Biomedical engineers from Tufts University have discovered a new protocol that can induce mesenchymal stem cells (MSCs) derived from bone marrow to differentiate into neuron-like cells by treating them with exosomes from cultured cells.
PC12 cells are neuron-like progenitor cells derived from rats that can be successfully grown in culture. The Tufts team, led by Qiaobing Xu, found that exosomes extracted from cultured PC12 cells at various stages of differentiation could drive MSCs to differentiate into neuron-like cells.
Exosomes are very small, hollow particles that a wide range of cells types secrete. These tiny vehicles contain proteins, RNA, and other small molecules, and serve as a vehicle for communication between cells. In the nervous system, exosomes guide the direction of nerve growth, and they control nerve connection and direct peripheral nerve regeneration.
Xu and his team showed that these exosomes contain microRNAs (miRNAs), which a small RNA molecules that regulate gene expression and are known to play a role in neuronal differentiation. They hypothesized that these miRNAs activate neuron-specific genes in the MSCs that receive them and this is the reason these cells begin their journey towards differentiating into neurons.
“In combination with synthetic nanoparticles, we may ultimately be able to use these identified miRNAs or proteins to make synthetic exosomes, thereby avoiding the need to use any kind of neural progenitor cell line to induce neuron growth,” said Xu.
This work was published in PLoS ONE 2015; 10(8): e135111 DOI: 10.1371/journal.pone.0135111.
A German research team at the University of Duisburg-Essen has published a study in the latest issue of STEM CELLS Translational Medicine that shows tiny membrane-enclosed structures that travel between cells work as well as adult stem cells to help the brain recover from a stroke.
Extracellular vesicles (EVs), which are small, membrane-enclosed structures that pass between cells, which are also referred to as exosomes, were given to one group of stroke-impaired mice and adult stem cells from bone marrow to another. After monitoring these mice for four weeks, both groups experienced the same degree of neurological repair. Besides promoting brain recovery in the mice, the EVs also down-regulated the post-stroke immune responses and provided long-term neurological protection.
This study could lead to a new clinical treatment for ischemic strokes, since exosomes carry far fewer risks than adult stem cell transplants, according to the co-leaders of this research, neurologist Thorsten Doeppner, and Bernd Giebel, a transfusion medicine specialist.
“We predict that with stringent proof-of-concept strategies, it might be possible to translate this therapy from rodents to humans, since EVs are better suited to clinical use than stem cell transplants,” said Doeppner and Giebel.
Scientists think that EVs carry biological signals between cells and direct a wide range of processes. Exosomes are under a good deal of scientific investigation for the role they could play in cancer, infectious diseases, and neurological disorders.
Other studies have shown that exosome administration can be beneficial after a stroke, but the Duisburg-Essen study is the first to supply evidence through a side-by-side analysis that they act as a key agent in repairing the brain.
“The fact that intravenous EV delivery alone was enough to protect the post-stroke brain and help it recover highlights the clinical potential of EVs in future stroke treatment,” Doeppner and Giebel said.
This study included contributions from ten different researchers from Duisburg-Essen’s Department of Neurology and Institute for Transfusion Medicine. The study was supported by the university, Volkswagen Foundation and German Research Council.
“The current research, combined with the previous demonstration that EVs are well tolerated in men, suggests the potential for using this treatment in conjunction with clot-busting therapies for treatment of stroke,” said Anthony Atala, M.D., editor of STEM CELLS Translational Medicine and Director of the Wake Forest Institute for Regenerative Medicine.
Scarring of the liver, otherwise known as “liver fibrosis” usually results when the liver is constantly assaulted by inflammation. Conventional treatments for liver fibrosis are usually not very effective. Therefore, mesenchymal stem cells (MSCs) is an attractive alternative due to their ability to suppress inflammation. Unfortunately, transplanted MSCs tend to show poor survival in the scarred liver, and they have an additional tendency to stimulate the formation of new scar tissue. These characteristics have bred skepticism among many investigators.
New work by Asok Mukhopadhyay and his colleagues from New Delhi, India has compared bone marrow (BM)-derived cells with MSCs as a treatment for liver fibrosis. They used CCl4 to induce liver fibrosis in laboratory mice. Then they treated liver-damaged mice with either BM-CD45 cells or fat-based MSCs.
Liver tests and tissue samples of both sets of mice clearly showed that the BM-CD45 cells did a much job attenuating liver scarring than did the fat derived MSCs. Interestingly, the anti-scarring capacity of the BM-CD45 cells was compromised by the presence of MSCs.
Why did the BM-CD45 cells do a better job? The bone marrow cells expressed rather high level expressions of matrix metalloproteinases. These enzymes chopped through scar tissue and also suppressed the hepatic stellate cells, which are responsible for making the scar tissue in liver. Apparently, the BM-CD45 cells induced the die off of the stellate cells. MSCs, however, released two growth factors (TGFβ and IGF-1) that are known to activate hepatic stellate cells, and promote the formation of scar tissue. As an added bonus, transplantation of CD45 cells led to functional improvement of the damaged liver, and this functional improvement seems to the result of improved liver repair and regeneration. Thus transplanted MSCs were pro-scarring while transplanted BM-CD45 cells were pro-regeneration, at least in the liver.
To summarize the results of these experiments, BM-derived CD45 cells appear to be a superior candidate for the treatment of liver fibrosis. The structural and functional improvement of CCl4-damaged livers was substantially better in animals that received transplants of BM-CD45 than those who received fat-derived MSCs.
A new study by researchers from Queen Mary University of London’s Barts Cancer Institute and the Spanish National Cancer Research Centre (CNIO) in Madrid shows that pancreatic cancer stem cells (PancSCs) are very dependent on oxygen-based metabolism, and can be “suffocated” with a drug that is already in use to treat diabetes.
Cancer cells typically rely on glycolysis; a metabolic pathway that degrades glucose without using oxygen. However, it turns out that not all cancer cells are alike when it comes to their metabolism.
PancSCs use another metabolic process called oxidative phosphorylation or OXPHOS to completely oxidize glucose all the way to carbon dioxide and water. OXPHOS uses large quantities of oxygen and occurs in the mitochondria. However, this is precisely the process that is inhibited by the anti-diabetic drug, metformin.
Ever crafty, some PancSCs manage to adapt to such a treatment by varying their metabolism, and this leads to a recurrence of the cancer. However, this English/Spanish team thinks that they have discovered a way to prevent such resistance and compel all PancSCs to keep using OXPHOS. This new discovery might open the door to new treatments that stop cancer stem cells using oxygen and prevent cancers from returning after conventional treatments. A clinical trial is planned for later next year.
Dr Patricia Sancho, first author of the research paper, said: “We might be able to exploit this reliance on oxygen by targeting the stem cells with drugs that are already available, killing the cancer by cutting off its energy supply. In the long-term, this could mean that pancreatic cancer patients have more treatment options available to them, including a reduced risk of recurrence following surgery and other treatments.”
PancSCs become resistant to metformin by suppressing a protein called MYC and increasing the activity of a protein called PGC-1α. However, this resistance mechanism of PancSCs can be abolished if a drug called menadione is given. Menadione increases the amount of reactive oxygen species in mitochondria. Additionally, resistance to metformin can be prevented or even reversed if the MYC protein is inhibited by genetic or pharmacological means. Therefore, the specific metabolic features of pancreatic Cancer Stem Cells are amendable to therapeutic intervention and can provide the basis for developing more effective therapies to combat this lethal cancer.
Pancreatic cancer is still one of the most difficult cancer types to treat. It rarely causes symptoms early on and does not usually trigger diagnosis until its later and more advanced stages. Unfortunately, many patients do not live longer than a year after being diagnosed. These cancers are also becoming more common due to obesity, which increases the patient’s risk for metabolic syndrome and diabetes, which are pancreatic cancer risk factors. Limited treatment options and a failure to improve survival rates mean that finding new treatment strategies is a priority.
PancSCs could be an important but as yet overlooked piece of this puzzle, since they compose only a small proportion of the tumor. PancSCs also have the potential to make new tumors, even if all the other cells are killed, and are prone to spreading around the body (metastasis). Therefore killing these PancSCs is a better way to treat such dangerous cancers.
