A Way to Get Stem Cells to Make Living Heart Valve Tissue?

What a benefit it would be to be able to replace diseased and defective heart valves with new heart valves. Thus, living tissue engineered heart valves (TEHV) would be a boon to children who require replacement heart valves that have the capacity to grow with the child and completely integrate into the child’s heart tissue. A persistent challenge for TEHV is accessible human cell source(s) that have the ability to mimic native valve cell phenotypes and possess matrix remodeling characteristics that are essential for long-term function.

Mesenchymal stem cells derived from bone marrow (BMMSC) or adipose tissue (ADMSC) are intriguing cell sources for TEHV. Unfortunately, they have not been compared to pediatric human aortic valve interstitial cells (pHAVIC) in relevant 3-dimensional culture environments.

In a recent study, Bin Duan from the Biomedical Engineering department at Cornell University compared the spontaneous and induced multipotency of ADMSC and BMMSC to that of pHAVIC using different induction culture systems within three-dimensional (3D) bioactive hybrid hydrogels that have similar material properties to those of aortic heart valve leaflets. pHAVICs possessed some multi-lineage differentiation capacity in response to induction media, but these cells were limited to the earliest stages and their differentiation capacity were less potent than either ADMSCs or BMMSCs. ADMSCs expressed cell phenotype markers that were similar to pHAVICs when they were grown in HAVIC growth media spiked with a growth factor called basic fibroblast growth factor (bFGF). BMMSCs generally expressed extra cellular matrix remodeling characteristics similar to pHAVICs.

Duan and his colleagues then chemically attached bFGF to components of the 3D hybrid hydrogels in order to further immobilize them. The immobilized bFGF upregulated vimentin expression and promoted the fibroblastic differentiation of pHAVIC, ADMSC and BMMSC. Since fibroblasts help make heart valves, these changes in gene expression might presage the ability of these cells to form new heart living heart valve tissue.

Thus, these findings show that even though mesenchymal stem cells retain a heightened capacity to form bone in 3D culture, this tendency can be shifted fibroblast cell fates by tethering bFGF to the 3-D matrix. Such a strategy is probably rather important for utilizing stem cell sources in heart valve tissue engineering applications.

This is an important finding.  Even though the production of TEHVs are some ways off, Duan’s findings might provide a strategy to begin cells on the path to making TEHVs.

Do Human Mesenchymal Stem Cell Therapies Help Older Patients with Ischemic Cardiomyopathy?

Joshua Hare from the Interdisciplinary Stem Cell Institute at the University of Miami Miller School of Medicine in Miami, Florida has conducted a variety of high-quality clinical trials that have tested the ability of mesenchymal stem cells to heal the hearts of patients with ischemic heart disease. Two of these trials, (Transendocardial Autologous Cells in Ischemic Heart Failure) and POSEIDON (Percutaneous Stem Cell Injection Delivery Effects on Neomyogenesis), injected mesenchymal stem cells from bone marrow directly into damaged heart muscle.

Both of these studies not only showed an increase in heart function after injection of mesenchymal stem cells compared to placebo, but further examination showed that mesenchymal stem cells induced shrinkage of the heart scar and replacement with living heart muscle tissue (see Alan Heldman and others, Transendocardial Mesenchymal Stem Cells and Mononuclear Bone Marrow Cells for Ischemic Cardiomyopathy: The TAC-HFT Randomized Trial, JAMA, Published online November 18th 2013). However, Hare wanted to compare the benefits experienced by younger patients with older patients in order to determine if age had any effect on the efficacy of this treatment.

To that end, Hare and his colleagues compared subjects from the TAC-HFT and POSEIDON clinical trials in 2 age groups: younger than 60 and 60 years of age and older. They used a 6-min walk distance to measure heart function and the Minnesota Living With Heart Failure Questionnaire (MLHFQ) to ascertain the quality of life of each patient.  Patients were tested at baseline (before the procedure), 6 months, and 1 year after the procedure.  Hare and his group also used particular cardiac imaging measurements, such as absolute scar size and compared the baseline size of the heart scar, and then again 1 year after the procedure.

These two tests, the 6MWD and the MLHFQ showed improvements in both age groups. These improvements were even significant in both groups. What this analyses show is that mesenchymal stem cell therapy helps patients with ischemic heart failure, regardless of their age. Older individuals did not have an impaired response to MSC therapy.

