well into the chronic stroke stage.

The transplanted stem cells predominantly homes to the spleen. In fact, Borlongan and his crew found 30-times more cells in the spleens of the animals than in the brain.

While in the spleen, the stem cells squelched the production of a protein called tumor necrosis factor, which is a major inflammatory signal that increases in concentration after a stroke. The reduction of the tumor necrosis factor signal prevented the macrophages and other immune cells from leaving the spleen and going to the brain. This reduced systemic inflammation and decreased the size of the lesions in the brain caused by the stroke. There was also a trend toward reduced neuronal death and smaller decreases in learning and memory in the laboratory animals.

Borlongan explained that during the chronic stage of stroke, macrophages seem to fuel inflammation. “If we can find a way to effectively block the fuel with stem cells, then we may prevent the spread of damage in the brain and ameliorate the disabling symptoms many stroke patients live with,” said Borlongan.

Borlongan and his team hope to test whether transplanting human bone marrow stem cells directly into the spleen will lead to behavioral recovery in post-stroke rats.

One drug that has been approved for the emergency treatment of stroke is tPA or tissue plasminogen activating factor, which activates the blood-based protein plasminogen to form the highly active enzyme, plasmin. Plasmin is a powerful dissolved of clots, but tPA must be administered less than 4.5 hours after the onset of ischemic stroke, and benefits only three to four percent of patients.

Even though more work needs to be done, evidence from the USF group and other neurobiology groups indicates that stem fells may provide a more effective treatment for stroke over a wider time frame.

Targeting the spleen with stem cells or the anti-inflammatory molecules they sec rate offers hope for treating chronic neurodegenerative diseases like stroke at later stages.

This study, which was published in the journal Stroke, shows that it is possible to arrest the chronic inflammation that characterizes chronic stroke 60 days after the initial stroke. If such a result can be replicated in human patients, it will indeed be a powerful thing, according the Sandra Acosta, the first author on this paper.

Laboratory-Grown Intestine Shows Promise in Mice and Dogs

David Hackam is a pediatric surgeon at the Johns Hopkins Children’s Center. Unfortunately, Dr. Hackam spends a good deal of his time removing dead sections of intestine from sick babies, but he would deeply love to be able to do more than just take out intestines but actually replace the dead or dying intestinal tissue. It is that desire that has driven Hackam and his colleagues to grow intestines in the laboratory.

They begin with stem cells taken from the small intestines of human infants and mice and apply them to intestine-shaped scaffolds. The stem cells dig in, grow and form mini-intestines that just might be able to treat disorders like necrotizing enterocolitis and Crohn’s disease someday. Transplantation experiments in laboratory animals have shown that this laboratory-grown tissue and scaffolding are not rejected, but integrate into the tissues of the animals. Experiments in dogs have shown that the scaffold allowed dogs to heal from damage to the colon lining, essentially restoring healthy bowel function.

The study is a “great breakthrough,” says Hans Clevers, a stem cell biologist at the Hubrecht Institute in Utrecht, the Netherlands, who was not involved in the new research. Clevers and his colleagues were the first to identify stem cells in the intestine, and his lab developed the technique Hackam’s team used to grow intestinal tissue.

Making replacement organs by growing cells on scaffolds molded into the shape of the organ is not a new idea, since other researchers have used exactly this technique to make bladders and blood vessels. However, the laboratory-grown intestines made by Hackam and his group come closer to the shape and structure of a natural intestine than anything created in the laboratory before. In previous experiments carried out in other laboratories, the gut lining has been grown on flat scaffolds or in culture flasks. Under these conditions, the tissue tends to roll up into little balls that have the absorptive surface on the inside. Hackam and his coworkers, however, overcame this problem by using a scaffold fabricated from materials similar to surgical sutures. This material can be molded into any desired intestinal size and shape, and in Hackam’s hands, the scaffolds formed a true tube-shaped (like a real gut), with tiny projections on the inner surface that can help the tissue form functional small intestinal villi (the small fingers of tissue that increase the surface area of the intestine to increase nutrient absorption. “They can now make sheets of cells that can be clinically managed,” Clevers says. “Surgeons can handle these things and just stick them in.”