Last year, a group of Japanese researchers, led by scientists from the high-regarded RIKEN Center for Developmental Biology, reported a break-through in stem cell technology. Their so-called STAP or stimulus-triggered acquisition of pluripotency cells could be derived from mature, adult cells by exposing those cells to stressful conditions. Even though the papers that reported these advances were published in the prestigious journal Nature, immediately, people found problems in the papers that could not be easily resolved. Several laboratories tried to replicate the STAP results, with no success. The papers were eventually retracted and an internal investigation by the RIKEN Center also suggested that foul play might have been at work. Amidst all this, a question that hung in the air was this, “Was there something to the original discoveries but it was overstated?”
That question has now been definitely answered in the negative, thus closing the door for good on this whole sordid affair. Two papers were published on 23 September in the journal Nature, which was the same journal that published the original, ill-fated papers early last year that showed that STAP cells should be called NE (never existed) cells.
The original STAP papers were published in January 2014 by a team led by researchers at the RIKEN Center for Developmental Biology (CDB) in Kobe, Japan, in collaboration with scientists from Harvard Medical School in Boston, Massachusetts. These two papers claimed that embryonic-like stem cells could be produced by exposing adult body cells to stress, such as acidic conditions or physical pressure. These papers dubbed their technology “stimulus-triggered acquisition of pluripotency,” or STAP. Unfortunately, other scientists quickly discovered problems with data in the research. These problems then generated an investigation, and these papers were eventually retracted.
The paper retraction, however, did not answer the nagging questions as to whether or not the STAP procedure might have worked, and where the pluripotent stem cells labelled STAP in the RIKEN laboratory came from.
Such questions were addressed by seven teams in four countries who tried to replicate the procedure under various conditions (De Los Angeles, A. et al. Naturehttp://dx.doi.org/10.1038/nature15513 (2015). These teams collaborated to generate 133 attempts to produce STAP cells, and all of these attempts failed. One of these teams was led by researchers at Harvard Medical School who had worked with one of the original STAP co-authors. In this laboratory, cells were engineered to express a fluorescent protein when a gene related to pluripotency was expressed. When cells were exposed to stressful conditions, they did find some fluorescence, which suggested that pluripotency genes were expressed when cells were subjected to such conditions. However further testing showed this result to be an artifact since cells can naturally emit light; a phenomenon known as autofluorescence. Six other groups also observed autofluorescence in stressed cells, but no convincing evidence of STAP conversion.
A group of RIKEN researchers that did not include any authors of the original STAP papers analyzed the genomes of purported STAP cell lines that had be derived at the CDB. These scientists discovered multiple instances of contradictory data that probably resulted from contamination of purported STAP cells by other known cell types. The RIKEN group’s analyses showed that all remaining purported STAP stem cell lines, for example, were genetically identical to embryonic stem cell lines that already existed in the laboratory.
Additionally the “chimeric” mice that were reportedly produced by injecting STAP cells into the embryo of a developing mouse were found to have been produced by injecting pre-existing embryonic cell lines, rather than STAP cells, into the embryo. The production of chimeric mouse embryos is an experiment that definitively shows that particular cells are truly pluripotent.
Cell contamination also explains one of the most puzzling features of the original work, and that has to do with why the alleged STAP cells were reported to be capable of forming placental tissue, which is something that embryonic stem cells are not able to do (De Los Angeles, A. et al. Nature 525, 469–478 (2015)). These most recent analyses show that mixtures of trophoblast stem cells (which form the placenta in a developing embryo) were mixed with embryonic stem cells and that this mixture was used in the mouse chimeric experiments, leading to the production of mouse placental and embryonic tissue.
Stem-cell scientist Rudolf Jaenisch of the Massachusetts of Technology in Cambridge, who was part of the replication efforts, originally suggested in April 2014 to Nature’s news team that contamination was the reason for the results in the STAP papers. Unfortunately, he did not have evidence at the time for his hypothesis, but this most recent work has vindicated Jaenisch’s hypothesis.
A lingering question is how these embryonic stem cells and trophoblast stem cells came to replace purported STAP cells when the chimeric mouse experiments were performed. So-called cross-examination, which is the accidental contamination of one cell culture by another type of cell, is a well-known problem in cell culture experiments and biological research that depends on cultured cells. However, to properly explain the results in the original STAP papers, multiple independent contamination events must be invoked. “It is very difficult to reconcile the data with simple contamination or careless mislabeling,” says stem-cell scientist George Daley at Harvard Medical School. Unfortunately, requests for clarifying comments from corresponding authors of the original papers went unanswered.
In a review article published in Nature, Daley, Jaenisch argue that all new reports of new types of pluripotency should be subjected to rigorous “forensic” analysis that examines the genomes of the cells under consideration before publication. According to the authors, besides the failed STAP papers, “numerous groups are reporting ever more nuanced states of pluripotency.” In particular, the article focuses on genomic analyses, which are enabled by advances in sequencing technology, that will help evaluate such cell types.
Daley says that these experiments bring some well-desired closure to the STAP. He ended, however, with a warning to scientists who are looking for ways to reprogram cells to an embryonic-like state: “We will all be a tad more cautious in evaluating such claims.”
A Jikei University School of Medicine research team based in Tokyo, Japan, led by Takashi Yokoo, in collaboration with scientists from Meiji University and St. Marianna University School of Medicine in Kawasaki, Keio University School of Medicine in Tokyo, and the School of Veterinary Medicine at Kitasato University in Towada, has shown that mini kidneys grown in vitro from human stem cells can be effectively connected to the excretory systems of rats and pigs.
This is not the first time that research groups have successfully grown mini kidneys in the laboratory. However, connecting these laboratory-grown organs to a laboratory animal’s excretory system constitutes a major technical challenge. The Jikei University team used an approach that employs a step-wise peristaltic ureter or SWPU to connect its lab-grown mini kidneys to the ureter of the transplant animal.
Previous attempts to use laboratory-grown kidneys in laboratory animals have failed because while the transplanted kidneys made urine, they were unable to pass that urine to the animal’s bladder and the kidneys swelled up and failed. Yokoo and his collaborators and colleagues used a stem cell method to make their mini kidneys, as others have in the past. However, he and his team grew more than just the kidney for the host animal; that also grew a drainage tube, known as a ureter, as well, in addition to a bladder to collect and store the urine.
Yokoo and others used laboratory rats as the incubators for their growing tissue. When they connected the new kidney and its tubular systems to the animal’s existing bladder, the system worked. Urine passed from the transplanted kidney into the transplanted bladder and then into the rat bladder. The transplant was still working well when they checked eight weeks later. Then Yokoo and others repeated their procedure in pigs, which are larger mammals than rats and better model systems for human beings. Fortunately, they achieved the same results.
Although this technology is still years away from clinical trials with human patients, this work provides a paradigm for making organs in the laboratory that will work in sick people. In the United Kingdom alone, more than 6,000 people are waiting for a kidney. Because of a shortage of kidney donors, fewer than 3,000 transplants are carried out each year, and more than 350 people die each year waiting for a transplant. Growing new kidneys using human stem cells could solve this problem.
“To our knowledge, this is the first report showing that the SWPU system may resolve two important problems in the generation of kidneys from stem cells: construction of a urine excretion pathway and continued growth of the newly generated kidney,” Yokoo and others wrote in their paper, which was published in the Proceedings of the National Academy of Sciences, USA, which was communicated to the journal by National Academy of Science member R. Michael Roberts from the University of Missouri-Columbia.
Stem cell expert Prof Chris Mason from University College London, said: “This is an interesting step forward. The science looks strong and they have good data in animals. But that’s not to say this will work in humans. We are still years off that. It’s very much mechanistic. It moves us closer to understanding how the plumbing might work. At least with kidneys, we can dialyse patients for a while so there would be time to grow kidneys if that becomes possible.”
A research group from the University of California, San Francisco have isolated and characterized human muscle stem cells. In addition, they have established that these stem cells can robustly replicate and repair damaged muscles when they are grafted onto an injured site. These remarkable findings might open the door to potential treatments for patients with severe muscle injuries, paralysis or genetic diseases that adversely affect skeletal muscles (e.g., muscular dystrophy).
