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Mechanisms underlying hypoxic vasodilatation in porcine coronary arteries

Elise Røge Hedegaard

Summary

Background and hypotheses: The major aim was to study mechanisms underlying hypoxia-induced vasodilatation in coronary arteries and identify potential targets for the treatment of coronary artery disease. The overall systemic vascular response to hypoxia is vasodilatation, which seems to be physiologically protective because it increases blood flow to endangered areas. We hypothesized that hypoxia-induced dilatation is affected by endothelial factors, such as endothelin-1 (ET-1), radical oxygen species (ROS), and nitric oxide (NO) as well as by K+ channel opening, force suppression, and by an increased response to adenosine. 

Experimental approach: Functional experiments were performed by mounting porcine left anterior descendent coronary artery segments with or without endothelium in myographs for isometric tension recording. Intracellular Ca2+ concentration ([Ca2+]i) was measured using fura-2AM; the presence of KV7 K+ channels was examined by PCR, sequence analysis, and immunoblotting; ET-1 was measured by an ELISA assay; NO was measured with an NO-sensitive electrode; cyclic AMP was determined by use of an ELISA assay; and phosphorylation of myosin light chain (MLC) and heat shock protein (HSP)20 was determined by immunoblotting.

Key results: We investigated the influence of potassium channels and found that several potassium channels play a role in hypoxia-induced dilatation but the most pronounced effect was observed by inhibition of KV7 channels. As the first research group to do so, we could demonstrate KV7.1, KV7.4, and KV7.5 expression in porcine coronary arteries. We  also found that hypoxia-induced dilatation consists of both a calcium-dependent and a calcium-independent part, because depolarization with K+ prevented reduction in [Ca2+]i, but without abolishing hypoxia-induced dilatation. Some of the KV7-mediated dilatation under severe hypoxia is mediated by hydrogen sulfide (H2S), because we observed that inhibitors of H2S-producing enzymes inhibited hypoxia-induced vasodilatation at 1% O2. We found that endothelin receptor activation reverses while endothelin receptor antagonism markedly enhances porcine coronary hypoxia-induced dilatation independently of the endothelium. Free tissue ET-1 concentration in the arterial wall was unchanged in 1% O2 versus 95% O2 and independent of the presence of endothelium. NO release was augmented during hypoxia, and an inhibitor of NO synthase, nitro-L-arginine (L-NOARG), inhibited hypoxia-induced dilatation, suggesting that NO contributes to hypoxia-induced dilatation. Furthermore, it was found that hypoxia induces vasodilatation at least in part by force suppression caused by increased cAMP, which leads to phosphorylation of HSP20. Increased adenosine receptor sensitivity under hypoxia compared to normoxia was observed; the results may suggest that adenosine causes dilatation by additional, unknown mechanisms besides the adenosine receptors. 

Conclusion: Potassium channels particularly KV7.4 and/or KV7.5 channels appear to play a major role in hypoxia-induced vasodilatation in coronary arteries, and they may be a potential target for treatment of hemodynamically significant coronary artery disease. We also found that ET-1 counteracts hypoxia-induced vasodilatation, possibly through inhibition of potassium channels. NO mediates part of the hypoxia-induced vasodilatation,  possibly through opening of potassium channels, whiled force suppression is mediated by phosphorylation of HSP20.