Role of Akt/GSK-3β signaling pathway in isoflurane preconditioning-induced inhibition of mPTP opening during myocardial ischemia-reperfusion in rats

Abstract

Objective To evaluate the role of serine-threonine kinase(Akt)/glycogen synthase kinase-3 beta(GSK-3β)signaling pathway in isoflurane preconditioning-induced inhibition of mitochondrial permeability transition pore protein(mPTP)opening during myocardial ischemia-reperfusion(I/R)in rats. Methods Ninety-six male Sprague-Dawley rats, aged 3-4 months, weighing 200-250 g, were randomly divided into 4 groups(n=24 each)using a random number table: control group(group C); I/R group; isoflurane preconditioning group(group IPC); Akt inhibitor MK-2206 group(group MK). Myocardial I/R was induced by occlusion of the anterior descending branch of the left coronary artery for 30 min followed by 2 h of reperfusion.In group IPC, 1.5% isoflurane was inhaled for 30 min followed by 45 min washout, and then the model of myocardial I/R injury was established.In group MK, MK-2206 300 μg/kg(in dimethyl sulfoxide)was injected intraperitoneally at 30 min before isoflurane inhalation.At 2 h of reperfusion, 8 rats were selected and sacrificed, and the hearts were removed for determination of myocardial infarct size.At 2 h of reperfusion, 8 rats were selected, and blood samples were collected from the right internal jugular vein for determination of serum cardiac troponin I(cTnI) concentrations.The rats were then sacrificed, and myocardial specimens were obtained for determination of the expression of phosphorylated GSK-3β(p-GSK-3β)in cytoplasm and mitochondria(by Western blot)and co-expression of p-GSK-3β with adenine nucleotide translocator(ANT), voltage-dependent anion channel or cyclophilin D in myocardial tissues(using co-immunoprecipitation). At 2 h of reperfusion, 8 rats were selected and sacrificed, myocardial cells were obtained, and the opening time of mPTP was determined with a laser scanning confocal microscope. Results Compared with group C, the myocardial infarct size and serum cTnI concentrations were significantly increased, and the expression of p-GSK-3β in cytoplasm and mitochondria was up-regulated in I/R and IPC groups, the co-expression of p-GSK-3β with ANT was significantly down-regulated, and the opening time of mPTP was shortened in group I/R, and the co-expression of p-GSK-3β with ANT was significantly up-regulated, and the opening time of mPTP was prolonged in group IPC(P 0.05). Compared with group IPC, the myocardial infarct size and serum cTnI concentrations were significantly increased, the expression of p-GSK-3β in cytoplasm and mitochondria was up-regulated, the co-expression of p-GSK-3β with ANT was significantly down-regulated, and the opening time of mPTP was shortened in group MK(P<0.05). No co-expression of p-GSK-3β with voltage-dependent anion channel or cyclophilin D was found in myocardial tissues. Conclusion The mechanism by which isoflurane preconditioning inhibits mPTP opening during myocardial ischemia-reperfusion is partially related to activation of Akt/GSK-3β signaling pathway in rats. Key words: Isoflurane; Ischemic preconditioning; Myocardial reperfusion injury; Mitochondrial membrane transport proteins; Protein-serine-threonine kinases; Glycogen synthase kinase 3