Skin-Based Stem Cells Repair Peripheral Nerves

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

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

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

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

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



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

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

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

Treating Crohn’s Disease Fistulas with Fat Stem Cells

All of us have probably heard of Crohn’s disease or have probably known someone with Crohn’s disease. While the severity of this disease varies from patient to patient, some people with Crohn’s disease simply cannot get a break.

Crohn’s disease is one of a group of diseases known as IBDs or “Inflammatory Bowel Diseases.” IBDs include Crohn;s disease, which can affect either the small or large intestine and rarely the esophagus and mouth, ulcerative colitis, which is restricted to the large intestine, and other rarer types of IBDs known that include Collagenous colitis, Lymphocytic colitis, Ischaemic colitis, Diversion colitis, Behçet’s disease, and Indeterminate colitis.

Crohn’s disease (CD) involves the patient’s immune system attacking the tissues of the gastrointestinal tract, which leads to chronic inflammation within the bowel. While the exact mechanism by which this disease works is still not completely understood and robustly debated, Crohn’s disease was originally thought to be an autoimmune disease in which the immune system recognizes some kind of surface protein in the gastrointestinal tract as foreign and then attacks it. However, genetic studies of CD, linked with clinical and immunological studies have shown that this is not the case. Instead, CD seems to be due to a poor innate immunity so that the bowel has an accumulation of intestinal contents that breach the lining of the gastrointestinal tract, resulting in chronic inflammation. A seminal paper by Daniel Marks and others in the Lancet in 2006 provided hard evidence that this is the case. When Marks and others tested the white blood cells from CD patients and their ability to react to foreign invaders, those cells were sluggish and relatively ineffective. Therefore, Crohn’s seems to be an overactivity of the acquired immunity to make up for poor innate immunity.

Given all that, one of the biggest, most painful consequences of CD are anal fistulas. If those sound painful it’s because they are. A fistula is a connection between to linings in your body that should not normally be connected. In CD patients, the anus and the attached rectum get kicked about by excessive inflammation and tears occur. These tears heal, but the healing can cause connections between linings that previously did not exist. Therefore fecal material not comes out of the body in more than one place. Sounds disgusting? It gets worse. Those areas that leak feces are not subject to extensive pus formation and they must be fixed surgically. But how do you fix something that is constantly inflamed? It’s an ongoing problem in medicine.

Enter stem cells to the rescue, maybe. In Spain, a multicenter clinical study has just been published that shows that fat-derived mesenchymal stem cells might provide a better way to treat these fistulas in CD patients. Mesenchymal stem cells have the ability to suppress inflammation, and for that reason, they are excellent candidates to accelerate healing in cases such as these.

Galindo and his group took 24 CD patients who had at least one draining fistula (yes, some have more than one) and gave them 20 million fat-derived mesenchymal stem cells. These cells were extracted from someone else, which is an important fact, since liposuction procedures on these patients might have added to their already surfeit of inflammation.

For this treatment, the cells were administered directly on the lesion, which is almost certainly important. If the closing of the fistula was incomplete after 12 weeks, then the patients were given another dose of 40 million fat-derived mesenchymal stem cells right on the lesion. All these patients were followed until week 24 after the initial stem cell administration.

The results were very hopeful. There were no major adverse effects six months after the stem cell treatment. This is a result seen over and over with mesenchymal stem cells – they are pretty safe when administered properly. Secondly, full analysis the data showed that at week 24 69.2% of the patients showed a reduction in the number of draining fistulas. Even more remarkably, 56.3% of the patients achieved complete closure of the treated fistula. That is just over half. Also, 30% of the cases showed complete closure of all existing fistulas. These results are exciting when you consider the criteria they used for complete closure: absence of draining pus through its former opening. complete “re-epithelization” of the tissue, which means that the lining of the tissue is healed, looks normal and is properly attached to the proper neighbors, and magnetic resonance image (MRI) scans of the region must look normal. For these patients, the MRI “Score of Severity,” which is a measure of the structural abnormality of the anal region, showed statistically significant reductions at week 12 with a marked reduction at week 24. Folks that’s good news.

Galindo interprets his results cautiously and notes that this is a small study, which is true. He also states that the goal of this study was to ascertain the safety of this technique, and when it comes to safety, this technique is certainly safe. When it comes to efficacy, another larger study is required that specifically examined the efficacy of this technique. Galindo is, of course, quite correct, but this is certainly a very exciting result, and hopefully these cells will get further chances to “strut their therapeutic stuff.”

See de la Portilla F, et al Expanded allogeneic adipose-derived stem cells (eASCs) for the treatment of complex perianal fistula in Crohn’s disease: results from a multicenter phase I/IIa clinical trial.  Int J Colorectal Dis. 2013 Mar;28(3):313-23. doi: 10.1007/s00384-012-1581-9. Epub 2012 Sep 29.

Stem Cell Fixes for the Heart

Two recent papers have provided very good evidence that pluripotent stem cells can help heal a heart that has experienced a heart attack. One of these papers used induced pluripotent stem cells from rats, and the other used embryonic stem cells.