Jason Pomerantz, MD is an assistant professor of plastic and reconstructive surgery at UCSF, and served as the managing author of this work. “We’ve shown definitively that these are bona-fide stem cells that can self-renew, proliferate and respond to injury,” said Pomerantz.
Badly damaged muscles can suffer terrible depletion of their native populations of stem cells or even obliteration of the stem cell niches and populations. Since such muscles have lost the very things that can heal them, these muscles will not be able to heal the damage they have sustained. This very fact represents a terrible hurdle for physicians who specialize in patients who have been crippled by muscle injury and paralysis. One of the worse cases is those conditions that cause damage or paralysis in the critical small muscles of the face, hand and eye, according to Pomerantz.
When muscles are badly damaged, they can lose the native populations of stem cells that are needed to heal. This has posed a major roadblock for treating patients crippled by muscle injury and paralysis, particularly in the critical small muscles of the face, hand and eye, Pomerantz said.
Fortunately, there have been remarkable surgical advances in restoring nerves in damaged muscles. Unfortunately, if the healing process takes too long, the stem cell pool is exhausted and the regenerative capacity is attenuated and eventually. Such injured muscles fail to connect to the nerve tissue and without accompanying motor and sensory nerves, skeletal muscles then to degenerate.
“This is partly why we haven’t had major progress in treating these patients in 30 years,” Pomerantz said. “We know we can get the axons there, but we need the stem cells for there to be recovery.”
A group of stem cells called “satellite cells” line the borders of muscle fibers and, in mice, can function as stem cells and contribute to muscle growth and repair. Until now, however, it wasn’t clear whether human satellite cells worked the same way. It was also terribly unclear how to isolate muscle satellite cells from human tissue samples or even adapt them to help treat patients with muscle damage.
Pomerantz and colleagues tackled this problem used muscle tissue from surgical biopsies of muscles of the head, trunk and leg. Then they used antibody staining to show that human satellite cells can be identified by the expression of the transcription factor PAX7 in combination with the cell-surface proteins CD56 and CD29. Pomerantz and his colleagues use this molecular signature to isolate populations of human satellite cells from these patient biopsies. Then they grafted these satellite cells into mice with damaged muscles whose own muscle stem-cell populations had been depleted. Five weeks after the transplantation, these human cells had successfully integrated into the mouse muscles and divided to produce families of daughter stem cells; effectively replenishing the stem cell niche and repairing the damaged muscle tissue.
This characterization of human muscle stem cells and the ability to transplant them into injured muscles has varied and wide-ranging implications for patients who are presently suffering from muscle paralysis, whose damaged muscles have lost the ability to regenerate. Additionally, protocols that allow us to isolate and manipulate human stem cells also may have applications for understanding why our muscles lose their regenerative capacity during normal aging or in the case of genetic diseases such as muscular dystrophy.
“This gives us hope that we will be able to extract healthy stem cells from other muscles in the patient’s body and transplant them at the site of injury,” Pomerantz said. “If replenishing a healthy muscle stem cell pool facilitates reinnervation and recovery, it would be a significant leap forward.”
These findings appeared the Sept. 8 edition in the open access Cell Press journal, Stem Cell Reports.
Healing the heart after a heart attack is a tough venture. Stem cell treatments have shown definite glimmers to success, but a lack of consistency is a persistent problem. Kick-starting the resident stem cell population in the heart is also a possibility but no single strategy has emerged as a tried and true method to treat a sick heart. Tissue engineering remains an engaging possibility and in the laboratory of Amit Patel at the University of Utah, the possibilities push the boundaries on your imagination.
Patel and his colleagues have been hammering at this problem for decades. The problem is how you replace dead tissue in a beating heart with live tissue that can beat in sync with the rest of the tissue. Unfortunately, you cannot ask the heart to take a vacation to help heal itself. Presently, Patel said that “The doctors say, ‘We’ll give you the beta blocker and the aspirin and the Lipitor and we can just hope to maintain you. But short of them getting worse or getting a heart transplant, there’s [sic] not too many options.”
Patel’s work, however, might change all that. He is presently leading trials on an experimental technology that might repair scarred heart tissue and even arrest or, perhaps, reverse heart failure.
His procedure is in a Phase 1 FDA clinical trial. The trial is designed to mix a powder that consists of a mixture of proteins and molecules isolated from heart muscle with saline or water, inject this mixture into the dead portions of the patient’s heart by means of a catheter, and then wait three to six months to determine if the patient’s heart muscle regenerates.
“Heart disease is the most common cause of death in the world, and the most prominent problem is heart failure,” said Tim Henry, the director of cardiology at the Cedars-Sinai Heart Institute. “Effectively, it’s basically one of the biggest problems in the U.S.” Curing the heart with stem cells is, according to Henry, “within our reach,” and Patel, is, to Henry’s thinking, “is clearly one of the most experienced stem cell people in the country”
After a heart attack, the dead regions of the heart form a scar that does not contract, does not conduct electrical impulses, and the rest of the heart has to work around. Reviving the heart scar, shrinking it or reprogramming it to live again has been the dream of stem cell therapy and gene therapy research. However, according to Patel, these venues have not proven to be very good at regenerating dead scar tissue.
Patel, however, noted that “endocardial matrix therapy” would probably be cheaper than stem cell or gene therapy, since it requires an off-the-shelf product that has the advantage of being mass-produced, is easily delivered clinically speaking, and can be easily commercialized and marketed.
This leads to a new question: “What is “extracellular matrix therapy?”
The extracellular matrix is a foundational material upon which cells sit. Extracellular matrix or ECM also provides the glue that attaches cells to each other, layers of cells to each other, and binds tissues together. In Patel’s rendering, ECM consists of everything in our tissues and organs except the cells. If you were to break down the ECM to its parts, you would end up with a concoction of proteins, minerals and a whole cadre of small molecules that can provide a scaffold for cells, nerves and vessels to attach.
To emphasize the importance of the ECM for the heart, Patel said: “A heart without scaffolding is just a bag of cells.” That pretty well nails it.
The ECM also plays a very important signaling role, since it acts as a repository for important signaling molecules that tell cells to grow and develop or divide and heal. The ECM is the milieu in which cells live and grow.
The foundational importance of the ECM gave Patel a revolutionary thought: to heal the heart the matrix has to come first before the cells can follow.
The powder form of heart-specific ECM was developed by scientists at the University of California, San Diego. This group removed the heart muscle from pig hearts, washed away all the cells, and then freeze-dried the remaining ECM into a powder. Using this work as their template, Patel and his team have also devised a protocol to make ECM power from human heart muscle.
When you add water or saline to this ECM powder, it forms a gooey substance called a “hydrogel.” This hydrogel has been called “VentriGel” and it is as flexible as native tissue. Hydrogels are the mainstay of tissue engineering experiments. VentriGel and hydrogels like it can mimic the molecular environment in which cells normally grow and develop. Fortunately, VentriGel has already been shown to successfully reduce scar tissue in the hearts of rats and pigs. To test VentriGel in human patients, Patel and his co-workers can come to the forefront.
Patel recruited a Utah woman who had suffered a heart attack six months ago. This episode reduced her overall heart blood pumping ability from 60 percent (normal) to less than 45 percent (well below normal). Patel and his colleagues made a virtual model of the inside of the patient’s heart to determine where her dead heart muscle resided. Then they marked out 18 different injection sites, and used a catheter to inject the matrix into her heart. The matrix injection procedure took less than two hours.
“This first patient was able to be done awake and safe and she’s already back to work,” Patel said. “She went home the next day.”
Patel plans to treat up to eighteen patients with his experimental procedure. Additionally, cardiologists at the Minneapolis Heart Institute in Minnesota, the only other site approved to test the new technology, performed the procedure on a second patient on Tuesday.
The risks of this procedure are well-known: When hydrogels are directly injected into the heart muscle, they can unintentionally interrupt the electrical conduction of the heart and cause irregular heartbeats. Also, the injected matrix can travel to other parts of the body where it can form a clot that could lead to a stroke. Clots in other parts of the body can also cause the patient’s blood vessels could collapse.
