Fig. 5.
Effect of TGF-β on the expression or phosphorylation levels of selected inhibitors and activators of cell proliferation in Cav-2-negative and -positive MLECs. pBABE and Cav-2-pBABE MLECs were plated at low density (3 × 105 per 150-mm dish), followed by incubation with complete medium without or with TGF-β (1 ng/ml) for 24 and 48 h. At indicated time points, cells were lysed and SDS-PAGE resolved protein lysates immunoblotted with the indicated antibodies to p27, phosporylated Rb protein (P-Rb), phosphorylated ERK1/2 (P-ERK 1/2), total ERK 1/2, phosphorylated Akt (P-Akt), total Akt, caveolins, and Hsp-90. Note that both 24 and 48 h treatment with TGF-β results in a greater increase in the expression levels of p27 in pBABE relative to Cav-2-pBABE cells (top panel and top graph). Also, note that 48 h treatment with TGF-β visibly reduces the P-Rb levels in pBABE but not Cav-2-pBABE (2nd immunoblot from the top and bottom graph). Finally, note that TGF-β does not affect P-ERK 1/2 nor P-Akt levels. B: densitometric ratios of p27 and P-Rb to Hsp90 quantified using Image J, which were selectively affected by TGF-β treatment in pBABE relative to Cav-2-pBABE cells and expressed as means ± SD (n = 3). *P < 0.05 vs. respective control sample; #P < 0.05 vs. respective TGF-β-treated sample at given time point.