To grow the gut lining in the lab, the researchers painted the scaffold with a sticky collagen-rich substance and then dripped onto it a solution of stem cells from the small intestine. This concoction was grown in a culture system for a week. Interestingly, Hackam and his team found that if they added connective tissue cells, immune cells, and probiotics (bacteria that help maintain a healthy gut), all of these things helped the stem cells mature and differentiate.

Hackam’s group also sutured intestines grown from mouse stem cells into the tissue surrounding the abdominal organs of the mouse. The lab-grown intestines developed their own blood supply and normal gut structures despite the fact that they were not connected to the animals’ digestive tract. “Using the mouse’s own stem cells, we can actually create something that looks just like the native intestine,” Hackam says. The next step, he says, is “to hook it up.”

Before “hooking it up,” Hackam needed to be sure that the scaffold could be tolerated in living animals. Therefore he tested the new scaffold in dogs. He removed sections of large intestinal lining and replaced it with pieces of scaffolding. The dogs made a complete recovery: their gut lining regrew onto the scaffold and functioned normally to absorb water from the colon. After a few weeks, the scaffolding had completely dissolved and was replaced with normal connective tissue. “The scaffold was well tolerated and promoted healing by recruiting stem cells,” Hackam says. “[The dogs] had a perfectly normal lining after 8 weeks.”

This technique could help more than just dogs and mice, but could aid human patients. According to Hackam, scaffolds could be custom-designed for individual human patients to replace a portion of an intestine or the entire organ. This could be a revolutionary treatment for patients with necrotizing enterocolitis, a condition that destroys intestinal tissue in about 12% of premature babies in the United States. It could also potentially repair the intestines of patients with Crohn’s disease, an inflammatory bowel disorder that can have life-threatening complications and that affects more than 500,000 people in the United States. However, these lab-grown intestines must pass several other tests before they are ready for human clinical trials, Hackam cautions.

The first test that these laboratory-grown intestines must pass is the absorption test. Laboratory-grown small intestines must be transplanted into live animals and they must properly absorb food. Also, the technology that is used will also require some adjustments. For example, Mari Sogayar, a molecular biologist at the University of São Paulo in Brazil, points out that the collagen product that helps the stem cells stick to the scaffold is not meant for use in people. In the next experiments, Hackam says, the researchers plan to use a surgical-grade alternative.

“I take care of children who have intestinal deficiencies, eating deficiencies, and they are very much at wits’ end,” Hackam says. “I think what we can offer in the scientific community is a path toward something that one day will help a child.”

Hydrogels Help Implanted Stem Cells Survive in the Heart

How do you get stem cells to survive after they have been transplanted? You can pre-condition them, but research from Johns Hopkins University has capitalized on a different strategy. The Hopkins team used hydrogel to protect and feed the stem cells that had been implanted into the heart.

They utilized a rat model system for this work. Rats that had been given heart attacks were given stem cell implants encased in a hydrogel. The hydrogel supported stem cells survival and also kept the stem cells at the site of their implantation where they re-muscularized the damaged heart muscle. 73% of the stem cells embedded in hydrogel survived whereas only 12% of the non-hydrogel-embedded stem cells survived after injection into the heart.

Previously, stem cell injections have been shown to aid damaged heart tissue, but the vast majority of the injected cells die or are washed from the heart into other tissues. Hydrogel, which mostly consists of water, allows the cells to live and grow while they integrate into the surrounding tissue and initiate healing.

Heart-damaged rats injected with hydrogel-loaded stem cells saw a 15% increase in pumping efficiency for the treated ventricle, compared with just 8% for regular stem cell therapies. Hydrogen can support both adult and embryonic stem cells, and if it’s not put inside a living being, the hydrogel can actually maintain 100% of the stem cells embedded in them.

