Rich Lowry at National Review has yet another take on Rick Perry’s treatment. It’s a good read.
Texas governor and Republican presidential candidate Rick Perry has received a fair amount of criticism for receiving a stem cell treatment for his back. The Rising Voice blog has another take on Perry’s treatment, which, by the way, was paid for with his own money and was his free choice to receive. Even if the procedure was somewhat experimental, Perry made his own informed decision to receive it. ll these criticism aside, some critics need to get over themselves. No one was harmed through Perry’s decision to receive this treatment, and Perry sought the treatment even though he knew it wasn’t FDA approved or endorsed. Apparently, the clinic that offered the treatment had garnered enough evidence to convince Perry, and he acquiesced to the treatment. If this inspires other people to seek similar treatments, then they will need to make their own informed decisions based on the evidence in hand. If the clinic is honest and transparent about their procedure and the evidence for it, then we should rejoice that Governor Perry received successful treatment without burdening the present health care system. That sounds like a win-win situation to me.
Scientists succeeded in making induced pluripotent stem cells in pigs (Zhao Wu, et al. Generation of Pig Induced Pluripotent Stem Cells with a Drug-Inducible System. J Mol Cell Biol 1, no. 1 (2009): 46-54). Pig iPSCs are a valuable research tool, since pig cells have a greater similarity to humans than do mouse cells. Also, pig iPSCs allow for the production of chimeric animals that are partially composed of cells descended from one cell type and also composed of cells from a different cell type. Chimeric animals that are partially made from iPSCs allow for the study of how effectively iPSCs cause tumors and other iPSC issues. This is an essential study because chimeric mice that partially consist of iPSCs showed that iPSC derived chimeras possessed large numbers of tumors. This result raises significant concerns about the safety of iPSC therapies. Additionally, pig iPSCs can generate chimeras whose gametes are made from iPSCs. This can potentially revolutionize the transgenic animal field by enabling complex genetic manipulations (e.g. knockout or knockin of genes) to produce biomedically important large animal models or improve livestock production.
In this study, Steen Stice’s research group demonstrated the germline transmission of iPSCs with the live birth of a transgenic piglet that possessed genome integration of two human genes. Additionally, gross and histological examination of necropsied porcine chimeras at 2, 7 and 9 months showed these animals lacked tumor formation and demonstrated normal development. The development of germline competent porcine iPSCs that do not produce tumors in young chimeric animals presents an attractive and powerful translational model to study the efficacy and safety of stem cell therapies and perhaps to efficiently produce complex transgenic animals.
In 2008, scientists from Thomas Jefferson University in Philadelphia, Pennsylvania and the University of Heidelberg, Germany published a study that shows the conversion of bone marrow stem cells into cells that look like heart muscle cells. They cultured rat bone marrow stem cells with heart muscle cells from baby rats and then added two growth factors that are commonly found in developing heart. These growth factors include fibroblast growth factor-2 (FGF-2) and bone morphogen protein-2 (BMP-2). They cultured the bone marrow stem cells with the baby rate heart muscle cells for ten days and added either BMP-2, FGF-2 or FGF-2 + BMP-2. Neither BMP-2 or FGF-2 alone could elicit any significant change in the bone marrow cells, but when FGF-2 and BMP-2 were added together, the caused the bone marrow stem cells to express heart muscle-specific genes.
Transformed bone marrow stem cells made transcription factors like Nkx2.5 and GATA-4. These two transcription factors are expressed in developing heart cells, and the production of these transcription factors in bone marrow stem cells demonstrates that they are differentiating into heart cells. These transformed bone marrow stem cells also produced connexion-43, which is a protein that allows calcium ions to pass from one heart muscle cell to another so that they beat simultaneously. Since heart muscle cells specifically make this protein, it suggests that the bone marrow stem cells are becoming heart cells.
Even more remarkably, the transformed bone marrow stem cells showed calcium-handling proteins and calcium currents that occurred as a result of stimulation. Heart muscle cells spontaneously form calcium currents in response to a flood of calcium ions from the neighboring cells. Heart muscle cells have calcium channels called L-type calcium channels. Particular drugs block these channels, and they are sometimes prescribed to lower blood pressure. The inducible calcium channels in the transformed bone marrow stem cells could be inhibited by these calcium channel blocking drugs, which argues that the calcium currents are due to proteins that are usually expressed in heart muscle cells.
These data argue that bone marrow stem cells can become heart muscle cells. Bone marrow treatments have been used in clinical trials for heart attack patients. However, the ability of bone marrow cells to differentiate into heart muscle cells is limited. Even though there is plenty of evidence that bone marrow stem cells can become heart muscle cells, they appear to do so at a very low-frequency. Therefore, changing bone marrow stem cells into heart muscle cells should greatly increase the therapeutic capacity of these cells. Treatment of human bone marrow stem cells with BMP-2 and FGF-2 might transform bone marrow stem cells into heart muscle cells, which would augment the ability of bone marrow stem cells to treat heart attack patients.
