Downregulation of endothelial nitric oxide synthase in rat aorta after prolonged hypoxia in vivo

Abstract

The goal of this study was to determine whether hypoxia alters expression of endothelial nitric oxide synthase (eNOS) in the systemic circulation. Rats breathed either air or 10% oxygen for 12 hours, 48 hours, or 7 days. Thoracic aortas were excised and either mounted in organ bath myographs or frozen in liquid nitrogen for later extraction of protein and RNA. eNOS protein (Western blotting) was decreased (20% of normoxic control) after 12 hours, 48 hours, and 7 days of hypoxia. eNOS mRNA (ribonuclease protection assay) was similarly reduced. Acetylcholine (10-4 mol/L) reversed phenylephrine (10-5 mol/L) preconstricfion by 53.3±5.6% in aortic rings from normoxic rats and 26.1±4.8% in rings from rats exposed to hypoxia for 48 hours (P<0.05), with comparable impairment of relaxation by the calcium ionophore A23187 (10-5 mol/L). Responses to diethylamine nitric oxide and 8-bromo-cGMP were unaffected. Aortic cGMP levels after incubation with acetylcholine (10-6 mol/L) averaged 14.0±1.8 fmol/mg in rings from normoxic rats compared with 8.7±1.0 fmol/mg in rings from hypoxic rats (P<0.05). Similarly, nitrate concentration (by capillary electrophoresis) in the media in which the rings were incubated was reduced in the hypoxic group (5.6±0.23 μmol/L for hypoxic rats and 7.8±0.7 μmol/L for normoxic rats). Impaired endothelial NO release may handicap the vascular responses that defend vital organ function during hypoxia.