ABSTRACT
Objective To evaluate the role of nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) singling pathway in reduction of myocardial ischemia-reperfusion (I/R) injury by zinc sulfate preconditioning in rats.Methods SPF healthy male Sprague-Dawley rats,weighing 250-280 g,aged 16-20 weeks,were used in this study.After the animals were anesthetized,their hearts were immediately removed and retrogradely perfused with an oxygenated K-H solution at 37 ℃ in a Langendorff apparatus.Forty isolated rat hearts were randomly divided into 4 groups (n=10 each):control group (group C),I/R group,zinc sulfate preconditioning group (group Zn) and zinc sulfate preconditioning plus Nrf2/ARE singling pathway blocker luteolin group (group Zn+Lut).After 20 min of equilibration,the hearts were continuously perfused for 100 min in group C,the hearts were perfused with 4 ℃ St.Thomas′ cardioplegic solution before ischemia and then subjected to 40 min global ischemia followed by 60 min reperfusion to establish the model of I/R in group I/R,200 μmol/L zinc sulfate 1.5 ml/kg was injected intraperitoneally,and 24 h later the model of I/R was established in group Zn,and in group Zn+Lut,the hearts were perfused for 3 min with K-H solution containing luteolin 50 μmol/L starting from the time point immediately before ischemia,and the other treatments were similar to those previously described in group Zn.Heart rate (HR),left ventricular end diabetic pressure (LVEDP),left ventricular developed pressure (LVDP) and the maximum rate of increase of left ventricular pressure (+dp/dtmax) were recorded at the end of equilibration and reperfusion.Coronary effluent was collected at the end of reperfusion to measure the levels of lactate dehydrogenase (LDH) and malondialdehyde (MDA).The expression of heme oxygenase-1 (HO-1),quinone oxidoreductase (NQO1),superoxide dismutase 1 (SOD1) and Nrf2 in myocardial tissues was detected by Western blot.Results Compared with group C,HR,+dp/dtmax and LVDP were significantly decreased,LVEDP was increased,and the levels of LDH and MDA in coronary effluent were increased at the end of reperfusion in I/R and Zn+Lut groups,and the expression of NQO1,HO-1,Nrf2 and SOD1 was up-regulated in I/R,Zn and Zn+Lut groups (P0.05).Conclusion The mechanism by which zinc sulfate preconditioning reduces myocardial I/R injury is related to activation of Nrf2/ARE singling pathway in rats.
ABSTRACT
Objective To evaluate the role of nuclear factor erythroid 2-related factor 2 (Nrf2)/ antioxidant response element (ARE) signaling pathway in reduction of myocardial ischemia-reperfusion (I/R) injury by ischemic preconditioning in the rats.Methods Healthy male Sprague-Dawley rats,weighing 250-300 g,aged 4-6 months,were used in the study.Their hearts were excised,and retrogradely perfused with K-H solution at 37 ℃ in a Langendorff apparatus.Forty isolated hearts were randomly divided into 4 groups (n=10 each) using a random number table:control group (group C),I/R group,ischemic preconditioning group (group IPC),and ischemic preconditioning +Nrf2/ARE signaling pathway blocker luteolin group (group IPC+L).After 20 min of equilibration,the hearts were continuously perfused for 100 min in group C.After 20 min of equilibration,the hearts were subjected to 40 min ischemia at 32 ℃ followed by 60 min of reperfusion in group I/R.In group IPC,ischemic preconditioning was induced by 6 cycles of 10 s ischemia followed by 10 s reperfusion starting from the time point immediately after 20 min of equilibration,and then the hearts were subjected to 40 min ischemia at 32 ℃ followed by 58 min of reperfusion.In group IPC+L,after 20 min of equilibration,the hearts were perfused with K-H solution containing lueolin 50 μmol/L for 3 min before ischemia,and the other treatments were similar to those previously described in group IPC.Left ventricular developed pressure (LVDP),left ventricular end-diastolic pressure (LVDEP),heart rate (HR),and the maximum rate of increase of left ventricular pressure (+dp/dtmax) were recorded at the end of equilibration and reperfusion.At the end of reperfusion,left ventricular myocardial tissues were obtained for examination of the ultrastructure of myocardial cells and for determination of the expression of Nrf2,heme oxygenase-1 (HO-1),quinone oxidoreductase 1 (NQO1),and superoxide dismutase 1 (SOD1) mRNA and protein (by real-time polymerase chain reaction and Western blot,respectively).Results Compared with group C,the HR,+ dp/dtmax and LVDP were significantly decreased,and LVEDP was significantly increased at the end of reperfusion in I/R and IPC+L groups,and the expression of Nrf2,HO-1,NQO1 and SOD1 mRNA and protein was significantly up-regulated in I/R,IPC and IPC+L groups (P＜0.05).Compared with group I/R,the HR,+dp/dtmax and LVDP were significantly increased,and LVEDP was significantly decreased at the end of reperfusion,the expression of Nrf2,HO-1,NQO1 and SOD1 mRNA and protein was significantly up-regulated (P＜0.05),and the pathological changes were significantly attenuated in group IPC,and no significant change was found in the parameters mentioned above in group IPC+L (P＞0.05).Compared with group IPC,the HR,+dp/dt and LVDP were significantly decreased,and LVEDP was significantly increased at the end of reperfusion,and the expression of HO-1,NQO1,SOD1 mRNA and protein was significantly down-regulated (P＜ 0.05),no significant change was found in Nrf2 mRNA and protein expression (P＞0.05),and the pathological changes were significantly aggravated in group IPC + L.Conclusion Ischemic preconditioning reduces myocardial I/R injury through activating Nrf2/ARE signaling pathway in the rats.