Abstract 18106: Reprogramming of Skeletal Myoblasts for Induction of Pluripotent Status: MicroRNA Profiling and Cardiomyogenesis
Background: Encompassing the advantages of SMs, we report SMs derived iPS (SM-iPS) cells and their microRNA (miR) profile during induction of pluripotent status and cardiogenesis in comparison with native SMs (NatSMs).
Methods and Results: The pMXs vector containing mouse cDNAs for Yamanaka's quartet of pluripotency factors were used for transduction of SMs purified from male Oct4-GFP+ transgenic mouse. Three weeks later, GFP+ colonies of SM-iPS cells, having morphological similarity with mouse embryonic stem (ES) cells, were isolated and propagated in vitro. SM-iPS cells were positive for alkaline phosphatase staining, expressed SSEA1 and displayed a panel of pluripotency markers similar to ES cells. SM-iPS derived embryoid bodies yielded beating cardiomyocyte-like cells that were positive for early and late cardiac specific markers such as myosin heavy chain, troponin-I, connexin-43, MEF-2c and NKX2.5. miR profiling showed abrogation of let-7 family in SM-iPS cells (P,0.05 vs native myoblasts). Of the five miR-200 family members, miR-200a-c were not detected in NatSMs and SM-iPS cells but were found in SM-iPS cell derived cardiomyocytes, which indicated that NatSMs were not totally differentiated, however, still retaining some undifferentiated characteristics. On the contrary, ES cell specific family of miR-290 to 295 which was regulated by Oct, Nanog and Sox, was upregulated in SM-iPS cells which indicated that SM-iPS cells were similar to ES cells in their miR profile. In vivo studies in an experimental animal model of acute myocardial infarction showed extensive survival of SM-iPS cells and SM-iPS cell derived cardiomyocytes in the mouse heart 7 days after transplantation. Our results from 4-week studies in control DMEM without cells (group-1), native SMs (group-2), SM-iPS cells (group-3) and SM-iPS cell derived cardiomyocytes (group-4) showed extensive myogenic integration of the transplanted cells in group-4 with attenuation of infarct size.
Conclusions: Successful generation of SM-iPS cells and their cardiac differentiation were observed in vitro. Mir profiling revealed down-regulation of differentiated tissue specific let-7 family and induction of ES cell specific miR-290–295 cluster during reprogramming of SM into iPS cells.
- © 2010 by American Heart Association, Inc.