Endocrine pharmacologyAcute restraint stress increases carotid reactivity in type-I diabetic rats by enhancing Nox4/NADPH oxidase functionality

Hyperglycemia increases the generation of reactive oxygen species and affects systems that regulate the vascular tone including renin-angiotensin system. Stress could exacerbate intracellular oxidative stress during Diabetes upon the activation of angiotensin AT1/NADPH oxidase pathway, which contributes to the development of diabetic cardiovascular complications. For this study, type-I Diabetes was induced in Wistar rats by intraperitoneal injection of streptozotocin. 28 days after streptozotocin injection, the animals underwent to acute restraint stress for 3 h. Cumulative concentration-response curves for angiotensin II were obtained in carotid rings pre-treated or not with Nox or cyclooxygenase inhibitors. Nox1 or Nox4 expression and activity were assessed by Western blotting and lucigenin chemiluminescence, respectively. The role of Nox1 and Nox4 on reactive oxygen species generation was evaluated by flow cytometry and Amplex Red assays. Cyclooxygenases expression was assessed by real-time polymerase chain reaction. The contractile response evoked by angiotensin II was increased in diabetic rat carotid. Acute restraint stress increased this response in this vessel by mechanisms mediated by Nox4, whose local expression and activity in generating hydrogen peroxide are increased. The contractile hyperreactivity to angiotensin II in stressed diabetic rat carotid is also mediated by metabolites derived from cyclooxygenase-2, whose local expression is increased. Taken together, our findings suggest that acute restraint stress exacerbates the contractile hyperreactivity to angiotensin II in diabetic rat carotid by enhancing Nox4-driven generation of hydrogen peroxide, which evokes contractile tone by cyclooxygenases-dependent mechanisms. Finally, these findings highlight the harmful role played by acute stress in modulating diabetic vascular complications.

Graphical abstractNox1/NADPH oxidase activity in carotid arteries from diabetic rats is enhanced; angiotensin II increases the generation of superoxide (O2−) and hydrogen peroxide (H2O2), which contribute to angiotensin II-induced contraction by activating the synthesis of contractile metabolites from cyclooxygenases (COX) (A). Acute restraint stress enhances the activity of Nox4/NADPH oxidase during type-1 Diabetes; Nox4 generates high levels of H2O2, which modulates the production of COX metabolites (B). Dashed arrow: activity; continuous arrow: generation; FAD, flavin-adenine dinucleotide; SOD, superoxide dismutase; O2− superoxide anion; H2O2, hydrogen peroxide; VSCM, vascular smooth muscle cell; AngII, angiotensin II; TXA2, thromboxane A2; PGH2, prostaglandin H2.Download high-res image (184KB)Download full-size image