This is an important result because heart disease is very often a condition of the aged and there are concerns as to whether or not older patients would benefit from regenerative medical procedures. Hare’s study suggests that older patients do benefit from these procedures. A caveat is that older patients have lower-quality mesenchymal stem cells, but these studies tended to use allogeneic mesenchymal stem cells or stem cells from donors. Therefore, allogeneic stem cell treatments may prove effective in older heart patients.

Gestational Diabetes Affects the Quality of Umbilical Cord Mesenchymal Stromal Cells

The laboratories of Drs. Jene Choi and Chong Jai Kim from the University of Ulsan College of Medicine in Seoul, South Korea have collaboratively shown that the therapeutic quality of umbilical cord mesenchymal stem cells is profoundly affected by gestational diabetes. Their work was published in a recent issue of the journal Stem Cells and Development and has profound implications for regenerative medicine.

Choi and Kim and their coworkers collected umbilical cords from mothers who had been given birth by Cesarian section and had also been diagnosed with gestational diabetes and mothers who had also just given birth by Cesarian section and showed normal blood sugar control. These umbilical cord tissues were processed and the mesenchymal stem cells from the cord tissue were isolated and cultured. These cells were grown and then subjected to a rather extensive battery of tests. These tests were a reflection of the ability of these to perform in regenerative treatments.

First umbilical cord mesenchymal stem cells (UCMSCs) from mothers with gestational diabetes (GD) did not grow as well as UCMSCs from mother who did not have GD.  As you can see in the graphs below, these are not small growth differences.  The UCMSCs from non-GD mothers (on the left) grow substantially better than those from GD mothers.  This result is also consistent for different cell lines.  This also means that transplanted cells would not grow very well if they were used for therapeutic purposes.

Umbilical cord mesenchymal stromal cells (UC-MSCs) derived from gestational diabetes mellitus (GDM) patients exhibit retarded growth proliferation. The growth of 7.5×103 UC-MSCs isolated from patients with normal pregnancies (A) and GDM (B) was monitored over a period of 12 days. GDM-UC-MSCs consistently showed decreased proliferation compared with normal pregnant women (N-UC-MSCs). Points represent the mean values from three independent experiments; bars denote standard deviation (SD).
Umbilical cord mesenchymal stromal cells (UC-MSCs) derived from gestational diabetes mellitus (GDM) patients exhibit retarded growth proliferation. The growth of 7.5×103 UC-MSCs isolated from patients with normal pregnancies (A) and GDM (B) was monitored over a period of 12 days. GDM-UC-MSCs consistently showed decreased proliferation compared with normal pregnant women (N-UC-MSCs). Points represent the mean values from three independent experiments; bars denote standard deviation (SD).

Secondly, UCMSCs from GD mothers showed a greater tendency to undergo premature senescence.  When MSCs are grown in culture, they usually grow rather well for several days and then the cells go to sleep and they stop growing.  This is called culture senescence and it is due to intrinsic properties of the cells.  When the cells go into senescence tends to be a cell line-specific property, but one thing is certain; the sooner cells become senescent, the few cells they will generate in culture.  The UCMSCs from GD mothers go into senescence early and easily and this is one of the reasons they grow so poorly relative to normal cells – because they are running to their beds to take a nap (so to speak).  Such cells are usually not good candidates for regenerative medicine.

Third, UCMSCs from GD mothers show poor lineage-specific differentiation.  MSCs have the ability to differentiate into fat cells, bone cells, and cartilage cells if particular well-established protocols are used.  However, UCMSCs from GD mothers showed inefficient differentiation and that is one of the things that MSCs must do if they are to repair bone or cartilage problems or if they are to help make smooth muscle for new blood vessels formation. 

Stem cell differentiation potentials are largely different between normal and GDM-affected UC-MSCs. Three different cell lines of normal and GDM-affected pregnancies were cultured in a control medium or induction medium for 5 days. Upregulation of the expression of the adipogenic-specific gene PPARγ (A) and the osteogenic genes alkaline phosphatase (ALP) (B), osteocalcin (OC) (C), and collagen type 1 alpha 1 (Col1α1) (D) was evaluated by real-time RT-PCR and normalized to GAPDH. All assays were performed in triplicate; bars denote SD (*P<0.05).
Stem cell differentiation potentials are largely different between normal and GDM-affected UC-MSCs. Three different cell lines of normal and GDM-affected pregnancies were cultured in a control medium or induction medium for 5 days. Upregulation of the expression of the adipogenic-specific gene PPARγ (A) and the osteogenic genes alkaline phosphatase (ALP) (B), osteocalcin (OC) (C), and collagen type 1 alpha 1 (Col1α1) (D) was evaluated by real-time RT-PCR and normalized to GAPDH. All assays were performed in triplicate; bars denote SD (*P<0.05).