“If you go through all the bad things that could happen, you’d be so depressed, you’d be like, ‘Really? You found somebody to go through this?'” Patel said. “The key is that the team that we have here, and many of my collaborators, we’re all at that same level of healthy enthusiasm mixed with extreme paranoia.”
All patients will be examined three and six months after the procedure out for evidence of muscle regrowth and revived heart function.
“We want to treat this before it ends up leading to permanent damage,” Patel said.
If the trial returns positive results, it will represent another step forward in a long journey to eradicate heart disease. Patel estimates, that if everything goes smoothly, the technology could become approved for clinical use within five to seven years.
When embryonic stem cell lines are made, they are traditionally grown on a layer of “feeder cells” that secrete growth factors that keep the embryonic stem cells (ESCs) from differentiating and drive them to grow. These feeder cells are usually irradiated mouse fibroblasts that coat the culture dish, but do not divide. Mouse ESCs can be grown without feeder cells if the growth factor LIF is provided in the medium. LIF, however, is not the growth factor required by human ESCs, and therefore, designing culture media for human ESCs to help them grow without feeder cells has proven more difficult.
Having said that, several laboratories have designed media that can be used to derive human embryonic stem cells without feeder cells. Such a procedure is very important if such cells are to be used for therapeutic purposes, since animal cells can harbor difficult to detect viruses and unusual sugars on their cell surfaces that can also be transferred to human ESCs in culture. These unusual sugars can elicit a strong immune response against them, and for this reason, ESCs must be cultivated or derived under cell-free conditions. However, to design good cell-free culture media, we must know more about the growth factors required by ESCs.
To that end, Bruno Reversade from The Institute of Molecular and Cell Biology in Singapore and others have identified a new growth factor that human ESCs secrete themselves. This protein, ELABELA (ELA), was first identified as a signal for heart development. However, Reversade’s laboratory has discovered that ELA is also abundantly secreted by human ESCs and is required for human ESCs to maintain their ability to self-renew.
Reversade and others deleted the ELA gene with the CRISPR/Cas9 system, and they also knocked the expression of this gene down in other cells with small interfering RNAs. Alternatively, they also incubated human ESCs with antibodies against ELA, which neutralized ELA and prevented it from binding to the cell surface. However Ela was inhibited, the results were the same; reduced ESC growth, increased amounts of cell death, and loss of pluripotency.
How does ELA signal to cells to grow? Global signaling studies of growing human ESCs showed that ELA activates the PI3K/AKT/mTORC1 signaling pathway, which has been show in other work to be required for cell survival. By activating this pathway, ELA drives human ESCs through the cell-cycle progression, activates protein synthesis, and inhibits stress-induced apoptosis.
Interestingly, INSULIN and ELA have partially overlapping functions in human ESC culture medium, but only ELA seems to prime human ESCs toward the endoderm lineage. In the heart, ELA binds to the Apelin receptor APLNR. This receptor, however, is not expressed in human ESCs, which suggests that another receptor, whose identity remains unknown at the moment, binds ELA in human ESCs.
Thus ELA seems to act through an alternate cell-surface receptor, is an endogenous secreted growth factor in human
This paper was published in the journal Cell Stem Cell.
Masayo Takahashi, an ophthalmologist at the RIKEN Center for Developmental Biology (CDB) in Kobe, Japan, has pioneered the use of induced pluripotent stem cells (iPSCs) to treat patients with degenerative retinal diseases.
Takahashi isolated skin cells from her patients, and then had them reprogrammed into iPSCs in the laboratory through a combination of genetic engineering and cell culture techniques. These iPSCs have many similarities with embryonic stem cells, including pluripotency, which is the potential to differentiate into any adult cell type.
Once induced pluripotent stem cell lines were established from her patient’s skin cells, they had their genomes sequenced for safety purposes, and then differentiated into retinal pigmented epithelial (RPE) cells. RPE cells lie beneath the neural retina and support the photoreceptors that respond to light. When the RPE cells die off, the photoreceptors also begin to die.
Takahashi watched the transplantation of the RPE cells that she had grown in the laboratory into the back of a woman’s damaged retina. This transplant would constitute the first test of the therapeutic potential of iPSCs in people. Takahashi described the transplant as “like a sacred hour.”
Takahashi has collaborated with Shinya Yamanaka, the discoverer of iPSC technology. She devised ways to convert the iPS cells into sheets of RPE cells. She then tested the resulting cells in mice and monkeys, jumped the various regulatory loops, recruited patients for her clinical trial, and practiced growing cells from those patients. Finally, she was ready to try the transplants in people with a common condition called age-related macular degeneration, in which wayward blood vessels destroy photoreceptors and vision. The transplants are meant to cover the retina, patch up the epithelial layer and support the remaining photoreceptors. Watching the procedure, “I could feel the tension of the surgeon,” Takahashi said.
This transplant surgery occurred approximately a year ago. Some new data on this patient is available.
As of 6 months after the transplant, the procedure appears to be safe. The one-year safety report should appear soon. Prior to the transplant, the patient was a series of 18 anti-vascular endothelial growth factor (anti-VEGF) ocular injections for both eyes to cope with the constant recurrence of the disease. However, data presented by Dr. Takahashi showed that the patient had subretinal fibrotic tissue removed during the transplant surgery in order to make room for the RPE cells. Once the RPE cells were implanted, the patient experienced no recurrence of neovascularization at the 6-month point. This is significant because she has not had any other anti-VEGF injections since the transplant. Her visual acuity was stabilized and there have been no safety related concerns to date.
I must grant that this is only one patient, but so far, these results look, at least hopeful. Hopefully other patients will be treated in this trial, and hopefully, they will experience the same success that the first patient is enjoying. We also hope and pray that the first patient will continue to experience relief from her retinal degeneration.
As to the treatment of the second patient of this trial, Takahashi has hit a snag. Some mutations were detected in the iPS cell-derived RPE cells prepared for the second patient. No one knows if these mutations make these cells dangerous to implant. Regulatory guidelines, at this point, are also no help. Apparently, the cells have three single-nucleotide change and three copy-number changes that are present in the RPE cells that were not detectable in the patient’s original skin fibroblasts. The copy-number changes were, in all cases, single-gene deletions. One of the single-nucleotide changes is listed in a database of cancer somatic mutations, but only linked to a single cancer. Further evaluation of these mutations shows that they were not in “driver genes for tumor formation,” according to Dr. Takahashi.
Tumorigenicity tests in laboratory animals has established that the RPE cells are safe. Remember that the presence of a mutation does not necessarily mean that these RPE cells can be tumorigenic.
However, Takahashi has still decided to not transplant these cells into the second patients. Part of the reason is caution, but the other reason is compliance with new Japanese law on Regenerative Medicine, which became effective after iPS trial was begun. This law, however, does not specify how safe a cell line has to be before it can be transplanted into a patient.
RIKEN’s decision to halt the trial is probably a good idea. After all, this is the first trial with iPSCs and it is important to get it right. Even though the RPE cells were widely thought to be safe to use, Takahashi decided not to implant another patient with RPEs derived from their own cells. Instead, they decided to use RPEs made from donated iPSC lines. Therefore, Takahashi is in discussions government officials to determine how this change of focus for the trial affects their compliance with Japanese law.
Frankly, this might be a very savvy move on Takahashi’s part. As Peter Karagiannis, a spokesperson for the Center for iPS Cell Research and Application, noted: “As of now, autologous would not be a feasible way of providing wide-level clinical therapy. At the experimental level it’s fine, but if it’s going to be mass-produced or industrialized, it has to be allogeneic.”
Therefore, the RIKEN institute is moving forward with allogeneic iPSC-derived RPEs. RIKEN will work in collaboration with the Center for iPS Cell Research and Application (CiRA) in Kyoto, Japan, which has several well characterized, partially-matched lines whose safety profiles have been established by strict, rigorous safety testing methods. However, immunological rejection remains a concern, even if these cells are transplanted into an isolated tissue like the eye where to immune system typically is not allowed. The simple fact is that no one knows if the cells will be rejected until they are used in the trial.
An additional concern is that CiRA has not typed its cells for minor histocompatibility antigens, which can cause T cell–mediated transplant rejection.
