Donor Fat-Based Stem Cells May Provide Augmented Healing of Rectovaginal Fistulas of Crohn’s Disease Patients

Fistulas are openings in organ systems that connect with another system. They usually result from wounds or erosions in the lining of a tube or duct that gets deeper and deeper and eventually opens into another tube or duct. Physical injuries can cause fistulas, but so can diseases such as Crohn’s disease. Anal fistulas result from erosions of the rectum that open to the outside and are typically very painful and do not readily heal.

Damián García-Olmo and his colleagues at the Universidad Autónoma de Madrid have conducted several clinical trials that have examined the ability of adipose-derived stem cells (ASCs) to facilitate the healing of fistulas in Crohn’s disease patients. A phase I study, which primarily examines safety, was published in 2005 (see García-Olmo D., et al., Dis Colon Rectum 2005; 48:1416-1423). According to this study, “No adverse effects were observed in any patient at the end of the follow-up period (minimum follow-up, 12 months; maximum follow-up, 30 months; follow-up average, 22 months).” The Phase II study was published in 2009 (García-Olmo, D., et al., Dis Colon Rectum 2009; 52:79-86). According to the results of this study, fistula healing was observed in 71 percent of patients who were treated with ASCs in combination with fibrin glue compared with 16 percent of patients who received fibrin glue alone. Quality of life scores were also higher in patients who received ASCs than in those who received fibrin glue alone. Once again, the stem cell treatments were well tolerated. The third study was a multicenter, randomized, single-blind clinical trial that enrolled 200 adult patients from 19 centers that were randomly assigned to three groups. The first group received 20 million stem cells (group A, 64 patients). The second group received 20 million adipose-derived stem cells plus fibrin glue (group B, 60 patients). The third group received only fibrin glue (group C, 59 patients). In treatment of anal fistulas in Crohn’s disease patients, a dose of 20 or 60 million adipose-derived stem cells alone or in combination with fibrin glue were demonstrably safe and did promote healing. However, there were no statistically significant differences between the three groups once the 3 groups were compared.

These studies suggest that stem cells from fat might have a place in the treatment of fistulas in Crohn’s disease patients. The application of the stem cells is feasible and safe, and requires no new equipment or skill. The stem cells also might augment the healing of these fistulas.

Unfortunately, anal fistulas are not the only type of fistulas that Crohn’s disease patients can experience. Female Crohn’s disease patients can have fistulas that open from their rectum into their birth canal. These rectovaginal fistula can deposit the contents of the gastrointestinal tract into the lower reproductive tract. While Crohn’s disease is not the only cause rectovaginal fistulas, Crohn’s disease patients are at higher risks for complications, which include: loss of control over stool deposition (fecal incontinence), hygiene problems combined with recurrent vaginal or urinary tract infections, inflammation of the birth canal and skin around the anus (perineum), abscess formation, which can become life-threatening if not treated, and recurrence of the fistula. Surgical treatment of rectovaginal fistulas requires that the tissue be free of inflammation before surgery, which can take time and cause extensive amounts of patient suffering.

Garcia-Olmo and his colleagues have conducted a small phase I-IIa clinical trial to evaluate the possibility of banked fat-based stem cells to treat recto-vaginal fistulas in female Crohn’s patients. This study has several limitations because it is so small and they have to exclude at least half of the participants because of complications beyond their control. Therefore, this study is not statistically significant. However, it does show what might be the beginnings of a stem-based treatment for this horrid condition.

The design of the study included 11 subjects who were initially enrolled in the study, but one of those recruited patients did not meet the criteria for the study. Therefore, ten subjects, all of whom suffered from Crohn’s disease and had rectovaginal fistulas were treated with 20 million fat-based stem cells that had been donated by a healthy volunteer in addition to surgical repair of their fistulas. These donated fat-based stem cells were provided by a Spanish biotechnology company called Cellerix S.L. Three months after this stem cell treatment, two patients were healed and the other either were given an additional treatment of 40 million fat-based stem cells. Of this group, four were healed. However, five of these patients experienced severe flare-ups of their Crohn’s disease that required treatment with biological agents, which disqualified these patients from further consideration from this study. The biological agents used to treat the Crohn’s disease flare-ups are very powerful medicines and can significantly influence the outcome of this study. Thus half of the subjects in this study had to be excluded. Of the five subjects that remained, 3 showed healing of their fistulas, and 2 did not.

