Abstract 014: Nox1-derived ROS Impairs Internal Pudendal Artery Function via Nrf2 and Rho Kinase in Diabetic Mice

Abstract

Oxidative stress plays an important role in vascular dysfunction in diabetes, an important risk factor for erectile dysfunction (ED). Functional and structural changes in internal pudendal arteries (IPAs), which provide blood supply to the corpus cavernosum, can lead to ED. We hypothesized that downregulation of Nrf2-regulated enzymes, consequent to increased NOX1-derived ROS, impairs IPAs function in diabetic mice. IPAs and cultured vascular smooth muscle cells (VSMC) from C57BL/6 (control) and NOX1 knockout (KO) mice were used; diabetes (DM) was induced by streptozotocin in C57BL/6 mice. Vascular function assessment, by wire myography, demonstrated that IPAs from diabetic mice displayed increased contractility to phenylephrine (Phe, Emax: control 138.5±9.5 vs. DM 191.8±15.5) and decreased endothelial-dependent relaxation to acetylcholine (ACh, Emax: control 98.3±2.65 vs. DM 76.1±4.5). These responses were corrected by incubation of IPAs with tiron (ROS scavenger), bardoxolone (Nrf2 activator), ML171 (NOX1 inhibitor) and Y27632 (Rho Kinase inhibitor). IPAs from diabetic mice exhibited decreased Y276732-induced vasodilatation (LogEC50: control 5.8±0.1 vs. DM 5.3±0.1). Cultured VSMC from IPAs, maintained in high glucose (HG) medium, displayed increased levels of superoxide anion (59.6%±10.43) and nitrotyrosine (40.8%±9.8), as well as decreased NO production (37.7%±10.42) and nuclear accumulation of Nrf2 (20.42%±3.6), effects not observed in VSMC from IPAs isolated from NOX1 KO mice. The expression of Nrf2-regulated genes was also decreased in VSMC from IPAs maintained in HG [catalase (25.6%±0.05), HO-1 (21%±0.1) and NQO-1 (22%±0.1)], an effect prevented by the incubation with ML171. HG decreased H2O2 levels in control VSMC (29%±5.0), but increased H2O2 in NOX1 KO VSMC (45.8%±7.1), effects not observed when cells were pre-incubated with GKT 137831 (NOX1/4 inhibitor). In conclusion, diabetes-associated IPAs dysfunction involves increased NOX-1 derived ROS, decreased expression of Nrf2-regulated enzymes and activation of the Rho kinase pathway. These data suggest that Nrf2 may be vasoprotective in diabetes-associated ED. Financial Support: FAPESP, Brazil.