Biomedical subjects
Publications and source records attributed to Sharon Etzion.
We review an experimental protocol for investigating concepts and methods for myocardial repair using fetal cardiomyocyte transplantation. We describe methods of cell isolation, culture, labeling, and assessment of the influence of the engrafted cells on left ventricular remodeling and function.
BACKGROUND: Systemic delivery of bone marrow-derived mesenchymal stem cells (BM-MSCs) is an attractive approach for myocardial repair. We aimed to test this strategy in a rat model after myocardial infarction (MI). METHODS AND RESULTS: BM-MSCs were obtained from rat bone marrow, expanded in vitro to a purity of >50%, and labeled with 99mTc exametazime, fluorescent dye, LacZ marker gene, or bromodeoxyuridine. Rats were subjected to MI by transient coronary artery occlusion or to sham MI. 99mTc-labeled cells (4x10(6)) were transfused into the left ventricular cavity of MI rats either at 2 or 10 to 14 days after MI and were compared with sham-MI rats or MI rats treated with intravenous infusion. Gamma camera imaging and isolated organ counting 4 hours after intravenous infusion revealed uptake of the 99mTc-labeled cells mainly in the lungs, with significantly smaller amounts in the liver, heart, and spleen. Delivery by left ventricular cavity infusion resulted in drastically lower lung uptake, better uptake in the heart, and specifically higher uptake in infarcted compared with sham-MI hearts. Histological examination at 1 week after infusion identified labeled cells either in the infarcted or border zone but not in remote viable myocardium or sham-MI hearts. Labeled cells were also identified in the lung, liver, spleen, and bone marrow. CONCLUSIONS: Systemic intravenous delivery of BM-MSCs to rats after MI, although feasible, is limited by entrapment of the donor cells in the lungs. Direct left ventricular cavity infusion enhances migration and colonization of the cells preferentially to the ischemic myocardium.
The field of tissue engineering, involving the reprogramming of stem cells or rejuvenation of specific differentiated cells, is emerging as a promising strategy to repair the damaged myocardium. The eventual goal is to be able to take a patient's own cells, expand them ex vivo, genetically engineer them to enhance specific properties, and then reintroduce them into the patient's heart to create a replacement tissue. Our review paper describes data that supports the potential of this strategy. This clinically relevant, combined strategy of genetic and tissue engineering could be of importance in treating elderly patients with massive myocardial damage, patients whose normal myogenic or angiogenic cells have been depleted or are inadequate in their growth potential, to prevent LV deterioration and heart failure.
BACKGROUND: The muscle-specific MyoD family of transcription factors function as master genes that are able to prompt myogenesis in a variety of cells. The purpose of our study was to determine whether MyoD could induce primary cardiac fibroblasts, isolated from infarcted myocardium or pericardium, to undergo myogenic conversion in a clinically relevant approach. METHODS AND RESULTS: Primary rat fibroblasts from 7-day-old infarcted myocardium or normal pericardium were transfected by an E1/E3-deleted adenoviral vector carrying both a human MyoD cDNA driven by a CMV promoter and a green fluorescent protein (GFP) reporter gene driven by a second CMV promoter. Expression of MyoD caused myogenic differentiation of cultured fibroblasts, as defined by elongation and fusion into multinucleated myotubes, typical cross striation as identified by electron microscopy, and positive immunostaining for sarcomeric actin, fast myosin heavy chain (MHC), and actinin. The myogenic cells (1.5x10(6)) were transplanted into the infarcted myocardium 7 days after coronary artery occlusion. By 1 month after transplantation, the converted fibroblasts gave rise to a cluster of myogenic cells that in a few hearts occupied a large part of the scar with positive immunostaining for the myogenic proteins fast-MHC and sarcomeric actin. A few cells expressed the gap junction protein connexin 43 in a disorganized manner. There was no positive staining in the control hearts treated with injections of untreated fibroblasts or culture medium. CONCLUSIONS: Our work shows that it is possible to exploit the unique capacity of MyoD to activate myogenesis in fibroblasts ex vivo and to create a vast source of autologous myogenic cells for transplantation.