Hydrogels are useful in biology because they are safe for use in living organisms. In fact, this study found that injecting the hydrogel alone, with no stem cells at all, had a mild benefit all its own by promoting new blood vessel growth.

These are the sorts of breakthroughs that will allow the stem cell technologies of today to become the amazing stem cell technologies of tomorrow.

Rejection of Induced Pluripotent Stem Cell Derivatives By the Immune System is a Function of Where They are Transplanted

Induced pluripotent stem cells (iPSCs) are made from mature, adult cells by a combination of genetic engineering and cell culture techniques. Master genes are transfected into mature cells, which are then cultured as they grow and revert to more immature states. Eventually, a population of cells grow in culture that have some, though not all of the characteristics, of embryonic stem cells. Because these cells are pluripotent, they should, theoretically have the ability to differentiate into any adult cell type. Also, since they are derived from a patient’s own cells, they should be tolerated by the patient’s immune system and should not experience tissue rejection.I

Or should they? Experiments with cells derived from iPSCs have generated mixed results. If C57BL/6 (B6) mice are transplanted with iPSC-derived cells, such cells show some levels of recognition by the immune system. However, another study has concluded that various lineages of B6 iPSC-derived cells are not recognized by the immune system when transplanted under the kidney capsule of B6 mice. Why the contradiction?

Yang Xu and his colleagues at the University of California, San Diego have attempted to resolve this controversy by utilizing a mouse model system. Xu and his colleagues used the same B6 transplantation model and transplanted a variety of different cells derived from iPSCs that were made from cells that came from the same laboratory mice.

Xu and others showed that iPSC-derived and embryonic stem cell (ESC)-derived cells are either tolerated or rejected, depending upon WHERE they are transplanted. You see the immune system depends upon a network of cells called “dendritic cells” to sample the fluids that circulate throughout the body and identify foreign substances. Some locations in our bodies are chock-full of dendritic cells, while other locations have a paucity of dendritic cells. When iPSC or ESC-derived cells are transplanted under the kidney capsule, they survive and thrive. The kidney capsule has a distinct lack of dendritic cells. However, if these same cells, which were so nicely tolerated under the kidney capsule, are transplanted under the skin or injected into muscles, they were rejected by the immune system. Why? These two sites are loaded with dendritic cells.

Therefore, the rejection of iPSC-derived cells by the patient’s body is more of a function of where the cells are transplanted than the cells themselves. Mind you, poor quality iPSCs can produce derivatives that are rejected by the immune system, but high-quality iPSCs can differentiate into cells that are accepted by the immune system, but it is wholly dependent on where they are transplanted.

Perhaps, transplanted IPSC derivatives will need the immune system suppressed for a short period of time and after they become integrated into the patient’s body, the immune suppression can be lifted. Alternatively it might be possible to induce tolerance to the transplanted cells with immunological tricks. Either way, understanding why iPSCs-derived cells are rejected or accepted by the patient’s immune system is the next step to using these amazing cells for regenerative medicine.

Xu’s paper appeared in the journal Stem Cells – DOI: 10.1002/stem.2227.

The Ideal Recipe for Cartilage from Stem Cells

Researchers at Case Western Reserve and Harvard University will use a 5-year, $2-million NIH grant to build a microfactory that bangs out the optimal formula for joint cartilage. Such an end product could one day potentially benefit many of the tens of thousands of people in the United States who suffer from cartilage loss or damage.

Joint cartilage or articular cartilage caps the ends of long bones and bears the loads, absorbs shocks and, in combination with lubricating synovial fluid, helps knees, hips, shoulders, and other joints to smoothly bend, lift, and rotate. Unfortunately, this tissue has little capacity to regenerate, which means that there is a critical need for new therapeutic strategies.