The authors present the data as a “final efficacy rate of 60%.” However, given the high rate of exclusion and the very low numbers of subjects in this study, all we can say with any confidence is that based on the previous successes of this treatment in other studies, there is precedent for such a technique to be safe and somewhat effective, and that the data in this study are in a favorable direction. However, that’s about it.

One feature of this study that differs from the other clinical trials done by this same group is that the previous studies utilized the patient’s own fat-based stem cells, whereas this study used stem cells from a healthy donor. The authors stress that this modification greatly simplifies the procedure and decreases its expense. Because of the ease of the treatments, it reduces postoperative hospitalization and is minimally invasive. This new trial suggests that further work is warranted and the results or even minimally hopeful.

This work was published in the journal Stem Cells Translational Medicine 2016; 5(11): 1441-1446.

Lead Induces Oxidative Stress in Neural Stem Cells

Researchers from the Harvard T.H. Chan School of Public Health have elucidated the potential molecular mechanism by which lead, a pervasive environmental toxin, harms neural stem cells and neurodevelopment in children.

The results of this study by Quan Lu and his colleagues suggest that exposure to lead leads to oxidative stress, which perturbs cell behavior. However, Lu and his coworkers found that lead also seems to disrupt the function of certain proteins within neural stem cells.

This study resulted from a collaboration between the Departments of Environmental Health, Biostatistics, and Genetics and Complex Diseases and the T.H. Chan School of Public Health, and the Department of Environmental Health Sciences at Columbia University Mailman School of Public Health, and Department of Preventative Medicine, Mount Sinai School of Medicine.

Epidemiological studies that conclusively linked lead exposure to specific health problems. Lu used these valuable studies are married the epidemiological data with the molecular data from his own work. In fact, this paper by Lu and others, is one of the first to integrate genetic analysis in the lab with genomic data from participants in an epidemiological study.

Lead exposure affects the early stages of neurodevelopment, but the underlying molecular mechanisms by which lead affects early childhood development remain poorly understood.

Lu and others in his laboratory identified one key mechanism that might lead to new therapeutic approaches to treat the neurotoxicity associated with lead exposure.

Numerous studies have suggested that lead exposure can harm the cognitive, language, and psychomotor development of children. Lead exposure also increases the risk that children will later engage in antisocial and delinquent behavior.

Although regulatory limits on the use of lead have definitely reduced blood lead levels in U.S., half a million children aged 1-5 in the U.S. have lead blood levels that are twice those deemed safe by the U.S. Centers for Disease Control. Recent incidents of lead contamination in drinking water in Flint, Mich., and several U.S. cities highlight the continued threat.

Outside the U.S., environmental levels of lead remain high in many countries where lead has not, or has only recently, been phased out from gasoline, paint, and other materials.

Lu and his coworkers explored the molecular mechanisms through which exposure to lead may impact neural stem cells. Neural stem cells can differentiate into other kinds of cells in the central nervous system and play a key role in shaping the developing brain.

In this paper, scientists in Lu’s laboratory and his collaborators conducted a genome-wide screen in neural stem cells for genes whose expression is changed during lead exposure. 19 different genes were identified, and many of these 19 genes are known to be regulated by a protein called NRF2. This is a significant finding, since the NRF2 proteins is known to control the oxidative stress response in cells. This led Lu and others to hypothesize that lead exposure induces an oxidative stress response in cells. However, the Lu group and their collaborators identified a new target of NRF2; a gene designated as SPP1 (also known as osteopontin).

Others involved in this work also conducted genetic analyses on blood samples from a group of infants who were part of the Early Life Exposures in Mexico and NeuroToxicology (ELEMENT) prospective birth cohort. The ELEMENT study was designed to assess the roles of environmental and social factors in birth outcomes and in infant and child development.

