Abstract 375: Stimulation Of cGMP Signaling Protects Reperfused Myocardium By Reducing Reperfusion Induced Barrier Failure Of The Coronary Endothelium
Ischemia-reperfusion (IR) provokes barrier failure of the coronary endothelium, leading to myocardial edema development that jeopardizes functional recovery of the heart. We showed that reperfusion induced barrier failure is due to a Ca2+ dependent activation of the endothelial contractile machinery which in turn leads to opening of intercellular gaps. The aim of the present study was to investigate whether stimulation of cGMP-signaling by activation of soluble guanylyl cyclase (sGC) with DEAnonoate or HMR1766, an activator of heme-oxidized sGC, can reduce reperfusion induced endothelial barrier failure and protect the heart against an imminent reperfusion injury and if this is due to an influence on cytosolic Ca2+ homeostasis during reperfusion. In cultured rat coronary endothelial monolayers stimulation of cGMP-signaling during reperfusion markedly reduced gap-formation (gaps [%]; normoxia:0±6; IR:205±38*; DEA:111±35**; HMR:57±23**; n>8; *p<0.05 vs.normoxia; **p<0.05 vs. IR) and attenuated the reperfusion induced increase in cytosolic Ca2+ (fura-2 ratio [a.u.]; normoxia:0.90±0.02; IR:1.12±0.02*; DEA:0.99±0.03**; HMR:0.95±0.03**; n>8; *p<0.05 vs. normoxia; **p<0.05 vs. IR). We found that reduction of cytosolic Ca2+ overload is due to an augmented Ca2+ sequestration into the endoplasmic reticulum, since thapsigargin releasable Ca2+ (n>5; p<0.05) and SERCA activity (n>5; p<0.05) were increased. Furthermore the decrease in cytosolic Ca2+ lead to decreased MLC-phosphorylation (n>5; p<0.05). In isolated saline perfused rat hearts, reperfusion induced increase of myocardial water content was significantly reduced (H2O/dry weight [ml/g]; normoxia:5.98±0.47; IR:7.90±0.23*; DEA:6.71±0.35**; HMR:6.94±0.34**; n>5; *<0.05 vs. normoxia; **p<0.05 vs. IR). In conclusion, activation of the cGMP signaling pathway effectively protects the coronary endothelium against reperfusion-induced barrier failure, in cultured coronary endothelium as well as in the coronary system in situ. This cGMP-mediated protection is due to reduction of cytosolic Ca2+ in endothelial cells and decreased activation of the endothelial contractile machinery, resulting in a stabilization of the endothelial barrier function during reperfusion.