. REGULATION OF MAPKs AND GENES EXPRESSION BY SOLANUM GLAUCOPHYLLUM LEAF?S EXTRACTS DURING DIFFERENTIATION OF MUSCLE CELLS

GONZALEZ PARDO V; BUITRAGO C; A. PAULA IRAZOQUI; AMUSQUIVAR A

Workshop; 20 Workshop on Vitamin D; 2017

Solanum glaucophyllum leaf?s extracts (SGE) enriched with 1,25(OH)2VitaminD3-glucosides are available for the organism due to the presence of endogenous glycosidases. We have shown that SGE alike 1,25(OH)2D3 (1,25D) induces myoblast differentiation. In this work we investigate Akt role in myoblast fusion to form myotubes during differentiation. The fusion index, was determine staining C2C2 cells with MitoTracker Red prior fixation and then cells were stained with DAPI and later examined by fluorescence microscopy. We found that the ability of vitamin D compounds, 1,25D or SGE to induce myotube formation was impaired in presence of Akt inhibitor LY294002. Since Akt inhibitor suppresses myotube formation, in parallel we investigate whether Akt participates in the late differentiation marker MHC2b expression founding that its mRNA expression was induced by 1,25D and SGE in an AKT dependent manner. The commitment of myogenic cells in skeletal muscle differentiation requires earlier irreversible interruption of the cell cycle. To evaluate the effect of SGE on the distribution of C2C12 cells into different phases of the cellular cycle we performed flow cytometry assays. The studies showed that SGE stimulation, similarly to 1,25D, prompts a peak of S-phase followed by an arrest in the G0/G1-phase, events which were dependent on the MAP kinases ERK1/2, p38 and JNK. We also investigated whether 1,25D or SGE regulate the immediate early gene c-myc in C2C12 cells undergoing proliferation. We found that 1,25D but not SGE induce c-myc mRNA expression. Significant differences of the data between control and treated conditions were analyzed by one way-ANOVA followed by Bonferroni test (p< 0.05) or t-test (*p