Data from the ELEMENT study showed that genetic variants in SPP1 in some blood samples that were statistically linked to abnormal cognition development in those children, whose neurodevelopmental progress was followed through age two. This suggests that lead exerts its deleterious effects, in part, through SPP1. Therefore, drugs that target SPP1 might provide protection against lead exposure in at-risk children.

This paper appeared here: Peter Wagner et al., “In Vitro Effects of Lead on Gene Expression in Neural Stem Cells and Associations between Upregulated Genes and Cognitive Scores in Children,” Environmental Health Perspectives, 2016; DOI: 10.1289/EHP265.

Inhibition of AKT Kinase Increases Umbilical Cord Blood Growth in Culture and Engraftment in Mice

Dr. Yan Liu from the Department of Pediatrics and the Herman B Wells Center for Pediatric Research at the Indiana University School of Medicine in Indianapolis, Indiana and his colleagues have increased the engraftment efficiency of umbilical cord hematopoietic (blood cell-making) stem cells in immunodeficient mice. The technique developed by Lui and his colleagues is simple and increases the proliferation of umbilical cord blood hematopoietic stem cells (UCB-HSCs) in culture, which potentially solves several long-standing problems with umbilical cord blood transplantation.

Umbilical cord blood has been used in the clinic for more than 40 years in hematopoietic stem cell transplantation therapies to treat patients with bone marrow diseases or to reconstitute the bone of those cancer patients who had to have theirs wiped out to cure their leukemia or lymphoma.

One of the problems with umbilical cord blood transplantations, however, is the small amount of material in a typical cord blood collection and, therefore, the small number of hematopoietic stem cells (HSCs) available for transplantation. To ameliorate these shortcomings, hematologists will transplant more than one lot of cord blood (a so-called “double umbilical cord blood transplantation”), which, unfortunately, also increases the risk of immunological rejection (so-called Graft Versus Host response).

A second strategy to get around the low numbers of UCB-HSCs is to expand them in culture, which has proven difficult. However, some experiments have given us more than enough hope to suspect this this is a feasible option (see Flores-Guzmán P, et al., Stem Cells Transl Med. 2013 Nov;2(11):830-8; Bari S., et al., Biol Blood Marrow Transplant. 2015 Jun;21(6):1008-1; Pineault N, Abu-Khader A. Exp Hematol. 2015 Jul;43(7):498-513).

Dr. Lui and his coworkers wanted to examine the role of the signaling protein AKT (also known and protein kinase B) in UCB-HSC expansion in culture. To this end, they used silencing RNAs to knock-down AKT gene expression in cultured UCB-HSCs. AKT knock-down enhanced UCB-HSC quiescence and growth in culture. In a separate experiment, Lui and others treated human UCB-HSCs (so-called CD34+ cells) with a chemical that specifically inhibits AKT activity. Then they subjected these cells to a battery of tests in culture and in laboratory mice.

The results were astounding.  Treatment of human UCB-HSCs did not affect the identity of the HSCs and enhanced their ability to form isolated colonies in cell culture growth tests known as “replating assays.”  Additionally, the short-term inhibition of AKT with drugs also enhanced the ability of UBC-HSCs to repopulate the bone marrow of immunodeficient mice.


In summary, inhibition of AKT increases human UCB-HSC quiescence, growth potential, and engraftment in laboratory mice.

These interesting pre-clinical results suggest that AKT inhibitor can increase the expansion of UCB-HSCs in culture and potential increase their tendency of these cells to engraft in patients.

Induced Pluripotent Stem Cell-Based Model System of Hypertrophic Cardiomyopathy Provides Unique Insights into Disease Pathology

A research team at the Icahn School of Medicine at Mount Sinai led by Bruce Gelb created a model of hypertrophic cardiomyopathy (HCM) by using human induced pluripotent stem cells.

Patients who suffer from an extreme thickening of the walls of the heart exhibit HCM. This excessive heart thickening is associated with a several rare and common illnesses. There is a strong genetic component to the risk for developing HCM. Can stem cell-based model system be used to study the genetics of HCM?

