Abstract 257: Bone Marrow Cells Form New Cardiomyocytes in the Infarcted Myocardium
The ability of c-kit-positive bone marrow cells (c-kit-BMCs) to form cardiomyocytes following delivery to the acutely infarcted myocardium is a matter of debate. In an attempt to resolve the controversy, we raised the possibility that c-kit-BMCs represent a functionally heterogeneous pool, containing cells with distinct transdifferentiation potential. To test this hypothesis, c-kit-BMCs were infected with EGFP-lentiviruses and injected in infarcted hearts. At 1 and 2 weeks, the regenerated myocardium was enzymatically digested and EGFP-labeled myocytes, endothelial cells (ECs), fibroblasts, and c-kit-cells were sorted. By employing a PCR-based method of detection of viral integrants, we searched for unique sites of viral insertion in the infected c-kit-BMCs and their progeny. Common insertion sites were found in the DNA of all cell populations, documenting that single c-kit-BMCs transdifferentiated into multiple cell lineages. However, the 31 detected clones shared 9 distinct sites of integration, suggesting that myocytes in different animals were generated by specific subsets of c-kit-BMCs, which retained the ability to transdifferentiate. To strengthen the observation that myocardial regeneration is oligoclonal in nature, freshly isolated FACS-sorted c-kit-BMCs were simultaneously transduced with 3 lentiviruses, each encoding red, green or blue (RGB) fluorescent proteins. Different combinations of inserted vectors resulted in the formation of numerous c-kit-BMC clones of mixed colors. RGB-infected HSCs were injected in infarcted rats; at 4 days, engrafted c-kit-BMCs showed a polyclonal pattern characterized by the presence of cells labeled by most of the color combinations seen in vitro. However, at 2 weeks, only a few of the homogeneously colored colonies seen at 4 days persisted and formed large patches of cells composed of myocytes, ECs, and smooth muscle cells. Our findings document that c-kit-BMCs are functionally heterogeneous and have different ability to survive, engraft, and transdifferentiate in the ischemic heart. At present, clonal c-kit-BMCs are being analyzed by RNA-sequencing for the identification of the unique phenotypical properties of c-kit-BMCs with high transdifferentiation potential.
- © 2013 by American Heart Association, Inc.