Abstract 364: Valproic Acid Induced Reprogramming of Adult Somatic Cells for Cardiac Differentiation
Aims: Owing to the ethical concerns for use of embryonic stem cells (ESC), adult somatic cells are attractive stem cell sources for reprogramming to pluripotency. We report here non-viral approach for reprogramming of skeletal myoblasts (SM) using a small molecule.
Methods and Results: SM purified from young male Oct3/4-GFP+ transgenic mouse were treated for 5 days with valproic acid (VPA), a histone deacetylase (HDAC) inhibitor. Three weeks later, GFP+ colonies of SM derived iPSC (Sk-iPS) resembling with mouse embryonic stem cells were observed and propagated in vitro. SiPS were positive for alkaline phosphatase, had normal karyotype, expressed SSEA1, and induced teratomas in nude mice containing tissue comprising all three germ layers. RT PCR analysis showed that Sk-iPS cells expressed Oct4, Sox2, KLF4, c-Myc, Nanog and ESC specific pluripotency genes. HDAC1 activity was significantly reduced in Sk-iPs generated with valproic acid treatment as compared to ES cells. Sk-iPS derived embryoid bodies (EBs) yielded spontaneously contracting cardiomyocytes with morphological, molecular, and ultrastructural features of developing cardiomyocytes. These cells were also positive for early and late cardiac markers such as myosin heavy chain, Gata4, Mef-2c and Nkx2.5, Connexin-43 (P<0.01vs native SM). Micro RNA (miR) profiling showed abolition of let-7 family in Sk-iPS whereas ESC specific family of miR-290-295 was upregulated which indicated that Sk-iPS possessed miR profile similar to ESC. However muscle specific miRNAs (miR -133, -206) were identified in Sk-iPS cells as compared to ES cells indicating that the Sk-iPS retained the epigenetic memory of myogenic origin.
Conclusions: We conclude that SM with endogenous expression of Sox2, KLF4, and cMyc are suitable candidates to generate iPS cells without viral vectors using a single small molecule.
- © 2012 by American Heart Association, Inc.