tretinoin and Myocardial-Ischemia

Bone marrow-derived CD34(+) cells are currently used in clinical trials in patients with ischemic heart disease. An option to enhance activity of injected progenitors may be offered by genetic engineering of progenitor cells with angiogenic growth factors. Recombinant adeno-associated viral vectors (rAAV) have emerged as a leading gene transfer systems. In contrast to other vector systems in use for genetic engineering of CD34(+) cells, rAAV-mediated gene expression does not depend on vector integration. This is relevant for application in regenerative medicine of ischemic tissues, where transient transgene expression is likely sufficient to achieve therapeutic benefits.. We compared three different human AAV serotypes, packaged as pseudotypes by a helper virus-free production method, for their transduction efficiency in human cord blood-derived CD34(+) cells. We further assessed the impact of vector genome conformation, of alpha(v)beta(5) and alpha(5)beta(1) integrin availability and of the transcription-modulating drugs retinoic acid and Trichostatin A on rAAV-mediated human CD34(+) cell transduction.. We provide, for the first time, evidence that hCD34(+) cells can be reproducibly transduced with high efficiency by self-complementary rAAV2 without inducing cytotoxicity or interfering with their differentiation potential. We further show the involvement of alpha(5)beta(1) integrin as a crucial AAV2 internalization receptor and a function for transcription-modulating drugs in enhancing rAAV-mediated transgene expression.. This study represents a first step toward translation of a combined cellular/rAAV-based therapy of ischemic disease.

Topics: Antigens, CD34; Antineoplastic Agents; Clinical Trials as Topic; Dependovirus; Fetal Blood; Genetic Vectors; Genome, Viral; Humans; Hydroxamic Acids; Integrin alpha5beta1; Myocardial Ischemia; Protein Synthesis Inhibitors; Stem Cell Transplantation; Stem Cells; Transcription, Genetic; Transduction, Genetic; Tretinoin

Cell transplantation to repair or regenerate injured myocardium is a new frontier in the treatment of cardiovascular disease. Most studies on stem cell transplantation therapy in both experimental heart infarct and in phase-I human clinical trials have focused on the use of undifferentiated stem cells. Based on our previous observations demonstrating the presence of multipotent progenitor cells in human adult skeletal muscle, in this study we investigated the capacity of these progenitors to differentiate into cardiomyocytes. Here we show an efficient protocol for the cardiomyogenic differentiation of human adult skeletal muscle stem cells in vitro. We found that treatment with Retinoic Acid directed cardiomyogenic differentiation of skeletal muscle stem cells in vitro. After Retinoic Acid treatment, cells expressed cardiomyocyte markers and acquired spontaneous contraction. Functional assays exhibited cardiac-like response to increased extracellular calcium. When cocultured with mouse cardiomyocytes, Retinoic Acid-treated skeletal muscle stem cells expressed connexin43 and when transplanted into ischemic heart were detectable even 5 weeks after injection. Based on these results, we can conclude that human adult skeletal muscle stem cells, if opportunely treated, can transdifferentiate into cells of cardiac lineage and once injected into infarcted heart can integrate, survive in cardiac tissue and improve the cardiac function.

Topics: Adult; Aged; Animals; Becaplermin; Cell Differentiation; Humans; Mice; Mice, Inbred Strains; Middle Aged; Muscle, Skeletal; Myocardial Ischemia; Myocytes, Cardiac; Phenotype; Platelet-Derived Growth Factor; Proto-Oncogene Proteins c-sis; Stem Cells; Tretinoin

Previous studies have shown that free polyunsaturated fatty acids (PUFA) reduce the excitability of cardiac myocytes and exert antiarrhythmic effects. Therefore, we hypothesized that retinoic acid (RA, vitamin A acid), which has structural characteristics similar to those of PUFA, may have similar antiarrhythmic effects. To test this hypothesis, we used an isolated, spontaneously beating, neonatal rat cardiac myocyte preparation to examine the effects of RA, added to the perfusion solution, on the cell contraction and arrhythmias induced by isoproterenol (ISO) or lysophosphatidylcholine (LPC). All-trans-RA (10-20 microM) induced a marked and reversible reduction in the contraction rate of the cell in 2-5 min without changing the amplitude of the contractions. Superfusion of the myocytes with either ISO (3 microM) or LPC (5 microM) induced sustained tachyarrhythmias characterized by spasmodic contractures and fibrillation. Addition of 15-20 microM all-trans-RA to the perfusion solution effectively prevented as well as terminated the arrhythmias induced by ISO and LPC. Furthermore, in a whole-animal model of arrhythmia in which the left anterior descending coronary artery (LAD) of the anesthetized rat was occluded for 15 min followed by reperfusion, both the incidence and severity of ventricular tachycardia and fibrillation (VT, VF) were significantly reduced during the ischemic and reperfusion periods by intravenous infusion of all-trans-RA. In contrast, other analogues, including retinol and retinal, and other fat-soluble vitamins, including vitamin D, E, and K, did not have such effects. Our results demonstrate that all-trans-RA can produce antiarrhythmic effects similar to those of PUFA, suggesting a novel role of RA as a potential antiarrhythmic agent.

Topics: Animals; Anti-Arrhythmia Agents; Arrhythmias, Cardiac; Cells, Cultured; Isoproterenol; Lysophosphatidylcholines; Myocardial Contraction; Myocardial Ischemia; Myocardial Reperfusion; Rats; Rats, Sprague-Dawley; Tretinoin