Nevertheless, Takahashi and her team deserve a good deal of credit for their work and vigilance.
Mesenchymal stem cells from bone marrow, fat, and other tissues have been used in many clinical trials, experiments, and treatment regimens. While these cells are not magic bullets, they do have the ability to suppress unwanted inflammation, differentiate into bone, cartilage, tendon, smooth muscle, and fat, and can release a variety of healing molecules that help organs from hearts to kidneys heal themselves.
Mesenchymal stem cell transplantation (MSCT) is the main means by which mesenchymal stem cells are delivered to patients for therapeutic purposes. However, the precise mechanisms that underlie the success of these cells are not fully understood. In a paper by from the University Of Pennsylvania School Of Dental Medicine published in the journal Cell Metabolism, MSCT were able to re-establish the bone marrow function in MRL/lpr mice. The MRL/lpr mouse is a genetic model of a generalized autoimmune disease sharing many features and organ pathology with systemic lupus erythematosus (SLE). Such mice show bone loss and poor bone deposition, a condition known as “osteopenia.” Because mesenchymal stem cells are usually the cells in bone marrow that differentiate into osteoblasts (which make bone) a condition like osteopenia results from defective mesenchymal stem cell function.
In this paper, Shi and his coworkers and collaborators showed that the lack of the Fas protein in the mesenchymal stem cells from MRL/lpr mice prevents them from releasing a regulatory molecule called “miR-29b.” This regulatory molecule, mir-29b, is a small RNA molecule known as a microRNA. MicroRNAs regulate the expression of other genes, and the failure to release miR-29b increases the intracellular levels of miR-29b. This build-up in the levels of miR-29b causes the downregulation of an enzyme called “DNA methyltransferase 1” or Dnmt1. This is not surprising, since this is precisely what microRNAs do – they regulate genes. Dnmt1 attaches methyl groups (CH3 molecules) to the promoter or control regions of genes.
Decrease in the levels of Dnmt1 causes hypomethylation of the Notch1 promoter. When promoters are heavily methylated, genes are poorly expressed. When very methyl groups are attached to the promoters, then the gene has a greater chance of being highly expressed. Robust expression of the Notch1 genes activates Notch signaling. Increased Notch signaling leads to impaired bone production, since differentiation into bone-making cells requires mesenchymal stem cells to down-regulate Notch signaling.
When normal mesenchymal stem cells are transplanted into the bone marrow of MRL/lpr mice, they release small vesicles called exosomes that transfer the Fas protein to recipient MRL/lpr bone marrow mesenchymal stem cells. The presence of the Fas protein reduces intracellular levels of miR-29b, and this increases Dnmt1-mediated methylation of the Notch1 promoter. This decreases the expression of Notch1 and improves MRL/lpr BMMSC function.
These findings elucidate the means by which MSCT rescues MRL/lpr BMMSC function. Since MRL/lpr mice are a model system for lupus, it suggests that donor mesenchymal stem cell transplantation into lupus patients provides Fas protein to the defective, native mesenchymal stem cells, thereby regulating the miR-29b/Dnmt1/Notch epigenetic cascade that increases differentiation of mesenchymal stem cells into osteoblasts and bone deposition rates.
Stanford University School of Medicine scientists have enabled the regeneration of damaged heart tissue in animals by delivering a protein to it by means of a bioengineered collagen patch.
“This finding opens the door to a completely revolutionary treatment,” said Pilar Ruiz-Lozano, PhD, associate professor of pediatrics at Stanford. “There is currently no effective treatment to reverse the scarring in the heart after heart attacks.”
Ruiz-Lozano and her colleagues published their data online in the journal Nature.
During a heart attack, cardiac muscle cells or cardiomyocytes die from a lack of blood flow. Replacing dead cells is vital for the organ to fully recover, but, unfortunately, the adult mammalian heart does not possess a great deal of regenerative ability. Therefore, scar tissue forms instead of heart muscle, and since scar tissue does not contract, it compromises the ability of the heart to function properly.
Heart attacks kill millions of people every year, and the number of heart attacks is predicted to rise precipitously in the next few decades. The number of heart attacks might even triple by 2030. Approximately, 735,000 Americans suffer a heart attack each year, and even though many victims survive the initial injury, the resulting loss of cardiomyocytes can lead to heart failure and even death. “Consequently, most survivors face a long and progressive course of heart failure, with poor quality of life and very high medical costs,” Ruiz-Lozano said. Transplanting healthy muscle cells and stem cells into a damaged heart have been tried, but these trials have mixed results, typically, and have yet to produce consistent success in promoting healing of the heart.
Previous heart regeneration studies in zebrafish have shown that the outer layers of the heart, known as the epicardium, is one of the driving tissues for healing a damaged heart. Ruiz-Lozano said, “We wanted to know what in the epicardium stimulates the myocardium, the muscle of the heart, to regenerate.” Since adult mammalian hearts do not regenerate effectively, Ruiz-Lozano and her co-workers wanted to know whether epicardial substances might stimulate regeneration in mammalian hearts and restore function after a heart attack.
She and her colleagues focused on Fstl1, which is a protein secreted by the epicardium, and acts as a growth factor for cardiomyocytes. Not only did this protein kick-start the proliferation of cardiomyocytes in petri dishes, but Ruiz-Lozano and others found that it was missing from damaged epicardial tissue following heart attacks in humans.
Next, Ruiz-Lozano and her colleagues reintroduced Fstl1 back into the damaged epicardial tissue of mice and pigs that had suffered a heart attack. They embedded a bioengineered patch on to the damaged heart tissue that was imbued with Fstl1. Then they sutured the patch, loaded with Fstl1, to the damaged tissue. These patches were made of natural material known as collagen that had been structurally modified to mimic certain mechanical properties of the epicardium.
Because the patches are made of collagen, they contain no cells, which mean that recipients do not need immunosuppressive drugs to avoid rejection. With time, the collagen material is absorbed into the heart. The elasticity of the material resembles that of the fetal heart, and seems to be one of the keys to providing a hospitable environment for muscle regrowth. New blood vessels regenerated there as well.
Within two to four weeks of receiving the patch, heart muscle cells began to proliferate and the animals progressively recovered heart function. “Many were so sick prior to getting the patch that they would have been candidates for heart transplantation,” Ruiz-Lozano said. The hope is that a similar procedure could eventually be used in human heart-attack patients who suffer severe heart damage.
The work integrated the efforts of multiple labs around the world, including labs at the Sanford-Burnham-Prebys Medical Discovery Institute in San Diego, UC-San Diego, Boston University School of Medicine, Imperial College London and Shanghai Institutes for Biological Sciences.
Stanford has a patent on the patch, and Ruiz-Lozano is chief scientific officer at Epikabio Inc., which has an exclusive option to license this technology.
When we think of our Immune systems, we normally entertain visions of white blood cells that fight off invading viruses and bacteria. However, recent work suggests that our immune systems may also being fighting a war against fat.
When laboratory mice are engineered to lack a specific type of immune cell, they become obese and show signs of high blood pressure, high cholesterol, and diabetes. Even though these findings have yet to be replicated in humans, they are already helping scientists understand the triggers of metabolic syndrome, a cluster of conditions associated with obesity.
A new study “definitely moves the field forward,” says immunologist Vishwa Deep Dixit of the Yale School of Medicine, who was not involved in the work. “The data seem really solid.”
Scientists have known for some time now that there is a correlation between inflammation—a heightened immune response—and obesity. Fat cells have the ability to release inflammatory molecules, which complicates these findings, since it is difficult to distinguish if the inflammation causes weight gain or is a side effect of weight gain.
Immunologist Yair Reisner of the Weizmann Institute of Science in Rehovot, Israel, came upon this new cellular link between obesity and the immune system while he was studying autoimmune diseases. Reinser was interested in an immune molecule called perforin, which kills diseased cells by boring a hole in their outer membrane. Reisner’s group suspected that perforin-containing dendritic cells might also be destroying the body’s own cells in some autoimmune diseases. To test their hypothesis, Reisner and his colleagues engineered mice that lacked perforin-wielding dendritic cells. Then they waited to see whether they developed any autoimmune conditions.
