Abstract 1134: Crucial Role of Cu/Zn-SOD in the Synthesis of Endothelium-Derived Hyperpolarizing Factor (EDHF) during Reactive Hyperemia in Mouse Mesenteric Microcirculation in Vivo

Abstract

Background: We have recently demonstrated that endothelial Cu/Zn-superoxide dismutase (SOD) plays an important role as an EDHF synthase. In this study, we examined the possible role of Cu/Zn-SOD in the synthesis of EDHF/H 2 O 2 during reactive hyperemia (RH) in mouse mesenteric microcirculation in vivo. Methods: Mesenteric arterioles from wild-type (wild) and Cu/Zn-SOD −/− mice (SOD-KO) (n=5 each, 22– 60 μm) were continuously observed with a pencil-typed intravital microscope (×600 magnification) during acetylcholine (ACh)-induced vasodilatation and RH (reperfusion after 60 sec of mesenteric artery occlusion) under cyclooxygenase blockade (indomethacin) under the following 3 conditions; control, NO synthase inhibitor (L-NMMA) and L-NMMA+catalase (a specific decomposer of H 2 O 2 ). Results: In mesenteric arterioles of wild-type mice, endothelium-dependent relaxations to ACh (34±4%) and vasodilatation during RH (40±5%) were resistant to L-NMMA (ACh 23±7% and RH 20±5%, both P<0.05), but were markedly inhibited by L-NMMA+catalase (ACh, 8±2% and RH, 4±3%, both P<0.01 vs. control). In mesenteric arterioles of SOD-KO mice, L-NMMA markedly inhibited the relaxations to ACh (30±4% to 8±2%, P<0.01) and vasodilatation during RH (40±5% to 5±2%, P<0.01), and the responses were not inhibited by the addition of catalase (ACh, 3±2% and RH, 4±2%, not significant vs. control). In the presence of L-NMMA, RH-induced increase in blood flow was 330±5% of baseline in the wild-type mice, whereas in the SOD-KO mice, the response was significantly reduced to 145±23% (P<0.01). In the mesenteric arterioles of SOD-KO mice, tempol, an SOD mimetic, significantly increased the ACh-induced vasodilation, which effect was abolished by catalase. Vascular H 2 O 2 production in response to ACh in mesenteric arterioles (fluorescent method) was significantly increased in wild-type mice but was markedly impaired in SOD-KO mice. By contrast, vascular NO production was significantly increased in wild-type mice and was unaltered in SOD-KO mice. Vascular level of BH 4 in the aorta was comparable between the two genotypes. Conclusions: These results indicate that endothelial Cu/Zn-SOD plays an important role as an EDHF synthase during RH in mouse mesenteric microcirculation in vivo.