FIG. 2.

Akt is induced upon myocyte differentiation. (A) Akt protein upregulation during myogenesis induced by mitogen deprivation. C2C12 cells were cultured in growth medium (GM) or in differentiation medium (DM) for 24 h (1d) or for 48 h (2d). Cell lysates containing 20 μg protein were subjected to Western immunoblot analysis using anti-Akt, p21, and Cdk4 antibodies. (B) Akt protein upregulation induced by prolonged cultivation of confluent cultures in high-mitogen medium. C2C12 cells were cultured in growth medium (GM) for 2, 4, and 6 days. Cell lysates were prepared, and 20 μg of extract protein was immunoblotted with anti-Akt, myogenin, and Cdk4. (C) Akt protein expression is sustained in terminally differentiated cells following serum restimulation. C2C12 cells were induced to differentiate into myotubes by incubating cultures for 4 days in differentiation medium. These cultures were then cultured in fresh differentiation medium (DM) or growth medium (GM) for 24 h. Cell lysates (20 μg) were immunoblotted with anti-Akt and Cdk4 antibodies.