“We were looking for conventional autoimmune diseases,” Reisner says. “Quite surprisingly, we found that the mice gained weight and developed metabolic syndrome.”
Mice lacking the dendritic cells with perforin had high levels of cholesterol, early signs of insulin resistance, and molecular markers in their bloodstreams associated with heart disease and high blood pressure. Furthermore, the immune systems of these laboratory animals revealed that they also had a peculiar balance of T cells—a type of white blood cell that directs immune responses.
Reisner and his colleagues report online in the journal Immunity that when they removed these T cells from the mice, the absence of dendritic cells no longer caused the animals to become obese or develop metabolic syndrome.
The results, according to Reisner, suggest that the normal role of the perforin-positive dendritic cells is to keep certain populations of T cells under control. In the same way that perforin acts to kill cells infected with viruses, it can be directed to kill subsets of unnecessary T cells. When the brakes are taken off those T cells, they cause inflammation in fat cells, which leads to altered metabolism and weight gain.
“We are now working in human cells to see if there is something similar going on there,” Reisner says. “I think this is the beginning of a new focus on a new regulatory cell.” If these results turn out to be true in humans, they could point toward a way to use the immune system to treat obesity and metabolic disease.
Daniel Winer, an endocrine pathologist at the University of Toronto in Canada and the lead author of a January Diabetes paper that links perforin to insulin resistance, says the new results overlap with his study. Winer and his group found that mice whose entire immune systems lack perforin developed the early stages of diabetes when fed a high-fat diet. This new paper builds on that by homing in on perforin-positive dendritic cells and showing the link even in the absence of a high-fat diet. “It provides further evidence that the immune system has an important role in the regulation of both obesity and insulin resistance.”
Even if the results hold true in humans, however, a treatment for Type 2 diabetes, obesity or metabolic disease are far off. Dixit said. “Talking about therapeutics at this point would be a bit of a stretch.” Injecting perforin into the body could kill cells beyond those T cells that promoting obesity. We can’t live without any T cells at all, since they are vital to fight diseases and infections.
However, research on what these T cells are recognizing when they seek out fat cells and cause inflammation in fat tissue could eventually reveal drug targets.
If we take tissue samples from the mouth and grow them in the laboratory and manipulate them, we might be able to cure the blind. Blind people who suffer from stem cell deficiency in the cornea might be able to see again by using stem cells isolated from the mouth. Furthermore, this treatment might not only restore vision, but it might also ameliorate pain in the cornea.
Ophthalmologist Tor Paaske Utheim has conducted research for over ten years on how to cure certain types of blindness by using stem cells harvested from tissue obtained from different parts of the body. He then transplants this cultured tissue into the damaged eye, and patients who suffer from blindness as a result of corneal stem cell deficiencies can regain their sight. Recently, Utheim’s research has utilized stem cells from the mouth to grow new corneal tissue, and has also tried to design optimal methods to store and transport this tissue to treat patients.
Utheim is the head of a research group at the Faculty of Dentistry at the University of Oslo (UiO) and the Department of Medical Biochemistry at Oslo University Hospital.
Using cells extracted from the mucous membrane lining the inside of the mouth (the oral mucosa) can restore vision is new to most people. Only ten years ago, this was considered impossible, but results confirm the potential of this method. Twenty clinical studies from various countries have, to date, shown good results, according to Utheim. These clinical trials, however, have only applied these cells to a group of diseases caused by stem cell deficiency in the cornea.
Utheim and his colleagues hope to treat patients with eye injuries caused by so-called limbal stem cell deficiencies. This disorder can be caused by such things as UV radiation, chemical burns, serious infections like trachoma, and various other diseases, some of which are heritable. The number of people worldwide affected by limbal stem cell deficiency is unknown, but in India alone there is an estimated 1.5 million. This disorder most often affects people living in developing countries.
Stem cells that are found at the outer edge of the cornea help to keep the surface of the cornea even and clear. In limbal stem cell deficiencies, the stem cells have been damaged, and they cannot renew the cornea’s outermost layer. Instead, other cells grow over the cornea, which clouds the cornea. The cornea can become fully or partially covered, explains Utheim, which leads to impaired vision or blindness.
Others suffer from severe pain as well. When one patient was interviewed by Norwegian national broadcaster NRK about his limbal stem cell deficiency, he responded: “I don’t know what’s worse: the pain, or losing my sight.”
Utheim explained that when stem cells do not work properly, ulcers can develop in the cornea, which exposed nerve fibers. Since the number of nerve fibers is far higher in the cornea than for example in the skin, it is not surprising that some patients experience severe pain.
A breakthrough within the field occurred about ten years ago when Japanese researchers showed that cells from the oral mucosa could be used to replace limbal stem cells in patients with limbal stem cell deficiency. Although it had been possible since the late 1990s to cure the disorder using cultured stem cells. The available treatment relied on the patient having a healthy eye from which to collect cells.
Further developments made it possible to harvest cells from a relative or deceased individual, but using limbal stem cells from other patients required the use of strong immunosuppressive drugs for the patients, which could cause serious side effects.
A milestone seemed to be reached when it became possible to use a patient’s own cells to treat blindness in both eyes without the need for immunosuppressive drugs. Strangely, this makes some sense because there are similarities between the oral mucosae and the surface of the eye (see Utheim TP. Stem Cells. 2015;33:1685-1695). Originally, using mouth mucosal cells to treat the eye required that the laboratory where the cells are cultured and the clinic where the patients are treated be quite close together. Because there were no protocols for storing extracted oral mucosal cells so that they can be easily kept and transported. This has made the treatment virtually inaccessible to many of the patients who need it the most, namely those in developing countries. However, this may be about to change.
Utheim’s research group is now on the brink of a development that will make it possible to cure both severe pain and blindness in patients who are spread over a larger geographical area than before (see Islam R, et al. PLoS One. 2015;10:e0128306.). “Today, cells from the mouth are cultured for use in the treatment of blindness in only a few specialized centers in the world. By identifying the optimal conditions for storing and transporting the cultured tissue, we would allow for the treatment to be made available worldwide, and not just close to the cell culture centers,” said Rakibul Islam, who is a PhD candidate in the Department of Oral Biology at the Faculty of Dentistry.
Islam is collaborating with Harvard Medical School to introduce this method of treating blindness to clinics around the world. Islam’s findings could also help improve treatment outcomes. “Being able to store the cultured tissue in a small sealed container for a week increases the technique’s flexibility significantly. It makes it easier to plan the operation and allows for quality assurance through microbiological testing of the tissue before transplantation,” Islam explained.
One of the things that Islam and his colleagues have discovered is the specific temperature range at which cells from the mouth should ideally be stored at after culture. Islam has shown that cultured mouth stem cells retain their quintessential properties best between 12 and 16 degrees Celsius (See Dolgin, Elie. Nature Biotechnolgy, 2015;33:224-225.).
During a brief stint at Harvard University, Islam also examined which areas of the mouth are best suited to use in regenerative medicine. In other words, Islam and his colleagues wanted to know which parts of the mouth contain cell layers that regenerate the fastest. Islam explained this using this example: “If you burn any part of your mouth on hot coffee, it heals so quickly that by the next morning you have forgotten about it. This is because the oral mucosa contains cells that multiply quickly. We wanted to investigate whether there were regional differences in the mouth that we could exploit for the treatment of limbal stem cell deficiency.”
Islam continued, “Our results show that the location from which the mucosal tissue is harvested has a striking impact on the quality of the cultured tissue.”
The results from this particular study have not yet been published.
This research can potentially give hope to the many blind that live far away from centralized cell culture laboratories. In work by Utheim in 2010, in collaboration with the ophthalmologist Sten Ræder, he developed storage technology for cultured stem cells that enables the cultured tissue to be transported in a small custom-made plastic container. Tissue from stem cells is thus freed from expensive and bulky laboratory equipment and provides a whole new level of flexibility.
Utheim said “The sample of cells from the mouth can be sent by air over long distances to specialist laboratories with first-class equipment and expertise. After a couple of weeks of laboratory cultivation, the sender may receive the tissue back ready for use. An ophthalmologist could then transplant the stem cells onto the patient’s eye.”
