Experimental Study Of Delayed Cardioprotection Of Sufentanil And Its Relations To COX-2 And Mitochondrial K_ATP Channels

Sufentanil is one of potent phenylpiperidine opioids most frequently used for anesthesia in the patients receiving cardiovascular surgery.It still remains unknown whether sufentanil can provide delayed cardioprotection against ischemia and reperfusion injury.As important mediators of cardioprotection induced by ischemic preconditioning and opioids,role of cyclooxygenase-2(COX-2) and mitochondrial adenosine triphosphate-sensitive potassium channels(mitoK_ATP channels) in the myocardial ischemic reperfusion process is being a hotspot subject studied.We therefore conducted the present study to investigate the delayed cadioprotection of sufentanil and the role of COX-2 and mitoK_ATP channels as well as the relations between them in mediating sufentanil-induced delayed cardioprotection.Our purpose was to further clarity the mechanism of late preconditioning.This study consists of the following two parts:Part One:Study of Delayed Cardioprotection of SufentanilAdult male Wistar rats weighing 250-300g were randomly divided into 7 groups.All the rats were pretreated with sufentanil or equal volume of saline 24 h before 45-min of left anterior descending coronary artery occlusion followed by 120-min of reperfusion.The detailed experimental protocols were as following:In group 1,rats were treated with normal saline 1 ml/kg intraperitoneally(I.P.);In groups 2,3,4 and 5,rats were treated with different doses of sufentanil(1,5,10,20μg/kg) I.P.,respectively;In groups 6 and 7,rats were treated according to the same experimental protocols as in groups 5 and 1 except receiving additional nonselective opioid receptor antagonist naloxone 1 mg/kg I.P.15 min before preconditioning with sufentanil 20μg/kg or saline,respectively.Heart rate(HR),mean arterial pressure(MAP),and a leadâ…¡electrocardiogram were continuously monitored during ischemia-reperfusion process.To determine plasma concentration of CK-MB, arterial blood sample was obtained immediately before ischemia,and at the end of ischemia and reperfusion.After a reperfusion period of 120 min,heart was removed for myocardial infarct size measurement.Infarct size was expressed as a percentage of the area at risk.The results showed no significant differences among all groups in the baselines of HR,MAP,RPP(rate-pressure product) and plasma concentrations of CK-MB before ischemia(P＞0.05).There were also no significant differences among all groups in HR,MAP and RPP at every measurement point during ischemia-reperfusion(P＞0.05).As compared to group 1,plasma concentration of CK-MB was significantly lower in group 3 at the end of ischemia and in group 4 and 5 at the end of ischemia and reperfusion(P＜0.05),and myocardial infarct size was smaller significantly in groups 3,4 and 5(P＜0.05).The plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were not different between groups 1 and 7(P＞0.05).The plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were significantly less in group 3,4 and 5 than in group 2(P＜0.05), and were also significantly less in group 4 and 5 than in group 3(P＜0.05). There were no significant differences in the plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size between groups 4 and group 5(P＞0.05).The plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were greater significantly in group 6 than in group 5(P＜0.05). Part Two:Roles of COX-2 and Mitochondrial K_ATP channels in Sufentanil-induced Delayed CardioprotectionAdult male Wistar rats weighing 200-300g were randomly devided into 6 groups.All the rats were pretreated with sufentanil or equal volume of saline 24 h before 45-min of left anterior descending coronary artery occlusion followed by 120-min of reperfusion.The detailed experimental protocols were as following:In group 1,rats were treated with normal saline 1 ml/kg I.P.; In group 2,rats were treated with sufentanil 20μg/kg I.P.;In groups 3 and 5,rats were treated according to the same experimental protocols as in groups 2 and 1,respectively except receiving additional selective COX-2 inhibitor NS-398 5 mg/kg I.P.30 min before left anterior descending coronary artery occlusion.In groups 4 and 6,rats were treated according to the same experimental protocols as in groups 2 and 1,respectively except receiving additional selective mitochondrial K_ATP channels blocker 5-HD (5-hydroxydecanoate) 10 mg/kg intravenously 10 min before left anterior descending coronary artery occlusion.HR,MAP,and a leadâ…¡electrocardiogram were continuously monitored during ischemia-reperfusion process.To determine plasma concentration of CK-MB,arterial blood samples were obtained immediately before ischemia,and at the end of ischemia and reperfusion.After a reperfusion period of 120 min,heart was removed for myocardial infarct size measurement. Infarct size was expressed as percentage of the area at risk.For measurement of the myocardial levels of prostaglandin(PG)E₂,6-keto-PGF_1α and COX-2 protein expression,myocardial tissue samples from each group were collected without myocardial ischemia and reperfusion in parallel series of experiments. Expression of COX-2 was assessed by Western blotting,and myocardial levels of PGE₂ and 6-keto-PGF_1α were measured using enzyme immunoassays.The results showed no significant differences among all groups in HR, MAP and RPP at every measurement point during the experiment(P＞0.05).Plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were significantly less in group 2 than in group 1(P＜0.05).Plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were not different among groups 1,5 and 6(P＞0.05).As compared to group 2,plasma concentrations of CK-MB at the end of ischemia and reperfusion and myocardial infarct size were significantly greater in groups 3 and 4 (P＜0.05).As compared to group 1,the myocardial COX-2 expression in groups 2,3 and 4,and myocardial contents of PGE₂ and 6-keto-PGF_1α in groups 2 and 4 were significantly greater(P＜0.05).The myocardial COX-2 expression and contents of PGE₂ and 6-keto-PGF_1α in groups 1,5 and 6 were comparable(P＞0.05). There was no significant difference in the myocardial COX-2 expression among groups 2,3 and 4.The myocardial contents of PGE₂ and 6-keto-PGF_1α in group 3 but not in group 4 were less significantly in comparison with group 2(P＜0.05).On the basis of the results from our experiments,the following conclusions can be drawn:1.Pretreatment with sufentanil at doses of 1,5,10 and 20μg/kg in intact rat induces a dose-related delayed cardioprotection,which nearly reachs the peak effect at dose of 10μg/kg.Sufentanil triggers the delayed cardioprotection via opioid receptors.2.Both COX-2 and mitoK_ATP channels play obligatory roles in mediating sufentanil-induced delayed cardioprotection.COX-2 locates upstream of mitoK_ATP channels in the singaling transduction of sufentanil-induced delayed cardioprotection.