Endothelium negatively modulates the vascular relaxation induced by nitric oxide donor, due to uncoupling NO synthase

Nitrosyl ruthenium complexes have been characterized as nitric oxide (NO) donors that induce relaxation in the denuded rat aorta. There are some differences in their vascular relaxation mechanisms compared with sodium nitroprusside. This study investigates whether the endothelium could interfere with the [Ru(terpy)(bdq)NO]3+-TERPY-induced vascular relaxation, by analyzing the maximal relaxation (Emax) and potency (pD2) of TERPY. Vascular reactivity experiments showed that the endothelium negatively modulates (pD2: 6.17 ± 0.07) the TERPY relaxation in intact rat aortic rings compared with the denuded rat aorta (pD2: 6.65 ± 0.07). This effect is abolished by a non-selective NO-synthase (NOS) inhibitor L-NAME (pD2: 6.46 ± 0.10), by the superoxide anion (O2-) scavenger TIRON (pD2: 6.49 ± 0.08), and by an NOS cofactor BH4 (pD2: 6.80 ± 0.10). The selective dye for O2- (DHE) shows that TERPY enhances O2- concentration in isolated endothelial cells (intensity of fluorescence (IF):11258.00 ± 317.75) compared with the basal concentration (IF: 7760.67 ± 381.50), and this enhancement is blocked by L-NAME (IF: 8892.33 ± 1074.41). Similar results were observed in vascular smooth muscle cells (concentration of superoxide after TERPY: 2.63 ± 0.17% and after TERPY + L-NAME: −4.63 ± 0.14%). Considering that TERPY could induce uncoupling NOS, thus producing O2-, we have also investigated the involvement of prostanoids in the negative modulation of the endothelium. The non-selective cyclooxygenase (COX) inhibitor indomethacin and the selective tromboxane (TXA2) receptor antagonist SQ29548 reduce the effect of the endothelium on TERPY relaxation (pD2 INDO: 6.80 ± 0.17 and SQ29548: 6.85 ± 0.15, respectively). However, a selective prostaglandin F2α receptor antagonist (AH6809) does not change the endothelium effect. Moreover, TERPY enhances the concentration of TXA2 stable metabolite (TXB2), but this effect is blocked by L-NAME and TIRON. The present findings indicate that TERPY induces uncoupling of eNOS, enhancing O2- concentration. This enhancement in O2- concentration induces COX activation, producing TXA2, which negatively modulates the rat aorta relaxation induced by the NO donor TERPY.