Ten Days of Aerobic Exercise Enhances Aortic Endothelium Dependent Relaxation through Depressed NADPH-oxidase Activity

Abstract

Tetrahydrobiopterin (BH4) is an essential co-factor of endothelial nitric oxide synthase (eNOS). BH4 can be depleted by oxidation, which leads to "uncoupling" of eNOS, where eNOS produces superoxide (O2-) rather than nitric oxide. In addition, enhanced O2- production by NADPH oxidase in the vasculature may create an oxidative environment that promotes eNOS uncoupling, thereby impairing endothelial function. PURPOSE: The purpose of this study was to investigate if 10 days of aerobic exercise can enhance aortic endothelium dependent relaxation (EDR) by BH4 preservation and/or alteration of NADPH oxidase activity. METHODS: Young, female Sprague-Dawley rats were assigned to either an exercise (Ex) or sedentary (Sed) group. Ex rats performed 10 consecutive days of exercise for 15/30/15 min on a treadmill at speeds of 15/30/15 m/min, respectively (6% grade). Rats were sacrificed 24 hrs following the final exercise bout and the aorta was excised and cut into ∼2 mm rings. Individual rings were mounted between 2 pins on a myograph system and bathed with physiological saline. EDR was assessed in rings pre-constricted with 0.3 μM phenylephrine (PE) by cumulative doses of acetylcholine (ACh) in the 0.001 - 10.0 μM range to generate a concentration-dependent response. The experiment was then repeated in each ring following a 30 min incubation in either 10.0 μM BH4 (Ex: n = 9; Sed: n = 2) or 1.0 mM apocynin (Ex: n = 4; Sed: n = 2), an NADPH oxidase inhibitor. The relaxation response was normalized to surface area and expressed as % of PE induced constriction. EDR was assessed as the EC50 value of the ACh dose-response curve. RESULTS: Exercise decreased the EC50 17% (0.171 ± 0.044 vs. 0.206 ± 0.051 μM; Ex vs. Sed). BH4 increased the EC50 35% in Ex (0.232 ± 0.047 vs. 0.171 ± 0.044 μM) and 25% in Sed (0.257 ± 0.074 vs. 0.206 ± 0.051 μM). Apocynin decreased the EC50 37% in Ex (0.158 ± 0.047 vs. 0.244 ± 0.085 μM) and 72% in Sed (0.041 ± 0.020 vs. 0.147 ± 0.039 μM). CONCLUSION: There is a trend toward enhanced EDR following 10 days of exercise. The trend of a greater improvement in ACh-sensitivity upon apocynin incubation in the Sed rings suggests that exercise training increases endothelium dependent relaxation at least partially via a suppression of NADPH oxidase activity. BH4 did not improve relaxation demonstrating a lack of detectable eNOS uncoupling.