A study published in the journal Blood, (Journal of the American Society of Hematology) shows that administration of special chemical called “granulocyte colony-stimulating factor (G-CSF)”, which mobilizes bone marrow stem cells and releases them from the bone marrow into the blood, is unlikely to put healthy stem cell donors at risk for later development of cancer-causing chromosomal abnormalities. The chromosome abnormalities in question involve loss or gains of chromosomes that have been linked to blood-based disorders such as myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML).
When given to stem cell donors, G-CSF moves the donor’s stem cells from the bone marrow into the blood stream. This process is called “mobilization.” Once in the blood, physicians and scientists can collect large doses of stem cells from the peripheral blood safely and without invasive surgery (this process is called “apheresis”). Apheresis avoids bone marrow harvests in the operating room, which can be painful, stressful, and require some recovery time. Research has shown that the large doses of mobilized stem cells, called peripheral blood stem cells or PBSCs, can repopulate the bone marrow and create new blood cells faster than stem cells collected directly from the bone marrow. Furthermore, several long-term follow-up studies have demonstrated that healthy donors are not at an increased risk of developing leukemia or other cancers following PBSC donation.
Betsy Hirsch, first author of the study and Associate Professor in the Department of Laboratory Medicine and Pathology at the University of Minnesota Medical School said: “In recent years, PBSCs have become the dominant source of stem cells for stem cell transplants and the number of transplants performed with PBSCs that have been mobilized with G-CSF has substantially increased. However, the potential for the therapy to cause DNA damage, mutations, or cancer had been suggested in a smaller and less comprehensive prior study, raising a serious concern within the transplant community and making a definitive study very important. Our study aimed at investigating potential effects of G-CSF on chromosomes in healthy donors,”
A research team from the University of Minnesota Medical School conducted a study to determine if G-CSF therapy is indeed a potential cause of chromosome loss or gain in stem cell donors. They asked if there was any risk with short-term, low-dose usage of G-CSF on healthy PBSC donors. Their study evaluated blood samples taken from 22 PBSC donors who had received G-CSF and 22 controls patients who had no history of cancer or prior exposure to G-CSF therapy over a 12 month period.
By using a technique called fluorescence in situ hybridization or FISH, the researchers evaluated the white blood cells of the study subjects for abnormal number of chromosomes. FISH is a sensitive laboratory technique that can detect specific targeted regions of DNA in an individual’s cells to identify chromosomal abnormalities. Loss and gain of chromosomes represent one form of chromosome instability that is frequently a step in the development of cancer. Specifically, the researchers focused on chromosome 7 and a series of other chromosomal regions that are documented to be associated with MDS and AML. The researchers also evaluated the white blood cells from these patients to determine if both copies of each chromosome replicated synchronously or asynchronously. Asynchronous replication can also signal genomic instability and a higher risk of chromosomal abnormalities, and if G-CSF stimulates asynchronous chromosomal replication, if could predispose stem cell donors to higher cancer rates.
The outcomes of this research, however are clear. According to Betty Hirsch: “Contrary to the previously published data, our study concludes that G-CSF stimulation does not result in replication asynchrony or induce the atypical levels of abnormality for chromosome 7 or other chromosomes that have been associated with MDS and AML, and we expect that these results can be generalized to all chromosomes,” said Dr. Hirsch.
Jeffrey McCullough, MD, senior study author and Professor in the Department of Laboratory Medicine and Pathology at the University of Minnesota Medical School, added, “Furthermore, our data support the conclusion that G-CSF does not induce chromosomal instability through the PBSC mobilization process and remains a safe therapy for healthy stem cell donors.”
An interesting paper was published in Asian Cardiovascular and Thoracic Annals: Soma Guhathakurta, Usha R Subramanyan, Ramesh Balasundari, Chandan K Das, Nainar Madhusankar, and Kotturathu Mammen Cherian, Stem Cell Experiments and Initial Clinical Trial of Cellular Cardiomyoplasty Asian Cardiovasc Thorac Ann, Dec 2009; 17: 581 – 586.
In this paper, several Indian scientists used bone marrow cells to test three possibilities: 1) to determine whether transplantation of a sheep’s own bone marrow stem cells into the infarcted myocardium could promote the differentiation of those bone marrow cells into beating heart muscles; 2) demonstrate that bone marrow mesenchymal cells could differentiate into heart muscle cells; 3) conduct a clinical trial with patient’s own bone marrow to determine if it could improve the heart function of patients with severe heart problems.
The first stage used five experimental sheep in which a heart attack had been induced (coronary artery ligation), and two control sheep (they started with nine, but one control animal and one experimental animal died). The control sheep did not show any improvement, but the experimental sheep, which received 100,000 to 1,000,000 bone marrow mononuclear cells (injected into the periphery of the infarcted area), showed improvements in blood circulation throughout the heart. Echocardiography demonstrated marginal improvements in ejection fraction at 3 months post-injection. Staining of the heart tissue from the injection site revealed that some of the heart tissue that should have died was still alive. An even more remarkable finding was that throughout the scar, islands of muscle tissue that could not be differentiated from the
surrounding muscle tissue. Since four of the experimental sheep had been injected with unmarked bone marrow, this large island of heart muscle tissue in the heart scar had a different look from the rest of the surround heart muscle.