The answer to this question seems to be yes, since laboratory-generated induced pluripotent stem cells lines that have been differentiated into heart cells that, in many cases, closely resemble human heart tissue. Studies with such stem cell-based model systems have reaped useful insights into disease mechanisms (see F Kamdar, et al., J Card Fail. 2015 Sep;21(9):761-70; Lee YK, Ng KM, Tse HF. J Biomed Nanotechnol. 2014 Oct;10(10):2562-85).

In this paper, Bruce Gelb and his colleagues examined a genetic disorder called cardiofaciocutaneous syndrome (CFC). CFC is caused by mutations in a gene called BRAF. It is a rare condition that affects fewer than 300 people worldwide, and causes head, face, skin, and muscular abnormalities, including abnormalities of the heart.

Gelb and his coworkers isolated skin cells from three CFC patients and reprogrammed them into induced pluripotent stem cells, which were then differentiated into heart cells. In this disease model system, the heart muscle cells enlarged, but this seemed to be due to the interaction of the heart muscle cells with heart-specific fibroblasts. Fibroblasts constitute a significant portion of total heart tissue, even though the heart muscle cells are responsible for the actual pumping activity of the heart. In their model system, Gelb and others observed that these fibroblast-like cells produce an excess of a protein growth factor called TGF-beta, which causes the cardiomyocytes to undergo hypertrophy or abnormal enlargement.

This model system has relevance for research on several related and more common genetic disorders, including Noonan syndrome, which is characterized by unusual facial features, short stature, heart defects, and skeletal malformations.

There is no cure for HCM in patients with these related genetic conditions, but if these findings are correct, then scientists might be able to treat HCM by blocking specific cell signals. This is something that scientists already know how to do. Approximately 40 percent of patients with CFC suffer from HCM (two of the three participants in this study had HCM). This suggests a pathogenic connection, though the link has never been adequately researched.

“We believe this is the first time the phenomenon has been observed using a human induced pluripotent stem cell model of the disease,” said Bruce Gelb.

Please see Rebecca Josowitz et al., “Autonomous and Non-Autonomous Defects Underlie Hypertrophic Cardiomyopathy in BRAF-Mutant hiPSC -Derived Cardiomyocytes,” Stem Cell Reports, 2016; DOI: 10.1016/j.stemcr.2016.07.018.

USC Researchers Isolate Human Nephon Progenitor Cells – Future Possibilities for Kidney Regeneration

Researchers at the Saban Research Institute of Children’s Hospital of Los Angeles and the University of Southern California (USC) have reported the isolation of human nephron progenitor (NP) cells. These results, which were published in the journal Stem Cell Translational Medicine, might very well elucidate how progenitor cells differentiate into become renal cells and then develop into kidneys. Such insights could, possibly provide new strategies to promote renal regeneration after chronic kidney failure or acute kidney injury.

Kidneys are composed of about a million tiny filtration units known as “nephrons.” These diminutive structures filter waste and concentrate those wastes into urine, which is leaked into the bladder. In humans, approximately 500,000 to 1,000,000 nephrons are generated before week 34 – 36 of fetal gestation. However, at this point in development, the NP cells are exhausted and kidney development (known as “nephrogenesis”) effectively ceases. If the kidney loses a large enough quantity of nephrons after this time period, such losses may lead to irreversible kidney failure, since no further cell repair or regeneration is possible.


In past studies, NPs were made from induced pluripotent stem cells, or by utilizing animal models. Scientists at USC and Children’s Hospital of Los Angeles (CHLA), chose a different tactic; they designed an efficient protocol by which they could directly isolate human NPs. To accomplish this, Dr. Laura Perin and her colleagues used RNA-labeling probes to obtain cells that expressed the SIX2 and CITED1 genes. Cells expressing both of these genes are almost certainly NPs, since SIX2 and CITED1 are master regulatory genes that promote renal development.

Dr. Perin, co-director of CHLA’s GOFARR Laboratory for Organ Regenerative Research and Cell Therapeutics in Urology, added, “In addition to defining the genetic profile of human NP, this system will facilitate studies of human kidney development, providing a novel tool for renal regeneration and bioengineering purposes.”