However, the container was just one step in the right direction: “Now we have identified those areas of the mouth that may be best suited for regenerative medicine, and developed a method for storing and transporting tissue from centralized, highly specialized tissue culture centers to clinics worldwide. Our findings are helping to simplify and streamline the clinical procedures, and to make the treatment far more accessible than it is today,” said Islam, who admitted that the transport potential of the project has been integral to his own enthusiasm. He continued, “Although the scientific and technical aspects of our project are very exciting, it has been especially motivating to think of the possibilities this storage technology brings to treating blindness in all parts of the world, including my homeland Bangladesh.”
A central laboratory for the growth of stem cells already exists in Italy. In fact, earlier this year the European Medicines Agency approved the procedure for the cultivation of stem cells from the cornea in EU laboratories. This is the first stem cell therapy to be approved by the European Medicines Agency, according to the journal Nature Biotechnology. Utheim described the approval as an important step towards the implementation of stem cell technology over larger geographical areas. To date, almost 250 people with limbal stem cell deficiency have undergone treatment involving transplantation of stem cells grown from their own mouth cells. “This provides a good basis for judging the success of the treatment” Utheim says.
He has recently published an article in the journal Stem Cells on the inherent potential of cells from the mouth to regenerative medicine. Roughly three out of four treatments are described as successful.
When patients have certain types of leukemia, they can be cured if they receive a bone marrow transplant from a healthy donor. The immune cells from the donated bone marrow will then attack the cancer cells vigorously, and the leukemia will slip into remission.
Such a strategy is called an “allogeneic bone marrow transplant,” and it is an effective way to treat some types of leukemia. However, this technique is risky and it usually involves some patient-related mortality. The problem is getting the transplanted cells to survive.
A new study from the University of Texas MD Anderson Cancer Center in Houston, Texas has examined over eighty patients who have received allogeneic bone marrow grafts for chronic myelomonocytic leukemia (CMML). Stefan O. Ciurea and his colleagues have identified a new pre-treatment that seems to decrease the degree of tumor relapse. Their study was published in the Biology of Blood and Bone Marrow Transplantation.
83 consecutive patients with some form of CMML received an allogeneic bone marrow transplants between April 1991 and December 2013 were examined in detail. They asked if pre-treatment of the bone marrow stem cells with chemicals called “hypomethylating agents” before transplant improved progression-free survival.
Seventy-eight patients received “induction treatment” before transplant, 37 received hypomethylating agents and 41 received cytotoxic chemotherapy. Patients treated with a hypomethylating agent had a significantly lower cumulative incidence of relapse at 3 years post-transplant (22%) than those treated with other agents (35%; p=0.03). However, the transplant-related mortality 1 year post-transplant did not significantly differ between these groups (27% and 30%, respectively; p=0.84). The lower relapse rate resulted in a significantly higher 3-year progression-free survival rate in patients treated with a hypomethylating agent (43%) than in those treated with other agents (27%; p=0.04).
This study supports the use of hypomethylating agents before allogeneic stem cell transplantation for patients with CMML to achieve remission and improve progression-free survival of patients. Of course future studies are needed to confirm these findings, but they suggest that pretreating bone marrow stem cells with hypomethylating agents prior to transplanting them will beef the cells up and help them life longer to fight tumors.
UC Berkeley scientists have discovered a new way to engineer the growth and expansion of energy-burning “good” fat. Subsequently they showed that this fat helped reduce weight gain and lower blood glucose levels in mice. According to Andreas Stahl, the senior author of this study, this technique could potentially lead to new approaches to combat obesity, diabetes and other metabolic disorders.
Stahl and his coworkers devised a specifically tailored hydrogel that acted as a scaffold for stem cells that are able to form brown fat. Implantation of this stem cell-laced scaffold could form a functional brown-fat-like tissue. White fat, which is associated with obesity, stores excess energy, but brown fat is a heat generator and burns calories in order to heat your body.
“What is truly exciting about this system is its potential to provide plentiful supplies of brown fat for therapeutic purposes,” said study lead author Kevin Tharp, a doctoral student in the Department of Nutritional Sciences and Toxicology. “The implant is made from the stem cells that reside in white fat, which could be made from tissue obtained through liposuction.”
“This is figuratively and literally a hot area of research right now,” said Stahl, who is an associate professor of nutritional sciences and toxicology. “We are the first to implant in mice an artificial brown-fat depot and show that it has the expected effects on body temperature and beneficial effects on metabolism.”
Several studies have shown that cold temperatures can increase the metabolic activity of brown fat. However, Stahl pointed out that exposure to cold usually leads to increases in food intake, as well, which potentially negating any calorie-burning benefits from brown-fat activity.
There are three basic types of fat tissue in our bodies. These include energy-storing white fat that many of us are most familiar with, and two kinds of energy-burning fat used to generate heat, which include brown fat, which arises during fetal development, and beige fat, which is brown-like fat that is formed within white fat tissue after exposure to cold and other stressful situations.
For this experiment, Stahl teamed up with Kevin Healy, a UC Berkeley professor of bioengineering. The goal was to increase brown-like beige fat without exposing the animals to cold temperatures. Stahl and Healy wanted to develop a system of physical cues to guide stem cell differentiation.
“It’s already known that for a number of organs, including the heart, the extracellular matrix in which a cell resides provides signals to guide growth and development,” said Dr. Healy. “We applied this concept to stem cells isolated from white-fat tissue.”
The specific matrix recipe for converting white-fat stem cells to brown fat was quite unclear but Stahl and Healy noted that previous studies suggested that stiffness of the surrounding environment was a factor. If white-fat stem cells are placed in a 3D environment that is soft, with little resistance, they become fat. However, if the surrounding environment is rather stiff, the stem cells grew into bone.
Healy and his postdoctoral research fellow Amit Jha generated a tightly knit 3D mesh that consisted of hydrogel, water, hyaluronic acid and short protein sequences associated with brown-fat growth and function. Hyaluronic acid is a naturally occurring acidic carbohydrate that helps make water thicker and gel-like (stiffer in other words). They then took white-fat stem cells (adipose tissue-derived multipotent stem cells or ADMSCs) from mice that had been genetically engineered to express an enzyme from fireflies that made the cells luminescent. Because these cells glowed when incubated with the right substrate, they could be easily traced.
These ADMSCs were then added to the hydrogel and, before the mixture thickened, injected them under the skin of genetically identical mice.
The gel polymerized after injection and stiffens up under the skin of the animal. These labeled cells were then monitored to determine how well they stayed put, how long they persisted in the body and whether they were metabolically functional.
Stahl and Healy and their colleagues noticed an increase in the core body temperature of the mice at ambient temperatures of 21 degrees Celsius and after 24 hours at 4 degrees Celsius. In both cases, the mice with the implanted cells were up to half a degree Celsius warmer than a control group of mice with no injection. The higher the concentration of cells, the larger the effect on temperature.
Next, they put these experimental mice on a high-fat diet. By the end of three weeks, the mice with injected beige fat gained half as much weight and had lower levels of blood glucose and circulating fatty acids compared with control mice.
“This is a feasibility study, but the results were very encouraging,” said Dr. Stahl. “It is the first time an optimized 3D environment has been created to stimulate the growth of brown-like fat. Given the negative health effects of obesity, research into the role of brown fat should continue to see if these findings would be effective in humans.”
North Carolina State University researcher have tested a faster, cheaper way to harvest and grow lung stem cells that have been extracted from patients’ own bodies. That makes such cells a perfect match for lung patients, according to a small proof-of-concept trial.
Ke Cheng, an associate professor of regenerative medicine at NC State, and his team tested this method with, a view toward eventually treating people with idiopathic pulmonary fibrosis, or IPF, a disease that causes inflammation in lung tissue that over time becomes thick and stiff. This scarring of tissue negatively affects lung function over time.
“In current stem cell harvesting, just the process of sorting the stem cells can damage them, wasting not only the cells, but also time and money,” said Cheng. “We wanted to see if we could take healthy stem cells from an organ while they were still in a supportive environment, recreate and enhance that environment outside the body to encourage stem cell reproduction, then reintroduce those cells into a damaged organ to treat disease.”
