Fig. 6.

eNOS signaling responses following siRNA-mediated knockdown of thiol-metabolizing proteins. A: immunoblots were analyzed in endothelial cell lysates prepared from bovine aortic endothelial cells (BAEC) transfected with control, GR, TrxR1, or TrxR2 siRNA targeting constructs as shown, and then treated with VEGF for the indicated times. Cell lysates were resolved by SDS-PAGE and probed in immunoblots using antibodies directed against phosphoserine¹¹⁷⁹-eNOS, phosphoserine¹¹⁶-eNOS, total eNOS, p-Akt, and total Akt. Shown are representative data of four independent experiments. B: results of densitometric analyses from pooled data, plotting the ratios of p-eNOS and p-Akt to total eNOS and Akt, respectively, relative to the signals present in the unstimulated cells. Each value represents the mean ± SE derived from four independent experiments. *P < 0.01 for VEGF vs. control cells. #P < 0.01 for TrxR1 siRNA-transfected cells vs. control siRNA-transfected cells control cells (ANOVA).