Sestrin2/adenosine monophosphate activated protein kinase regulated autophagy pathway in myocardial ischemia-reperfusion injury in diabetic rats and the protective effect of tert-butyl hydroquinone

Abstract

Objective To investigate Sestrin2/adenosine monophosphate-activated protein kinase (AMPK) induced autophagy in myocardial ischemia-reperfusion injury in diabetic rats and the protective effect of tert-butyl hydroquinone (TBHQ). Methods Healthy adult male SD rats were randomly divided into 6 groups (n=10): control rats with sham surgery group (NS), control rats with ischemia-reperfusion (NIR) group, control rats with ischemia-reperfusion+TBHQ group (NIPC), diabetes rats with sham surgery group (DS), diabetic rats with ischemia-reperfusion(DIR) group, diabetic rats with ischemia-reperfusion+TBHQ group(DIPC). Diabetes mellitus was induced by intraperitoneal injection of 60 mg/kg streptozotocin. TBHQ (40 mg/kg·d) was injected intraperitonealy to rats in TBHQ groups for 5 days before ischemia-reperfusion surgery. The myocardial ischemia-reperfusion model was induced by temporary ligation of left anterior descending branch of coronary artery for 30 min followed by 120 min reperfusion. The change of myocardial tissue were observed by hematoxylin-eosin staining. The activities of serum creatine kinase isoenzyme (CK-MB) , lactate dehydrogenase (LDH) and superoxide dismutase (SOD) were determined by enzyme linked immunosorbent assay. The expression of Sestrin2, phosphorylated AMP activated protein kinase (p-AMPK) , Beclin1 and cysteine aspartic acid specific protease 3 (Caspase3) were detected by Western blotting. T-test or ANOVA analysis was used for data analyses. Results Compared with group NIR, the activities of CK-MB and LDH were decreased(343±37 vs 453±36, 428±65 vs 576±45, t=0.006 6,0.013 4, all P<0.05), whereas the level of SOD was increased in group NIPC (706±17 vs 566±21, t<0.000 1, P<0.05). Sestrin2, p-AMPK, Beclin1 level were statistically higher and Caspase 3 was lower in NIPC group (t=0.000 4-0.005 6, P<0.05) compared with NIR group (t=0.000 9, P<0.05). Compared with group NS, the activities of CK-MB, LDH, SOD were highly expressed in group NIR and group NIPC (F=17.04-413.16, all P<0.05). Compared with group DS, the activities of CK-MB, LDH, SOD were highly expressed in group DIR and group DIPC (F=91.83-183.28, P<0.05), whereas Sestrin2, p-AMPK, Beclin1, Caspase3 were increased in groups NIR and NIPC (F=8.63-128.69, P<0.05). Compared with group DIR, the activities of CK-MB, LDH, Caspase3 were decreased in group DIPC (t=0.008 0-0.022 7, all P<0.05), but SOD, Sestrin2, p-AMPK, Beclin1 were increased in group DIPC (t=0.000 3-0.006 9, all P<0.05). Conclusion TBHQ alleviates myocardial ischemia-reperfusion injury by up-regulating Sestrin2/AMPK pathway which then activates authophagy in mitochondria. Key words: Diabetes mellitus; Myocardial ischemia-reperfusion injury; Tert-butyl hydroquinone; Sestrin2; Adenosine monophosphate-activated protein kinase