One experimental animal, however, had been infected with a virus that causes the bone marrow to glow (Green Fluorescent Protein). When this experimental animal was examined with UV light, the island of heart muscle tissue in the scar glowed, demonstrating that the new heart muscle is from the implanted bone marrow.
In the second stage of their experiment, the researchers tried to grow bone marrow cells in culture and differentiate them into heart muscle cells. The technique they used is apparently undergoing a patent process. Therefore, they did not reveal the technique. Nevertheless, the bone marrow cells formed structures that looked like heart muscle, expressed many heart muscle genes (GATA-4, Nkx 2.5, hANP, MLC-2v and MLC-2a), and in the electron microscope, had a definite heart muscle-like structure, but without the structures that connect heart muscle cells together (intercalated discs).
A marginal improvement in myocardial function was noted at 3 months (mean increase in ejection fraction, 6% ±1%), although it plateaued at 6 months. The trial proved tobe safe because there was no procedure-related mortality. There is growing optimism that stem cell therapy may delay heart transplantation.
Finally, they transplanted bone marrow cells into the hearts of 29 patients who suffered from “dilated cardiomyopathy,” which simply means that they have an enlarged heart, or “endstage ischemic cardiomyopathy,” in which the blood vessels of the heart are so clogged that the patient’s heart cannot receive adequate amounts of oxygen even at rest. Also 11 other patients with similar heart problems were injects with endothelial progenitor cells (EPCs), which are also found in bone marrow, but make blood vessels, and occasionally heart muscle. One particular patient was 5-months old and had a congenital case of cardiomyopathy. In this patients, EPC injections caused a spike in the ejection fraction from 32% to 58%. Other EPC-injected patients produced mixed results,from no improvement to a 7% increase in ejection fraction, with no mortality. The differences in the results is probably due to the fact that they used three different ways to deliver these stem cells; intracoronary delivery, direct injection into the heart muscle, and infusion through the pulmonary arteries. A marginal improvement in heart function was noted at 3 months (mean increase in ejection fraction, 6% ±1%), although it plateaued at 6 months. The trial proved to be safe because there was no procedure-related mortality.
This study is a nice example of the combination of animals, test-tube, and clinical studies. The problem is that the clinical study is too unfocused to properly interpret. It is small, and non-randomized. There is no placebo, and the patients showed a wide variety of chronic heart conditions. The fact that no one became horribly ill because of the procedure is encouraging, but the results are all over the board, as are the patients and their conditions, which makes the clinical trial inconclusive with respect to efficacy. This study definitely justifies a larger, more focused, randomized study. Also, the animal and in vitro portion of the study gives excellent, positive evidence that mesenchymal stem cells from bone marrow can transdifferentiate into heart muscle cells. However, the in vitro work shows that the heart muscle made by the mesenchymal stem cells did not make connexion-43, which is essential for gap junctions and physiological connection between the heart muscle cells. This lack of physiological integration can lead to functional isolation of the new heart muscle tissue, which can generate arrhythmias. Engineering these cells with Connexin-43 would be a good start, which has been shown in skeletal muscle cells to prevent functional isolation and arrythymias (see Tolmachov O, et al. Overexpression of connexin 43 using a retroviral vector improves electrical coupling of skeletal myoblasts with cardiac myocytes in vitro. BMC Cardiovasc Disord. 2006 Jun 6;6:25). The second concern is that these presumptive heart muscle cells were not tested for calcium handling proteins. This is important, since some publications that shown that bone marrow stems cells can form cells that approximate heart muscle cells, but these cells lack the calcium-handling machinery necessary to generate a heart beat (Scherschel JA, Soonpa MH, Srour EF, Field LJ and Rubart M (2008). Adult bone marrow–derived cells do not acquire functional attributes of cardiomyocytes upon transplantation into peri-infarct myocardium. Mol Ther 16: 1129–1137). These scientists should go back to look at this as well.
All in all this is a fascinating study that provides some excellent evidence, but also leaves many hanging questions.
Christopher Centeno, who performs stem cells treatments for patients who are considering knee replacement, has submitted another paper that examines the outcomes of his techniques. His former publication examined 227 patients who were followed for between 3 months and 2-3 years. This paper also utilized data from 50-60 ultra-high field MRIs of the re-implant sites. The results were quite encouraging.
Their new paper has 339 patients followed for up to 3-4 years with 210 MRIs of the places where cells were re-implanted. This paper also includes knee outcome data for the “Regenexx-C” procedure. The data included in this work shows that the Regenexx-C procedure is dramatically safer than the knee replacement surgeries it helped many patients avoid. The paper is still in the galley proof stage, and therefore, it will be a few months before it makes it into press.
Centeno and his colleagues also have other publications on the way. There is a paper that focuses on knee/hip patient outcome, which will be published sometime this year or early next (the paper is winding it’s way through the scientific review process). Stem cell orthopedic treatments come in all sorts of shapes and sizes, but many of them are subjected to peer-review. Make sure the specific therapy you’re being offered has data showing it works.