Artificial substitutes cannot match real cartilage and attempts to engineer articular cartilage have been stymied by the complexities of directing stem cells to differentiate into chondrocytes and form the right kind of cartilage.

Stem cells are quite responsive to the environmental cues presented to them from their surroundings. What this research project hopes to determine are those specific cue that drive stem cells to differentiate into chondrocytes that make the right kind of cartilage with the right kind of microarchitecture that resembles natural, articular cartilage. To do this, they will engage in a systematic study of the effects of cellular micro-environmental factors that influence stem cell differentiation and cartilage formation.

Bone marrow- and fat-derived mesenchymal stem cells have been differentiated into cartilage-making chondrocytes in the laboratory. These two stem cell populations are distinct, however, and required different conditions in order to drive them to differentiate into chondrocytes. This research group, however, has designed new materials with unique physical properties, cell adhesive capabilities, and have the capacity to deliver bioactive molecules.

By controlling the presentation of these signals to cells, independently and in combination with mechanical cues, this group hopes to identify those most important cues for driving cells to differentiate into chondrocytes.

Ali Khademhosseini specializes in microfabrication and micro-and nano-scale technologies to control cell behavior. He and his team will develop a microscale high-throughput system at his laboratory that will accelerate the testing and analysis of materials engineered in another laboratory.

This research cooperative hopes to test and analyze more than 3,000 combinations of factors that may influence cell development, including differentiation, amounts of biochemicals, extracellular matrix properties, compressive stresses, and more. Khademhosseini and his colleagues hope to begin testing comditions identified from these studies in animal models by the of the grant term.

Scientists Reprogram Adult Skin Cells to Make Mini Kidneys

Japanese and Australian researchers have used induced pluripotent stem cell (iPSC) technology to reprogram human skin cells to make the most mature human kidneys yet to be grown in a culture. These mini kidneys have hundreds of filtering units (nephrons) and blood vessels and appear to be developing just as kidneys would in an embryo.

“The short-term goal is to actually use this method to make little replicas of the developing kidney and use that to test whether drugs are toxic to the kidney,” said lead researcher Professor Melissa Little, of the Murdoch Children’s Research Institute. “Ultimately we hope we might be able to scale this up so we can … maybe bioengineer an entire organ.”

In other previous research, Professor Little and her co-workers generated cells that self-organized into the nephrons and collecting ducts needed for the kidney to filter blood and produce urine. They used a precise combination of called growth factors to direct embryonic stem cells to develop into the different cell types.

In the journal Nature, Professor Little and her collaborators report they have made a developing kidney from a type of skin cell called a fibroblast. Little and her team reprogrammed adult fibroblasts to become “induced pluripotent stem cells,” which act like embryonic stem cells, and can become any cell in the body. By adopting their growth factor recipe, Little and others were able to grow these cells into larger and more complex, three-dimensional kidneys than previously made.

“These kidneys have something like 10 or 12 different cell types in them … all from the one starting stem cell,” said Professor Little. “What we had previously were little flat structures over the surface of a dish … Now we have an organoid that is about 5-6 millimetres across, has about 100 filtering units in it, and is starting to form blood vessels. It’s starting to mature and the cell types are starting to do more of the functions of the final kidney.”

Scientists in Little’s laboratory demonstrated that the genes expressed in the mini kidneys as they formed faithfully recapitulated the expression of those same genes in a developing kidney in a first trimester embryo.

“It is actually mirroring what is happening in human development,” said Professor Little.

Little and her group also found that the laboratory-grown kidney was damaged when it was treated with known renal toxins. Little suggested that the iPSCs cells they had created were functioning as a kidney, but further tests would be required to demonstrate that.

It might be possible to use these bioengineered kidneys to test the renal toxicity of drugs. Likewise, the production of mini kidneys using cells from kidney patients might provide a way to study inherited forms of kidney disease.

