FIGURE 3.

Hypertrophic agonists stimulate increases in tubulin content and stability in cardiac myocytes. Primary cultures of neonatal rat cardiac myocytes were treated for 24 h with PE (100 μm), LIF (10 ng/ml), OSM (10 ng/ml), or 0.1% (w/v) BSA in PBS as a non-stimulated control (Con). A, protein lysates from treated cardiac myocytes were immunoblotted for α-tubulin and post-translationally modified glu- and acetyl-tubulin. Blotting for GAPDH demonstrated equivalent protein loading. Glu-tubulin (B) and acetyl-tubulin (C) bands from immunoblots (n = 5) were quantitated by densitometric analysis, normalized for α-tubulin, and expressed as average fold change + S.E. Significant changes in glu- and acetyl-tubulin compared with Con are denoted by †, p < 0.05 and *, p < 0.05, respectively. D, immunostaining for glu-tubulin (upper panels) and co-staining with TRITC-conjugated phalloidin (lower panels) revealed the stabilized MT network and the organization of contractile actin filaments, respectively. E, Con or LIF-stimulated cardiac myocytes were treated with nocodazole (NZ, 2 μm, 1 h) before immunostaining for α-tubulin. Scale bars, 50 μm.