AbstractŽ y . Objective: Previous studies suggest a role of superoxide anion radicals PO in impaired endothelium-dependent relaxation of 2 diabetic blood vessels; however, the role of secondary reactive oxygen species remains unclear. In the present study, we investigated a role of various potential reactive oxygen species in diabetic endothelial dysfunction. Methods: Thoracic aortic rings from 8-week streptozotocin-induced diabetic and age-matched control rats were mounted in isolated tissue baths. Endothelium-dependent relaxation to Ž . Ž . acetylcholine ACH and endothelium-independent relaxation to nitroglycerin NTG were assessed in precontracted rings. Results: ACH-induced relaxation was impaired in diabetic compared to control rings and was not improved with either indomethacin or daltroban. ACH-induced relaxation in both control and diabetic rings was completely blocked with the nitric oxide synthase inhibitors, Ž . L -nitroarginine methyl ester or L-nitroarginine L-NA . NTG-induced relaxation was insensitive to L-NA and was unaltered by diabetes.Ž . Pretreatment with superoxide dismutase SOD at activities which did not alter contractile tone failed to alter responses to ACH in diabetic rings. Similar results were obtained using either catalase or mannitol. In contrast, the combination of SOD plus catalase or Ž . DETAPAC, an inhibitor of metal-facilitated hydroxyl radical POH formation, markedly enhanced relaxation to ACH in diabetic but not in control rings. Neither the combination of SOD plus catalase nor DETAPAC altered the sensitivity or relaxation to NTG in control rings with or without endothelium. In diabetic rings with endothelium, both DETAPAC or SOD plus catalase increased sensitivity but not maximum relaxation to NTG. In diabetic rings without endothelium, relaxation and sensitivity to NTG were unaltered by either treatment. In L-NA-treated diabetic rings with endothelium, sensitivity and relaxation to NTG was unaltered by either DETAPAC or SOD plus catalase. Conclusion: Diabetic endothelium produces increases in both PO y and H O leading to enhanced intracellular production of 2 2 2 P OH. Thus, POH are implicated in diabetes-induced endothelial dysfunction.

Oxidative stress is believed to play an important role in the development of vascular complications associated with diabetes mellitus. In this study, we examined the efficacy of long-term treatment with the antioxidant, N-acetylcysteine, in preventing the development of defective endothelium-dependent relaxation in streptozotocin-induced, Sprague-Dawley diabetic rats. At 48 h after injection of streptozotocin, a portion of diabetic rats received 250 mg/L N-acetylcysteine in drinking water for a total duration of 8 weeks. Oral administration did not alter the increase in blood glucose or the reduction in serum insulin but did modestly reduce total glycosylated hemoglobin. In precontracted thoracic aortic rings suspended in isolated tissue baths, endothelium-dependent relaxation to acetylcholine was impaired in diabetic rings compared with control rings. Endothelium-independent relaxation to nitroglycerin was unaltered. Long-term oral administration of N-acetylcysteine did not alter responses to nitroglycerin but completely prevented the defective relaxation to acetylcholine. These studies indicate a dissociation between glycemic control and correction of endothelial dysfunction and suggest that long-term exposure to reactive oxygen subsequent to diabetes rather than hyperglycemia per se is responsible for the development of endothelial dysfunction in diabetes mellitus.

Recent studies have indicated impaired endotheliumdependent relaxation in arteries of insulin-dependent [1] diabetic (IDDM) patients. The mechanisms for this defect in human blood vessels is not yet understood. Several studies have observed defects in endothelium-dependent relaxation in both conduit and resistance arteries of streptozotocin (STZ)-or alloxaninduced diabetic rats or rabbits (see reviews [2,3]).Another experimental diabetic model which is not as frequently investigated is the spontaneously diabetic BB (Bio-Breeding) rat despite the fact that this model more closely approximates IDDM in man. Currently, only a limited number of studies regarding endothelial function have been conducted in this important model [4][5][6][7]. More importantly, there is no available information regarding specific defects in nitric oxide (NO) production contributing to defective endothelium-dependent relaxation in this model. In this study, we evaluated the efficacy of supplementation with l -arginine (l-ARG) to improve NO synthase-dependent, endothelium-dependent relaxation in the diabetic BB rat. Materials and methodsExperiments were performed in compliance with the National Institutes of Health "Principles of Laboratory Animal Care" (NIH publication No. 85-23, revised 1985). Progeny of the original BB rat colony from the University of Pennsylvania were used in breeding pairs to develop a local colony of diabetic and non-diabetic BB rats. Onset of diabetes was regularly monitored using test strips and glucometer and defined by 2 successive days of elevated glucose concentration less than 11 mmol/l. Diabetic animals received s. c. injections of NPH insulin which was varied to maintain a low level of hyperglycaemia (i. e. approximately 12-14 mmol/l). Female diabetic BB rats and age-matched, diabetic-resistant littermates were Diabetologia (1997) 40: 910-915 Reversal by L-arginine of a dysfunctional arginine/nitric oxide pathway in the endothelium of the genetic diabetic BB rat Summary We examined the effects of acute supplementation with arginine in vitro on endothelium-dependent relaxation in aortic rings taken from female genetic, diabetes-prone BB rats. Sensitivity to norepinephrine-induced contraction was unaltered in rings of diabetic BB rats compared to rings from non-diabetic littermates. In precontracted rings, acetylcholine produced a concentration-dependent relaxation which was impaired by diabetes. This relaxation was blocked by l -arginine had no effect on acetylcholine-induced relaxation in control rings. In contrast, relaxation-induced by nitroglycerin in diabetic rings without endothelium was not altered by l -arginine treatment. Thus, a defect in the utilization of arginine by nitric oxide synthase exists in the endothelium of the diabetic BB rat. [Diabetologia (1997) 40: 910-915]