The first paper comes from the laboratory of Yoshiki Sawa, who is a professor in the Department of Surgery at the Osaka University Graduate School of Medicine in Osaka, Japan. In this paper, Sawa’s group made induced pluripotent stem cells (iPSCs) from mice and cultured them under conditions known to induce differentiation into heart muscle cells. Beating cells were detected and grown on gelatin-coated plates with Delbecco’s medium. When these cells were tested for gene expression, they made all the same genes as those found in a mouse heart.

To get the cells to form sheets of heart muscle cells, Sawa and his team plated his iPSCs on UpCell plates that are coated with a chemical that causes the cells to adhere to it at normal temperatures, but when the temperature is dropped, the cells detach from the plate. Sawa used another innovation with this culture system; he grew cell without any sugar. This caused all the non-heart cells to die off. The result was a sheet of heart muscle cells that contracted in unison.

Next, the Sawa team took induced heart attacks in a Japanese rat strain. 2 weeks after suffering the heart attack, the sheet of heart muscle cells were placed on the heart scar in half of the rats and the other half received no implants.

Four weeks after implantation of the heart muscle sheet, the differences in heart function were stark. The ejection fraction in the hearts of the animals that had received the iPSC-derived heart muscle sheets increased almost 10%. The fractional shortening, which is the degree to which the heart muscle shortens when it contracts, also increased more than 5%. Also, the amount of stretching during pumping decreased, which indicates that the heart is pumping more efficiently.

When the heart muscle from the implants were examined, they were also filled with molecules associated with the production of new blood vessels. Thus the implanted heart muscle sheets also helped heal the heart by inducing the formation of new blood vessels.

A danger of using iPSC-derived heart muscle cells is the tendency to miss undifferentiated cells and have undifferentiated cells that cause tumors. In this experiment, they noticed tumors if they only grew the cells in the sugar-free medium for a little while. However, if they grew the iPSC-heart muscle cells in sugar-free media for at least three days, all the tumor-causing cells died and implants from these sheets never formed any tumors.

This paper demonstrated the efficacy and plausibility of using patient-specific iPSCs to treat a heart that has had a heart attack some time ago.

The second paper comes from the laboratory of Marisa Jaconi in Geneva, Switzerland. In this paper, Jaconi and her gang of stem cell scientists at the Geneva University Hospitals and the Ecole Polytechnique Fédérale de Lausanne used a “cardiopatch” seeded with cardiac-committed embryonic stem cells to treat a heart attack in rats.

Because the injection of stem cells can induce arrhythmias (irregular heart beats), narrowing of blood vessels, blood vessel obstruction, and other types of damage, these two papers tried to use sheets of cells or cells embedded in biodegradable patches to treat the heart. In this paper, Jacobi and others used a hydrogel made from fibrin, which is the same material found in blood clots. Into that fibrin hydrogel, they placed mouse embryonic stem cells that had been treated with a protein called BMP-2, which drives pluripotent stem cells toward a heart cell fate.

To use these cardiopatches, Jacobi and her group induced heart attacks in a French rat strain and then applied the patch to the heart. They had two groups of rats; those that had been given heart attacks and those that had not. The sham group received either a patch with cells, a patch with iron particles (for detection with MRI) or not patch. The heart attack group received the same.

The results are a little hard to interpret, but the patch + cells definitely improved heart function. First, the hearts that had received patches with cells showed in increase in small blood vessels and blood vessel-making (CD31+) cells. Therefore the patches + cells improved heart circulation. Second, the hearts with the patch + cells showed the presence of new heart muscle cells and much mess thinning of the walls of the heart. Third, the heart functional parameters were better preserved in the patch + cells hearts. The ejection fraction decreased substantially in the hearts that did not receive cells, but in the hearts that received patch + cells, the amount of blood left in the heart after pumping and at rest did not increase nearly as much as in the other groups. These parameters are in indication of the efficiency with which the heart is pumping. The fact that the heart + cells hearts did not decrease in efficiency nearly as precipitously as the others shows that the stem cells are healing the heart.

While these results may not seem terribly robust, we must remember that the cardiopatch was only placed over a small portion of the heart. Therefore, we would not expect to see large increased in function. The fact that we do see new heart muscle cells, new blood vessels, and an arrest in the functional free fall of the heart is significant, given the small area of the heart that was cover with the cells.

The cardiopatch is a new technology and this experiment showed that the patch biodegrades quickly and without incident. It also showed that embedding cells in the patch is feasible, and that the patch is a plausible vehicle to deliver cells to the heart. This procedure also induced the formation of new heart muscle cells in the heart scar and new blood vessels too. Perhaps even more encouraging is the absence of tumors reported in this paper. Even though the ESCs were not differentiated completely into heart muscle cells, the cardiac-directed cells were differentiated enough to form either blood vessels, smooth muscle, or heart muscle. This seems to be enough to prevent the cells from forming tumors. Also, the fibrin scaffold was not deleterious to the heart, even though some studies have used other scaffolds that are damaging to the heart.

Thus cardiopatches and cardiac muscle sheets are perfectly good strategies for treating heart with stem cells. More work needs to be done, but the results are encouraging.