Abstract 2099 Pulmonary: General Lung Biology Poster Symposium, Sunday, 5/2

Introduction: The production of the free radical superoxide in blood vessels has been shown to affect vascular smooth muscle tone through the inactivation of nitric oxide. We explored the vascular effects of superoxide in pulmonary arteries isolated from late gestation fetal and juvenile lambs. Methods: Fifth generation pulmonary arteries were isolated from fetal (138 days, n=5) and 4-6 week old juvenile (n=5) lambs, and studied using conventional tissue bath techniques. Vessels were pretreated with indomethacin and propranolol, and preconstricted with a submaximal concentration of norepinephrine. Concentration response curves were then developed for polyethelene glycol-superoxide dismutase (PEG-SOD, 0.75-75 U/mL), a membrane permeable SOD which catalyzes the dismutation of superoxide anion to hydrogen peroxide. Repeated measures ANOVA was used for statistical comparisons. Results: We found that PEG-SOD significantly relaxed PA from both fetal and juvenile lambs (maximal relaxations 88 ± 6% and 92 ± 2%, respectively). Lowering the oxygen tension supplied to tissue bath from 94% to 3% significantly blunted, but did not eliminate these relaxations. PEG-SOD relaxations were completely blocked by pretreatment with the superoxide scavenger Tiron (10-5 M). Pretreatment with a membrane soluble form of catalase (PEG-catalase, 1200 U/mL) partially inhibited relaxations to PEG-SOD, indicating they were potentially mediated by hydrogen peroxide formation. Further experiments were performed to test the effect to blockade of nitric synthase (L-NA, 10-3 M) and soluble guanylate cyclase (ODQ, 3×10-6 M, a concentration which inhibits relaxations to exogenous NO and NO donors). L-NA completely blocked PEG-SOD relaxations in both fetal and juvenile arteries (p<0.05). However, ODQ pretreatment had no effect on relaxations to PEG-SOD. Preliminary experiments, relaxations were not altered following pretreatment with inhibitors of xanthine oxidase (allopurinol, 10-4 M), or NADPH oxidase(diphenyliodonium, 10-6 M). Conclusions: We conclude that superoxide anion is generated in pulmonary arteries isolated from both fetal and juvenile lambs, and acts as an endogenous vasoconstrictor. The finding that inhibition of soluble guanylate cyclase does not affect these relaxations suggests that the effect of PEG-SOD is independent of the NO/sGC signal pathway, although our data do not rule out the possibility of relaxations mediated by a cGMP independent action of NO or the generation of hydrogen peroxide. The most likely source of superoxide in isolated pulmonary arteries appears to be nitric oxide synthase, an enzyme which is known to synthesize superoxide under specific cellular conditions.

Funded by NIH 54705 and AHA 9740024N