TY - JOUR
T1 - Efficient non-viral reprogramming of myoblasts to stemness with a single small molecule to generate cardiac progenitor cells
AU - Pasha, Zeeshan
AU - Haider, Husnain Kh
AU - Ashraf, Muhammad
PY - 2011
Y1 - 2011
N2 - The current protocols for generation of induced pluripotent stem (iPS) cells involve genome integrating viral vectors which may induce tumorgenesis. The aim of this study was to develop and optimize a non-viral method without genetic manipulation for reprogramming of skeletal myoblasts (SMs) using small molecules. Mzethods and Results: SMs from young male Oct3/4-GFP + transgenic mouse were treated with DNA methyltransferase (DNMT) inhibitor, RG108. Two weeks later, GFP + colonies of SM derived iPS cells (SiPS) expressing GFP and with morphological similarity of mouse embryonic stem (ESCs) were formed and propagated in vitro. SiPS were positive for alkaline phosphatase activity, expressed SSEA1, displayed ES cell specific pluripotency markers and formed teratoma in nude mice. Optimization of culture conditions for embryoid body (EBs) formation yielded spontaneously contracting EBs having morphological, molecular, and ultra-structural similarities with cardiomyocytes and expressed early and late cardiac markers. miR profiling showed abrogation of let-7 family and upregulation of ESCs specific miR-290-295 cluster thus indicating that SiPS were similar to ESCs in miR profile. Four weeks after transplantation into the immunocompetent mice model of acute myocardial infarction (n = 12 per group), extensive myogenesis was observed in SiPS transplanted hearts as compared to DMEM controls (n = 6 per group). A significant reduction in fibrosis and improvement in global heart function in the hearts transplanted with SiPS derived cardiac progenitor cells were observed. Conclusions: Reprogramming of SMs by DNMT inhibitor is a simple, reproducible and efficient technique more likely to generate transgene integration-free iPS cells. Cardiac progenitors derived from iPS cells propagated extensively in the infarcted myocardium without tumorgenesis and improved cardiac function.
AB - The current protocols for generation of induced pluripotent stem (iPS) cells involve genome integrating viral vectors which may induce tumorgenesis. The aim of this study was to develop and optimize a non-viral method without genetic manipulation for reprogramming of skeletal myoblasts (SMs) using small molecules. Mzethods and Results: SMs from young male Oct3/4-GFP + transgenic mouse were treated with DNA methyltransferase (DNMT) inhibitor, RG108. Two weeks later, GFP + colonies of SM derived iPS cells (SiPS) expressing GFP and with morphological similarity of mouse embryonic stem (ESCs) were formed and propagated in vitro. SiPS were positive for alkaline phosphatase activity, expressed SSEA1, displayed ES cell specific pluripotency markers and formed teratoma in nude mice. Optimization of culture conditions for embryoid body (EBs) formation yielded spontaneously contracting EBs having morphological, molecular, and ultra-structural similarities with cardiomyocytes and expressed early and late cardiac markers. miR profiling showed abrogation of let-7 family and upregulation of ESCs specific miR-290-295 cluster thus indicating that SiPS were similar to ESCs in miR profile. Four weeks after transplantation into the immunocompetent mice model of acute myocardial infarction (n = 12 per group), extensive myogenesis was observed in SiPS transplanted hearts as compared to DMEM controls (n = 6 per group). A significant reduction in fibrosis and improvement in global heart function in the hearts transplanted with SiPS derived cardiac progenitor cells were observed. Conclusions: Reprogramming of SMs by DNMT inhibitor is a simple, reproducible and efficient technique more likely to generate transgene integration-free iPS cells. Cardiac progenitors derived from iPS cells propagated extensively in the infarcted myocardium without tumorgenesis and improved cardiac function.
UR - http://www.scopus.com/inward/record.url?scp=80051728762&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=80051728762&partnerID=8YFLogxK
U2 - 10.1371/journal.pone.0023667
DO - 10.1371/journal.pone.0023667
M3 - Article
C2 - 21886809
AN - SCOPUS:80051728762
SN - 1932-6203
VL - 6
JO - PloS one
JF - PloS one
IS - 8
M1 - e23667
ER -