The figure above shows the disparity between these established UCMSC cell lines.  The dark, solid bars indicate non-induced cells that were grown in normal culture media, and the striped bars are cells grown in media that designed to induce the differentiation of these cells into either bone, fat, or cartilage cells.  The cell lines with “N” in their name are from non-GD mothers and those with “D” in their designations are from GD mothers.  These assays are for genes known to be strongly induced when cells begin to differentiate into fat (PPARgamma), bone (ALP or osteocalcin or collagen 1 alpha 1).  As you can clearly see, the Ns outdo the Ds every time.

Finally, when the mitochondria, the compartments in cells that generate energy, from these two cell populations were examined it was exceedingly clear that UCMSCs from GD mothers had mitochondria that were abnormal and did not make every very well.  Mitochondria from UCMSCs taken from GD mothers showed decreased expression of the energy-making components.  Thus the energy-making pathways in these cell compartments were sub-par from a structural perspective.  Functional assays for mitochondria showed that mitochondria from UCMSCs from GD mothers consistently underperformed those from UCMSCs taken from non-GD mothers.  Also, when markers of mitochondrial dysfunction were measured (reactive oxygen species and indicators of mitochondrial damage from reactive oxygen species), such markers were consistently higher in mitochondria from UCMSCs from GD mothers relative to those from non-GD mothers.  This shows that the energy-making or powerhouses of the cells are dysfunctional in UCMSCs from GD mothers.  Without the ability to properly make energy from food molecules, the cells have a diminished capacity to heal damaged tissues and organs.

Several studies have established a positive link between mitochondrial dysfunction and accelerated aging.  Therefore, these cells, because they have more extensive indications of mitochondrial damage, may show profound accumulation of mitochondrial damage and accelerated aging.

In summary, this study shows that integral biological properties of human UC-MSCs differ according to obstetrical conditions.  These data also stress the importance of maternal–fetal conditions in biological studies of hUC-MSCs and the development of future therapeutic strategies using hUC-MSCs.

Stem Cells from Adult Nose Tissue Used to Cure Parkinson’s Disease in Rats

For the first time, German stem cell scientists from the University of Bielefeld and Dresden University of Technology have used adult human stem cells to “cure” rats with Parkinson’s disease.

Parkinson’s disease results from the death of dopamine-using neurons in the midbrain, and the death of these midbrain-based, dopamine-using neurons causes a loss of control of voluntary motion. Presently, no cure exists for Parkinson disease.

In this study, which was published in STEM CELLS Translational Medicine, the German team produced mature dopamine-using neurons from inferior turbinate stem cells (ITSCs). ITSCs are stem cells taken from tissues that are normally discarded after an adult patient undergoes sinus surgery. The German team tested how ITSCs would behave when transplanted into a group of rats with a chemically-induced form of Parkinson’s disease. Prior to transplantation, the animals showed severe motor and behavioral abnormalities. However, 12 weeks after transplantation of the ITSCs, the cells had not only migrated into the animals’ brains, but their functional ability was fully restored and significant behavioral recovery was also observed. Additionally, none of the treated animals shows any signs of tumors after the transplantations, something that also has been a concern in stem cell therapy.

“Due to their easy accessibility and the resulting possibility of an autologous transplantation approach, ITSCs represent a promising cell source for regenerative medicine,” said UB’s Barbara Kaltschmidt, Ph.D., who led the study along with Alexander Storch, M.D., and Christiana Ossig, M.D., both of Dresden University. “The lack of ethical concerns associated with human embryonic stem cells is a plus, too.”

“In contrast to fighting the symptoms of Parkinson’s disease with medications and devices, this research is focused on restoring the dopamine-producing brain cells that are lost during the disease,” said Anthony Atala, M.D., Editor-in-Chief of STEM CELLS Translational Medicine and director of the Wake Forest Institute for Regenerative Medicine. These cells are easy to access and isolate from nasal tissue, even in older patients, which adds to their attraction as a potential therapeutic tool.”

This is certainly a very exciting animal study, but treating chemically-induced Parkinson’s disease in rodents and treating Parkinson’s disease in aged human patients is two very different things. Thus while this study is important, work in human wild require more testing and studies in larger animals.