“You can take a fibroblast [from someone with inherited kidney disease], make a stem cell out of it, generate a little kidney and use that as our model for their disease,” said Professor Little.

Perhaps most exciting, laboratory-generated kidneys might one day provide rejection-free transplants for patients, and gene editing could be used to fix the genetic defect that caused an inherited kidney disease.

Professor Jamie Davies of the University of Edinburgh, who was not involved with this work, but commented on it for Nature, emphasized this was not a full-fledged, functional kidney. “The structure’s fine-scale tissue organization is realistic, but it does not adopt the macro-scale organization of a whole kidney. For example, it is not ‘plumbed’ into a waste drain, and it lacks large-scale features that are crucial for kidney function, such as a urine-concentrating medulla region. There is a long way to go until clinically useful transplantable kidneys can be engineered, but [this] protocol is a valuable step in the right direction.”

Davies also mentioned that these mini kidneys had the potential to replace “poorly predictive” animal drug safety tests, and called on researchers to team up with toxicologists to test the potential of their system.

Transplantation of Unique, Newly Discovered Stem Cells May Lead to Promising Stroke Therapy

Stroke treatments have seen some remarkable advances in the past few years. Stem cell treatments for stroke have even seen some successes in clinical trials, showing that stem cell transplantation aimed at neural repair after a stroke is a possible way to ameliorate the effects of stroke.

Now, collaboration between teams of American and Japanese researchers has shown that a newly-identified stem cell has the ability to successfully treat stroke in rats. When administered to rats who have suffered from an experimentally-induced stroke, MUSE or multilineage-differentiating stress-enduring cells induced the regeneration of neurons and resulted in “significant improvements in neurological and motor functions” compared to control groups that were not transplanted with MUSE cells. MUSE cells also do not cause tumors.

The study has increased the number of therapeutic arrows in the quiver of neurologists and neuroscientists and lengthens the list of cells that might one day be considered for human clinical trials if continued pre-clinical tests prove successful. Future clinical studies aimed at regenerating neurological and motor function in patients who have suffered ischemic stroke.

The paper describing this study appeared in a recent issue of Stem Cells (Sept. 2015).

“Muse cells are unique stem cells that are able to self-renew and display high-efficiency for differentiating into neuron-like cells,” explained lead author Dr. Cesar V Borlongan, Distinguished Professor and Vice-Chairman for Research at the University of South Florida (USF) College of Medicine Department of Neurosurgery and Brain Repair and Director of USF’s the Center of Excellence for Aging and Brain Repair. “Unlike mesenchymal stem cells (MSCs) that have previously been used in stem cell transplantation in stroke-related clinical trials, in the present study Muse cells were found to possess functional characteristics of neurons as they attain the attributes of the host microenvironment. When MUSEcells were transplanted into to the brains of rats modeled with stroke, they attained neuronal characteristics.”

MUSE cells are found in many different tissues, including bone marrow, skin and fat. Since these cells can be derived from dermal fibroblasts (a type of connective tissue cell that provides the structural framework for animal tissues and plays a critical role in wound healing), they can be accessed with relative ease, without the need for the painful, invasive procedures required for obtaining other kinds of stem cells. Furthermore, while some stem cells used in stem cell transplantation studies have been found to cause cancer, MUSE cells do not produce tumors and exhibit exceptional tissue repair potential when introduced into the blood stream.

Some researchers think that fetal stem cells might be better candidates for replacing lost neural circuitry. The main reason in favor of fetal stem cells is that they preferentially differentiate into neuronal cells. However, the accessibility to fetal stem cells is limited and, like embryonic stem cells, the immaturity of these cells may present safety issues, such as tumor development. Additionally, the use of fetal and embryonic stem cells has many ethical difficulties to say the least. Since MUSE cells can be derived from adult tissue rather than fetal or embryonic tissue, the ethical quandaries associated with using them is minimal.