On a rather sanguine note, Perin noted that these experiments, which constitute proof-of-concept work, may create new applications to researchers who might be able to use her laboratory’s techniques to isolated progenitor cells for other organs, the pancreas, heart, or lung. “This technique provides a ‘how to’ of human tissue during development,” said Perin.

“It is an important tool that will allow scientists to study cell renewal and differentiation in human cells, perhaps offering clues to how to regulate such development,” added first author of this paper, Stefano Da Sacco.

Factor From Umbilical Cord Blood Could Treat Harmful Inflammation

Umbilical cord blood turns out to have a factor that can potentially fight inflammation, according to scientists at the University of Utah School of Medicine. This study was published online Sept. 6, 2016, in The Journal of Clinical Investigation.

“We found something we weren’t expecting, and it has taken us to new strategies for therapy that didn’t exist before,” says Guy Zimmerman, M.D., a professor of internal medicine at the University of Utah School of Medicine, who was also the senior author of this work. Dr. Zimmerman collaborated with associate professor of pediatrics, Christian Con Yost, M.D., and their colleagues for this work.

Inflammation is well-known to anyone who has whacked their leg, been stung by a bee or a wasp, or anyone who over-stressed their muscles. The redness, heat pain, and swelling are signs that the body is cleaning up damaged cells and their debris, fighting invading microorganisms, and beginning the healing process. However, under certain circumstances, inflammation can go overboard and turn against us and seriously and chronically damage healthy tissues. Out-of-control inflammation is probably the culprit behind several different conditions ranging from rheumatoid arthritis to sepsis. In fact, the inflammatory overreaction to infections is one of the most common causes of hospital deaths.

Dr. Yost and his coworkers successfully isolated a cord blood factor, called “neonatal NET inhibitory factor” or nNIF. This name comes from the ability of this factor to inhibit “NETs” or neutrophil extracellular traps. NETs or neutrophil extracellular traps are composed of processed chromatin bound to granular and selected cytoplasmic proteins that are released by white blood cells called neutrophils. NETs seem to be a kind of last resort that neutrophils turn to in order to control microbial infections. Even though NETs usually help our bodies ward off infectious bacteria and viruses, they can also damage blood vessels and organs during sepsis.


As physicians who have treated critically ill patients suffering from out-of-control inflammation, Drs. Zimmerman and Yost recognized the therapeutic potential of nNIF. “We knew we were onto something that could be very meaningful,” recalls Yost.

To test if this cord blood-based factor could control sepsis, Zimmerman and Yost and others treated groups of mice that suffered from laboratory-induced inflammatory disease. In the absence of treatment, only 20 percent of the mice survived longer than two to four days. However, 60% of those mice treated with nNIF survived after the same amount of time.

“Sepsis is a case where the body’s reaction to infection is lethal,” says Yost. “nNIF is offering insights into how to keep the inflammatory response within prescribed limits.” He adds that they will carry out additional studies to test the therapeutic properties of nNIF.

Anti-Inflammatory Agent Isolated From Umbilical Cord Blood Infection fighting cells from umbilical cord blood (left) and circulating blood three days after birth (right) from the same prematurely born baby. Umbilical cord blood has high levels of a factor, called neonatal NET inhibitory factor (nNIF), which inhibits a specific inflammatory response called NETs. Within two weeks after birth, nNIF levels drop and NETs can form. True to their name, they consist of a net-like substance that traps infectious agents like bacteria, as seen on the right. nNIF is showing promise as a potential therapy against harmful inflammation and sepsis.
Anti-Inflammatory Agent Isolated From Umbilical Cord Blood
Infection fighting cells from umbilical cord blood (left) and circulating blood three days after birth (right) from the same prematurely born baby. Umbilical cord blood has high levels of a factor, called neonatal NET inhibitory factor (nNIF), which inhibits a specific inflammatory response called NETs. Within two weeks after birth, nNIF levels drop and NETs can form. True to their name, they consist of a net-like substance that traps infectious agents like bacteria, as seen on the right. nNIF is showing promise as a potential therapy against harmful inflammation and sepsis.

nNIF seems to be present for just a brief window of time at the beginning of life. It circulates in cord blood and persists in the baby’s own bloodstream for up to two weeks after birth. However, after two weeks, nNIF disappears and is not found in older babies and is completely absent from the blood of adults. Scientists in Yost’s laboratory also discovered that the placenta also contains a similar, albeit less potent, anti-inflammatory agent. The evanescent nature of these factors possibly indicates that inflammation is under tight control during this time, since the fragility of young babies might make extensive amounts of inflammation deleterious to their health.

