2013 - CTS 2013 Congress

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Hot Topics in Cell Transplantation

3.2 - Effect of DNA demethylation on myoblasts differentiation and myogenesis

Presenter: Ileana , Terruzzi, Milan, Italy
Authors: Pamela Senesi1,2,3, Livio Luzi1,2,3, Anna Montesano1,2,3, Ileana Terruzzi1,2,3

DNA demethylation enhances myoblasts hypertrophy during the late phase of myogenesis activating the IGF-1 pathway

Pamela Senesi1,2,3, Livio Luzi1,2,3, Anna Montesano1,2,3, Ileana Terruzzi1,2,3

1Department of Biomedical Sciences for Health., University of Milan, Milan, Italy; 2Research Centre, San Donato Hospital and Scientific Institute, San Donato Hospital and Scientific Institute, Milan, Italy; 3Nutrition-Metabolism Unit, San Raffaele Scientific Institute, Milan, Italy

Skeletal muscle regeneration and hypertrophy are important adaptive responses to both  physical activity and pathological stimuli. This research was performed to investigate the action of DNA demethylation on the late phase of muscle differentiation and early stage of hypertrophy.
The epigenetic process involved in myogenesis was studied with the DNA-demethylating agent 5-azacytidine (AZA). We induced muscle differentiation in C2C12 mouse myoblasts in presence of 5µM AZA and growth (GM) or differentiation (DM) medium for 48, 72 and 96 hours. To study a potential AZA hypertrophic effect, we stimulated 72h differentiated myotubes with AZA for 24h. Unstimulated cells were used as control. By Western blot and immunofluorescence analysis, we examined AZA action on myogenic regulatory factors expression, hypertrophic signaling pathway and myotube morphology.
During differentiation, protein levels of myogenic markers, Myf6 and Myosin Heavy Chain (MyHC), were higher in AZA stimulated cells compared to control. Myostatin and p21 analysis revealed morphological changes which reflect a tendency to hypertrophy in myotubes. In AZA stimulated neo formed myotubes, we observed that IGF-1 pathway, kinases p70 S6 and 4E-BP1 were activated. Furthermore, AZA treatment increased MyHC protein content in stimulated neo myotubes.
Our work demonstrates that DNA demethylation could play an important role in promoting the late phase of myogenesis, activating endocellular pathways involved in protein increment and stimulating the hypertrophic process.

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