Not only do MUSE cells also have the practical advantage of being non-tumorigenic, they are readily accessed commercially and can also be easily collected from patient skin biopsies. MUSE cells also do not have to be “induced,” or genetically manipulated in order to be used, since they already display inherent stem cell properties after isolation. MUSE cells also spontaneously home toward the stroke-damaged sites.

“Ours is the first study to show that human skin fibroblast-derived Muse cells can have neuron-like function, possess an inherent ability to assume ‘stemness’ properties, and to readily differentiate into neural-lineage cells after integration into the stroke brain,” said co-lead author Dr. Mari Dezawa, Department of Stem Cell Biology and Histology, Tohoku University Graduate School of Medicine in Sendai, Japan. “Our results show that Muse cells are a feasible and promising source for cell-based approaches to ischemic stroke therapy.”

Clincal Trial Validates Stem Cell-Based Treatments of Sickle Cell Disease in Adults

Santosh Saraf and his colleagues at the University of Illinois have used a low-dose irradiation/alemtuzumab plus stem cell transplant procedure to cure patients of sickle-cell disease. 12 adult patients have been cured of sickle-cell disease by means of a stem cell transplantation from a healthy, tissue-matched donor.

This new procedure obviates the need for chemotherapy to prepare the patient to receive transplanted cells and offers the possibility of curing tens of thousands of adults from sickle-cell disease.

Sickle cell disease is an inherited disease that primarily affects African-Americans born in the United States. The genetic lesion occurs in the beta-globin gene that causes hemoglobin molecules to assemble into filaments under low-oxygen conditions. These hemoglobin filaments deform red blood cells and cause them to plug small capillaries in tissues, causing severe pain, strokes and even death.

Fortunately, a bone marrow transplant from a healthy donor can cure sickle-cell disease, but few adults undergo such a procedure because the chemotherapeutic agents that are given to destroy the patient’s bone marrow leaves from susceptible to diseases, unable to make their own blood cells, and very weak and sick.

Fortunately, a gentler procedure that only partially ablate the patient’s bone marrow was developed at the National Institutes of Health ()NIH) in Bethesda, Maryland. Transplant physicians there have treated 30 patients, with an 87% success rate.

In the Phase I/II clinical trial at the University of Illinois, 92% of the patients treated with this gentler procedure that was developed at the NIH.

Approximately 90% of the 450 patients who received stem cells transplants for sickle-cell disease have been children. However, chemotherapy has been considered too risky for adult patients who are often weakened far more than children by it.

Adult sickle-cell patients live an average of 50 years with a combinations of blood transfusions and pain medicines to manage the pain crisis. However, their quality of life can be quite low. Now, with this chemotherapy-free procedure, adults with sickle-cell disease can be cured of their disease within one month of their transplant. They can even go back to work or school and operate in a pain-free fashion.

In the new procedure, patients receive immunosuppressive drugs just before the transplant, with a very low dose of whole body radiation. Alemtuzumab (Campath, Lemtrada) is a monoclonal antibody that binds to the CD52 glycoprotein on the surfaces of lymphocytes and elicits their destruction, but not the hematopoietic stem cells that gives rise to them.  Next, donor cells from a healthy a tissue-matched sibling or donor are transfused into the patient. Stem cells from the donor home to the bone marrow and produce healthy, new blood cells in large quantities. Patients must continue to take immunosuppressive drugs for at least a year.

In the University of Illinois trial, 13 patients between the ages of 17-40 were given transplants from the blood of a healthy, tissue-matched sibling. Donors must be tested for human leukocyte antigen (HLA) markers on the surfaces of cells. Ten different HLA markers must match between the donor and the recipient for the transplant to have the best chance of evading rejection. Physicians have transplanted two patients with good HLA matches, to their donor, but had a different blood type than the donor. In many cases, the sickle cells cannot be found in the blood after the transplant.

