Figure 1.

E2β stimulates dynamic S-nitrosylation in CFL1 in HUVECs via eNOS-derived NO. A, Subconfluent HUVECs were treated by 10 nM of E2β with or without 1 mM of L-NAME for the indicated time periods. Total protein extracts were harvested and subjected to biotin switch and avidin capture. The biotin-labeled SNO proteins were analyzed by immunoblotting with anti-CFL1 antibody. *, P < .05 vs untreated control. B, HUVECs were transfected with or without scrambled or eNOS-specific siRNAs, followed by treatment with or without E2β for 30 minutes. SNO-CFL1 was measured. Representative blots of SNO-CFL1, total CFL1, eNOS, and β-actin of a typical experiment are shown. Graphs summarize data (mean ± SEM) from three independent experiments using cells from different fetuses. Data were normalized and expressed fold to control. Bars with different letters differ from each other significantly; paired comparisons of importance were indicated with an asterisk (P < .05).