Cheng and others placed healthy, human adult lung stem cells in a multicellular spheroid, a three-dimensional structure with stem cells in the middle surrounded by layers of support cells. Spheroids are typically used in the laboratory to culture cancer or embryonic cells.
They then used mice with IPF and injected cultured human stem cells into the animals. These injected stem cells produced decreases in inflammation and fibrosis, which Cheng said matched the condition of lungs in the study’s control group that did not have IPF.
Cheng hopes that stem cells isolated from biopsies in human patients can be used to grow and harvest additional cells. Such a procedure should be able to decrease the number of invasive procedures necessary for treatment.
“Picture the lung as a garden and the stem cells as seeds,” Cheng said. “In an IPF environment, with inflammation, the soil is bad, but the seeds are still there. We take the seeds out and give them a protected place to grow. Then when we put them back into the lung, they can grow into mature lung cells to replace the damaged lung tissues in IPF. They can also wake the other seeds up, telling them to help fight the inflammation and ‘improving’ the soil.”
The study was published in the journal STEM CELLS Translational Medicine.
Karolinska Institutet researcher Paolo Macchiarini has been cleared of misconduct charges after what was described as a “a detailed and lengthy investigation,”
The investigation of Dr. Macchiarini was based on his work that was published in seven papers that described the successful transplantation of bioengineered tracheas into patients with tracheal defects. These tracheas were reportedly made from the patient’s own stem cells. The complaints against Macchiarini were lodged by four physicians at Karolinska University Hospital; all of whom were part of the research environment and, in some cases, were even co-authors of the articles in question.
The complainants charged that the findings reported in the first six papers did not agree with the medical records of the patients who were treated. Additionally, there was also a problem with the seventh paper, which describes the implantation of a synthetic esophagus into a rat and a subsequent computed-tomography scan to verify its function. The initial investigation claimed, among other things, that Dr. Macchiarini “selectively described” certain minor problems in the patients while omitting serious complications.
These complaints were submitted in June, August and September 2014, and they prompted an inquiry by Karolinska Institutet’s vice-chancellor in accordance with the provisions of the Higher Education Ordinance and the university’s own established rules for dealing with cases of alleged scientific misconduct.
The recent decision by the university, announced Aug. 28, overturned the decision of an independent investigator commissioned by Karolinska. Bengt Gerdin. Dr. Gerdin is a general surgeon and professor emeritus at Uppsala University in Sweden, and he had concluded in May that Dr. Macchiarini committed misconduct, saying there were problems with those papers that described the transplant procedure in humans. Specifically, there was no evidence for the dramatic improvement claimed in a 2011 Lancet paper concerning such things as blood-vessel formation that penetrated into the grafted trachea.
In response to that investigation, in June the Swedish Research Council announced a freeze on grant payouts to ACTREM (the Centre for Regenerative Medicine) at Karolinska Institutet, which is directed by Dr. Macchiarini.
However, last week’s “no misconduct” ruling was based on more than 1,000 pages of documents submitted by Dr. Macchiarini and his co-authors in response to Dr. Gerdin’s report, in additional to the documents Dr. Gerdin had initially reviewed. In making this latest decision, Vice-Chancellor Anders Hamsten said that while the work in question “does not meet the university’s high quality standards,” it did not constitute misconduct.
“Now that we have examined the allegations of scientific misconduct in all seven indicted articles, we have found that they contain certain flaws but nothing that can be considered scientific misconduct,” the vice-chancellor said.
Proving that someone wilfully deceived the scientific community can be difficult to do. However, this investigation certainly demonstrated that Macchiarini’s work is fraught with problems. Certainly making an artificial trachea is a remarkable advance, but it seems as though Dr. Macchiarini’s synthetic tracheas do not work nearly as well as he claimed they do. This means that further research is required in order to properly make tracheas from a patient’s own stem cells or biomaterials or a combination thereof to treat patients with tracheal defects.
Bone contains a wide variety of stem cells whose potential are only beginning to be tapped. One cell population possesses a cell surface protein called c-kit, and these c-kit+ progenitor cells seem to support myocardial regeneration. Do c-kit+ cells from umbilical cord blood have the same capacity?
Luciana Teofili from the Catholic University of the Sacred Heart in Rome, Italy and her colleagues purified c-kit+ cells from umbilical cord blood by means of magnetic beads that were coated with c-kit-binding antibodies. Teofili and others induced heart muscle differentiation in these cells with several different protocols. Then the expression of cardiac markers (GATA4, GATA6, β-myosin heavy chain, α-sarcomeric actin and cardiac Troponin T) was investigated, and whole-cell current and voltage-clamp recordings were performed.
The c-kit+ cells from umbilical cord blood showed a rather immature gene profile, and by themselves, they did not differentiate into heart muscle-like cells in culture. In contrast, if whole mononuclear cells from umbilical cord blood were subjected to the same treatment, several if the employed protocols produced large, adherent cells that expressed several heart muscle-specific genes and exhibited an excitability much like that of heart muscle cells.
Formation of these heart muscle-like cells was prevented if the c-kit+ cells were removed from the other cells. Tracking studies showed that the c-kit+ cells were the ones that differentiated into heart muscle-like cells, but they only did so when they were together with c-kit– cells.
Thus umbilical cord blood contains progenitors endowed with the ability to differentiate into heart muscle-like cells. The cells with this potential reside in the c-kit+ fraction but they require the presence of abundant accessory cells to differentiate properly.
These preliminary observations suggest that it is a good idea to consider the storage of the umbilical cord blood of patients with prenatal diagnosis of congenital heart diseases. Such conditions might be potentially amenable to myocardial regenerative therapies with umbilical blood-based stem cells.
This paper was published in the journal Cytotherapy, but it must be said that the evidence that these cells differentiated into heart muscle cells was not completely convincing.
Prof Lonnie Shea, from the Department of Biomedical Engineering at the University of Michigan and his team have designed a small sponge-like implant that has the ability to mop up cancer cells as they move through the body. This device has been tested in mice, but there is hope that the device could act as an early warning system in patients, alerting doctors to cancer spread. The sponge-like implant also seemed to stop rogue cancer cells from reaching other areas where they could establish the growth of new tumors. Shea and others published their findings in the journal Nature Communications.
According to Cancer Research UK, nine in 10 cancer deaths are caused by the disease-spreading to other areas of the body. Stopping the spread of cancer cells, or metastasis, is one of the ways to prevent cancers from becoming worse. Complicating this venture is the fact that cancer cells that circulate in the bloodstream are rare and difficult to detect.
Shea’s device is about 5mm or 0.2 inches in diameter and made of a “biomaterial” already approved for use in medical devices. So far, this implant has so far been tested in mice with breast cancer. Implantation experiments showed that it can be placed either in the abdominal fat or under the skin and that it tended to suck up cancer cells that had started to circulate in the body.
The implant mimicked a process known as chemoattraction in which cells that have broken free from a tumor are attracted to other areas in the body by immune cells. Shea and others found that these immune cells are drawn to the implant where they “set up shop.” This is actually a natural reaction to any foreign body, and the presence of the immune cells also attracts the cancer cells to the implant.
Initially, Shea and others labeled cancer cells with fluorescent proteins that caused them to glow under certain lights, which allowed them to be easily spotted. However, they eventually went on to use a special imaging technique that can distinguish between cancerous and normal cells. They discovered that they could definitively detect cancer cells that had been caught within the implant.
Unexpectedly, when they measured cancer cells that had spread in mice with and without the implant, they showed that the implant not only captured circulating cancer cells, but it also reduced the numbers of cancer cells present at other sites in the body.
Shea, said that he and his team were planning the first clinical trials in humans fairly soon: “We need to see if metastatic cells will show up in the implant in humans like they did in the mice, and if it’s a safe procedure and that we can use the same imaging to detect cancer cells.”
Shea and his coworkers are continuing their work in animals to determine what the outcomes if the spread of the cancer spread was detected at a very early stage, which is something that is presently not yet fully understood.
Lucy Holmes, Cancer Research UK’s science information manager, said: “We urgently need new ways to stop cancer in its tracks. So far this implant approach has only been tested in mice, but it’s encouraging to see these results, which could one day play a role in stopping cancer spread in patients.”