Figure 4.

Exogenous IL-6 activates Stat3-SOCS3 signaling during myogenic differentiation. C2C12 myoblasts were cultured in GM until subconfluence and then shifted to DM or DM containing 25 ng/mL of recombinant IL-6. After 24 h or 48 h in the differentiation media, cells were harvested and total protein was extracted, or total RNA was extracted and first-strand cDNA was generated from 1 μg of total RNA. (a) Cell lysates (70 μg of total proteins) were subjected to Western blotting analysis with the indicated specific antibodies. The experiment shown is representative of three independent determinations. (b) Real-time PCR was performed on cDNA, using the indicated specific Taqman probes. Expression data were normalized using actin beta (ACTB) as housekeeping gene. The experiment shows means ± standard deviation of three independent determinations. **P < 0.005; n.s., not significant (Student's t test).