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Figure 4 Effect of French red wines on endothelial-type nitric oxide synthase (eNOS) protein expression and eNOS activity in EA.hy 926 cells. (A) Western blot analyses using a polyclonal anti-eNOS- and a monoclonal anti-ß-tubulin-antibody (for normalization). Cells were incubated for 48 h with ethanol (EtOH-Co, 1.25%, v/v) or two different French red wines ("Ch.Bonnet reserve" and "L.Chevaliers", 1%, 3%, and 10%, v/v). The blots represent four independent experiments each. (B) This panel shows the effect of a 48-h incubation with "L.Chevaliers" and "Ch.Bonnet reserve" on the nitric oxide (NO) production in EA.hy 926 cells stimulated for 2 min with the calcium ionophore A23187. Conditioned medium from EA.hy 926 cells was transferred to RFL-6 reporter cells and incubated for 2 min. Thereafter, cGMP content in RFL-6 cells was determined by radioimmunoassay. Basal cGMP content of the RFL-6 cells (1.7 ± 0.16 pmol/10⁶ cells) was subtracted from all samples. The NO production of ethanol-treated EA.hy 926 cells (EtOH-Co, 13.1 ± 0.19 pmol cGMP/10⁶ RFL-6 cells) was set 100%. Data represent means ± SEM of three independent experiments. Asterisks indicate significant differences from ethanol-treated cells (*p < 0.05; **p < 0.01; by ANOVA followed by the Fisher PLSD test).