In all 13 patients, the transplanted cells successfully engrafted into the bone marrow of the patients, but one patient failed to follow the post-transplant therapy regimen and reverted to the original sickle-cell condition.

One year after the transplantation, the 12 successfully transplanted patients had normal hemoglobin concentrations in their blood and better cardiopulmonary function. They also reported significantly less pain and improved health and vitality,

For of the patients were able to stop post-transplantation immunotherapy, without transplant rejection or other complications.

“Adults with sickle-cell disease can be cured with chemotherapy – the main barrier that has stood in the way for so long,” said Damiano Rondelli, Professor of Medicine and Director of the Stem Cell Transplantation Program at the University of Illinois. “Our data provide more support that this therapy is safe and effective and prevents patients from living shortened lives, condemned to pain and progressive complications.”

These data were published in the journal Biology of Blood and Marrow Transplantation, 2015; DOI 10.1016/j.bbmt.2015.08.036.

Rebooting Pancreatic Cells Can Normalize Blood Sugar Levels in Diabetic Mice

Type 1 diabetes results from the inability of the endocrine portion of the pancreas to secrete sufficient quantities of the hormone insulin. The cells that make insulin, beta cells, have been destroyed. Consequently, type 1 diabetics must inject themselves with insulin routinely in order to stay alive. Is there a better way?

A new strategy suggests that maybe pancreatic cells can be “rebooted” to produce insulin and that sure reprogramming could potentially help people with type 1 diabetes manage their blood sugar levels without the need for daily injections. This therapeutic approach is simpler and potentially safer than giving people stem cells that have been differentiated into pancreatic beta cells.

Philippe Lysy at the Cliniques Universitaires Saint Luc, which is part of the Catholic University of Louvain in Belgium, and his colleagues have reprogrammed pancreatic duct cells extracted from dead donors who were not diabetic at the time of death. The duct cells do not produce insulin, but they have a natural tendency to grow and differentiate into specific types of cells.

Lysy and his team grew the cells in the laboratory and encouraged them to become insulin-producing cells by exposing them to fatty particles. These fatty particles are absorbed into the cells after which they induce the synthesis of the MAFA transcription factor. MAFA acts as a genetic “switch” that binds to DNA and activates insulin production.

Implantation of these altered cells into diabetic mice showed that the cells were able to secrete insulin in a way that controls blood sugar levels. “The results are encouraging,” says Lysy.

Lysy’s colleague, Elisa Corritore, reported these results at this week’s annual meeting of the European Society for Pediatric Endocrinology in Barcelona, Spain. Lysy and others are preparing to submit their results for publication.

This work, if it continues to pan out, might lead to the harvesting of pancreatic ducts from deceased donors and converted in bulk into insulin-making cells. Such “off-the-shelf” cells could then be transplanted into people with type 1 diabetes to compensate for their inability to make their own insulin.

“We would hope to put the cells in a device under the skin that isolates them from the body’s immune system, so they’re not rejected as foreign,” says Lysy. He says devices like this are already being tested for their ability to house insulin-producing cells derived from stem cells.

Previous attempts to get round this problem have included embedding insulin-producing cells in a seaweed derivative prior to transplantation in order to keep them from being destroyed by the recipient’s immune system.

Lysy thinks that since insulin-producing cells originate from pancreatic tissue, they have an inherently lower risk of becoming cancerous after the transplant. This has always been a worry associated with tissues produced from embryonic stem cells, since these have the capability to form tumors if any are left in their original state in the transplanted tissue.

The basic premise of the work looks solid, says Juan Dominguez-Bendala, director of stem cell development for Translational Research at the University of Miami Miller School of Medicine’s Diabetes Research Institute in Florida. “However, until a peer-reviewed manuscript is published and all the details of the work become available to the scientific community, it is difficult to judge if this advance represents a meaningful leap in the state of the art.”

Lysy expects it will take between three and five years before the technique is ready to be tested in human clinical trials.