“The beginning of life is a delicate balance,” says Yost. “Our work is showing that it is important to have the right defenses, but they have to be controlled.”

AUF1 Gene Important Inducer of Muscle Repair

A new study in the laboratory of Robert J. Schneider at NYU Langone and his collaborators has uncovered a gene that plays integral roles in the repair of injured muscle throughout life. This investigation shows that this previously “overlooked” gene might play a pivotal role in “sarcopenia,” which refers to the loss of muscle tissues with age.

This collaboration between scientists at NYU Langone Medical Center and the University of Colorado at Boulder showed that the levels of a protein called AUF1 determine if stem cell populations retain the ability to regenerate muscle after injury and as mice age.

Changes in the activity of AUF1 have also been linked by past studies to human muscle diseases. More than 30 genetic diseases, known collectively as myopathies, show defective muscle regeneration and these anomalies cause muscles to weaken or waste away.

For example, muscular dystrophy is a disease in which abnormal muscles fail to function properly and undergo normal repair. Although the signs and symptoms of Duchenne Muscular Dystrophy vary, in some cases wildly, this disease develops in infants and affects and weakens the torso and limb muscles beginning in young adulthood. Sarcopenia, in healthy individuals occurs in older patients.

Skeletal muscles have a stem cell population set aside for muscle repair known as satellite cells. These cells divide and differentiate into skeletal muscle when skeletal muscle is damaged, and as we age, the capacity of muscle satellite cells to repair muscle decreases.

AUF1 is a protein that regulates muscle stem cell function by inducing the degradation of specific, targeted messenger RNAs (mRNAs). According to Robert Schneider, “This work places the origin of certain muscle diseases squarely within muscle stem cells, and shows that AUF1 is a vital controller of adult muscle stem cell fate.” He continued: “The stem cell supply is remarkably depleted when the AUF1 signal is defective, leaving muscles to deteriorate a little more each time repair fails after injury.”

The experiments in this study demonstrated that mice that lack AUF1 display accelerated skeletal muscle wasting as they age. These AUF1-depleted mice also showed impaired skeletal muscle repair following injury. When the molecular characteristics of these AUF1-depleted muscle satellite cells were examined, Schneider and his collaborators showed that auf1−/− satellite cells had increased stability and overexpression of so-called “ARE-mRNAs.” ARE mRNAs contain AU-rich elements at their tail-ends. AUF1 proteins bind to these ARE mRNAs and induce their degradation. In the absence of AUF1, muscle satellite cells accumulate ARE mRNAs. One of these ARE mRNAs includes that which encodes matrix metalloprotease, MMP9. Overexpression of MMP9 by aging muscle satellite cells causes degradation of the skeletal muscle matrix, which prevents satellite-cell-mediated regeneration of muscles. Consequently, the muscle satellite cells return to their quiescent state and fail to divide and repair skeletal muscle.

When Schneider and his coworkers and collaborators blocked MMP9 activity in auf1−/− mice, they found that they had restored skeletal muscle repair and maintenance of the satellite cell population.

These experiments suggest that repurposing drugs originally developed for cancer treatment that blocks MMP9 activity might be a way to dial down age-related sarcopenia.

“This provides a potential path to clinical treatments that accelerate muscle regeneration following traumatic injury, or in patients with certain types of adult onset muscular dystrophy,” said Schneider.

This work was published here: Devon M. Chenette et al., “Targeted mRNA Decay by RNA Binding Protein AUF1 Regulates Adult Muscle Stem Cell Fate, Promoting Skeletal Muscle Integrity,” Cell Reports, 2016; DOI: 10.1016/j.celrep.2016.06.095.