Novastem Treats Its First Stroke Patient With Stemedica’s Mesenchymal and Neural Stem Cell Combination

The biotech company Novastem is a leader in regenerative medicine and has announced the treatment of its first patient in its clinical study for ischemic stroke at Clinica Santa Clarita, Mexico. This clinical trial is testing cell products made by Stemedica. In particular, Stemedica’s ischemia-tolerant mesenchymal stem cells (itMSCs) were administered in combination with ischemia-tolerant neural stem cells (itNSCs); both of which are proprietary products of Stemedica.

Stemedica‘s itMSCs and itNSCs are unique because of the manner in which they are manufactured – they are grown under conditions that make them resistant to low-oxygen conditions. Experiments conducted with these cells in culture and in living animals have definitely shown that when these cells are exposed to low-oxygen conditions, they show greater homing and engraftment than cells grown under normal conditions. Compared to other MSCs and NSCs, Stemedica’s stem cells secrete higher levels of growth factors and other important proteins associated with angiogenesis and healing.

According to the American Stroke Association, ischemic strokes account for 87 percent of all stroke cases. Novastem is continuing to enroll qualified patients in their study. This clinical trial is entitled “Internal Research Protocol in Combination Therapy of Intravenous Administration of Allogeneic Mesenchymal Stem Cells and Intrathecal Administration of Neural Stem Cells in Patients with Motor Aphasia due to Ischemic Stroke.” All participants in this clinical trial will receive a unique, combination stem cell therapy consisting of cells made by Stemedica Cell Technologies.

Novastem is sponsoring this clinical trial and Novastem is the only company licensed to use Stemedica’s stem cell products for studies in Mexico. Novastem’s Clinica Santa Clarita facility is federally licensed to use stem cell therapies, and this trial marks the first time ischemic stroke is being treated with a patented medical method that comprises administration of hypoxically-grown neural stem cells into the cerebrospinal fluid in combination with intravenous administration of hypoxically-grown mesenchymal stem cells. This combination approach is designed to treat the after effects of ischemic strokes.

“Novastem and Clinica Santa Clarita are committed to advancing the research of neurodegenerative disease, and we are pleased to be working with internationally-recognized physician Clemente Humberto Zuniga Gil, MD as the principal investigator and study designer,” says Rafael Carrillo, Novastem’s President. “Our medical team believes that Stemedica’s mesenchymal and neural stem cells, used in this unique combination therapy, will restore and build new vascularization, improve the blood supply, reconnect damaged neural networks and improve functionality of areas affected by our patients’ ischemic stroke.”

The aim of this Novastem study is to evaluate functional changes on subjects after the administration of ischemia-tolerant mesenchymal and neural stem cells. The protocol in use in this clinical trial has been approved by the Research Ethics Committee of Clinica Santa Clarita, which is federally registered and licensed by the Federal Commission for the Protection against Sanitary Risk (COFEPRIS), a division of Mexico’s Ministry of Health.

Patient progress will be tracked at the beginning of the study before any cells have been administered, at 90 days after stem cell administration, and then again at 180 days after administration. Patient improvement will be ascertained with the United States National Institute of Health Stroke Score (NIHSS), Stroke and Aphasia Quality of Life Scale-39 (SAQCOL-39) and the Boston Diagnostic Aphasia Examination (BDAE) neuropsychological evaluation for diagnosis. Additionally, MRIs taken with a gadolinium-based contrast agent (GBCA) will examine the structural integrity of the brain before and after stem cell administration. At the endpoint, the treatment will be evaluated for safety and tolerance of the two-cell treatment. Additionally, patients will be evaluated for changes in neurological functionality.

New Bone Marrow-Based Stem Cell Identified in Mice that Regenerates Bones and Cartilage in Adults

Researchers at Columbia University Medical Center (CUMC) have discovered a bone marrow-based stem cell capable of regenerating both bone and cartilage in mice. The discovery appeared in the online issue of the journal Cell.

These cells have been called osteochondroreticular (OCR) stem cells, and they were identified in experiments that tracked a protein expressed by these cells. By using this specific protein as a marker for OCR stem cells, the Columbia team found that OCR cells self-renew and produce key bone and cartilage cells, including osteoblasts and chondrocytes. Furthermore, when OCR stem cells are transplanted to a fracture site, they dutifully contribute to bone repair.

“We are now trying to figure out whether we can persuade these cells to specifically regenerate after injury. If you make a fracture in the mouse, these cells will come alive again, generate both bone and cartilage in the mouse—and repair the fracture. The question is, could this happen in humans,” says Siddhartha Mukherjee, MD, PhD, assistant professor of medicine at CUMC and a senior author of the study.

Since mice and humans have similar bone biology, Mukherjee and his colleagues are quite confident that OCR stem cells exist in human bone marrow. Further studies could uncover new and effective ways to exploit OCR cells to provide greater ways to prevent and treat osteoporosis, osteoarthritis, or bone fractures.

“Our findings raise the possibility that drugs or other therapies can be developed to stimulate the production of OCR stem cells and improve the body’s ability to repair bone injury—a process that declines significantly in old age,” says Timothy C. Wang, MD, the Dorothy L. and Daniel H. Silberberg Professor of Medicine at CUMC, who initiated this research. Wang and his team previously found an analogous stem cell in the intestinal tract and observed that it was also abundant in the bone.

“These cells are particularly active during development, but they also increase in number in adulthood after bone injury,” says Gerard Karsenty, MD, PhD, the Paul A. Marks Professor of Genetics and Development, chair of the Department of Genetics & Development, and a member of the research team.

Mukherjee and his coworkers also showed that adult OCR stem cells are distinct from mesenchymal stem cells (MSCs). MSCs play essential roles in bone generation during development and adulthood. Therefore, researchers thought that MSCs gave rise to all bone, cartilage, and fat, but recent studies have shown that MSCs do not generate young bone and cartilage. This study by Mukherjee and his colleagues suggests that OCR stem cells actually make young bone and cartilage, but both OCR stems cells and MSCs contribute to bone maintenance and repair in adults.

Mukherjee also suspects that OCR cells may play a role in soft tissue cancers.

A research team from Stanford University School of Medicine just released a similar study that used a different methodology to identify the same stem cell type.

Brigham and Women’s Hospital Researchers Reverse Type 1 Diabetes in Diabetic Mice

Brigham and Women’s Hospital (BWH) is a Harvard University-affiliated institution with a robust research program. In particular, several BWH are interested in mesenchymal stem cells and their ability to suppress inflammation and mediate healing in injured organs.

To that end, a research team led by Robert Sackstein from BWH’s Departments of Dermatology and of Medicine and Reza Abdi from BWH’s Department of Medicine and Transplantation Research Center, has published a stupendous report in the journal Stem Cells. In this paper, Sackstein and his coworkers used mesenchymal stem cells (MSCs) to successfully treat laboratory animals that suffered from type 1 diabetes.

Type 1 diabetes is, to a large extent, a disease of the immune system, since a large majority of type 1 diabetes patients have immune cells that recognize the insulin-secreting beta cells as foreign and these immune cells attack and obliterate them. MSCs are a type of adult stem cell that has shown potent immune-suppressing and anti-inflammatory effects in animal and human clinical studies. Previous preclinical trials with diabetic-prone mice have demonstrated that intravenous administration of MSCs can tamp down pancreatic injury and reduce the blood sugar levels without insulin administration. However, these effects were modest and temporary.

Sackstein and his team suggested that if more MSCs could be inserted into the pancreatic islets, then more islets would be spared from immune destruction. This would yield a more complete reversal of diabetes.

MSCs tend to lack a key cell surface adhesion molecule called HCELL. HCELL mediates the homing of cells in the bloodstream to inflammatory sites. Unfortunately, direct injection of MSCs directly into pancreatic islets is not clinically feasible because the pancreas is fragile and the damage caused by injection would cause the release of hydrolytic enzymes that would degrade the rest of the pancreas and other tissues as well. In order to move intravenously administered MSCs to the sites of the immune attack, Sackstein and others engineered MSCs that expressed the HCELL homing molecule. The presence of HCELL on the surfaces of these MSCs directed them to the inflamed pancreatic islets.

The BWH team found that administering these HCELL-bearing MSCs into diabetic mice caused the MSCs to lodge in the islets. These cells decreased inflammation in the pancreas and durably normalized blood sugar levels in the mice, which eliminated the need for insulin administration; in other words they caused a sustained reversal of diabetes

Sackstein concluded that while further studies of the effects of MSCs are warranted, this preclinical study represents an important step in the potential use of mesenchymal stem cells in the treatment of type 1 diabetes and other immune-related diseases.