Role Of AMPK Mediated Restoration Of Autophagic Flux In Slow-Releasing H₂S Donor ADT Postcondition Protects Rat Hearts Against Ischemia–reperfusion Injury In Vivo

ObjectiveTo investigate if slow-releasing H2 S donor ADT postcondition protects rat hearts against ischemia-reperfusion injury in vivo by restoring impaired autophagic flux in an AMP-activated protein kinase activation-dependent manner. MethodsTwo hundred and two male rats were established acute myocardial I/R injury model in vivo. All rats were randomly divided to nine groups: Sham group, I/R + ADT group, I/R + vehicle group, Sham + ADT group, I/R + ADT + CC group, I/R + ADT + DMSO group, I/R + ADT + CQ group, I/R + ADT + NS group, I/R + CQ group. All rats except Sham group and Sham + ADT group received left anterior descending coronary artery occlusion followed by 4 hours reperfusion in vivo. ADT(50 mg/kg), the same volume of control vehicle for ADT, CQ(10 mg/kg) and the same volume of normal saline(NS) were administered via intraperitoneal injection while Compound c(250 μg/kg) and the same volume of DMSO were administered via right jugular vein injection. CQ and NS were given 1 h before surgical operation, all the rest of drugs were administered immediately after reperfusion. At the end of reperfusion, infarction was analyzed with TTC histology and hematoxylin-eosin staining. The generation of reactive oxygen species(ROS) was assessed with the oxidant-sensitive fluorogenic probe. AMPK activation and autophagosome accumulation were assessed by analyzing AMPK and S6 ribosomal protein(S6) phosphorylation, microtubule-associated protein 1 light chain 3(LC3), lysosome associated membrane protein-2(LAMP-2), P62/SQSTM1 and beclin-1. ResultsCompared with the Sham and Sham + ADT group, infarct size and the production of ROS were increased, pathological damage was aggravated, the expression of Beclin-1, LC3 II/I and P62 were up-regulated while the expression of LAMP-2 was down-regulated in the other groups(P < 0.05).Compared with the I/R + vehicle group, infarct size and the production of ROS were decreased, pathological damage was alleviated, the expression of p-AMPK/AMPK and LAMP-2 were up-regulated while the expression of p-S6/S6, Beclin-1, LC3 II/I and P62 were down-regulated in the I/R + ADT group(P < 0.05).Compared with the I/R + ADT group, infarct size and the production of ROS were increased, pathological damage was aggravated, the expression of p-AMPK/AMPK and LAMP-2 were down-regulated while the expression of p-S6/S6, Beclin-1, LC3 II/I and P62 were up-regulated in the I/R + ADT + CC group(P < 0.05).Compared with the I/R + ADT group, infarct size was increased, the expression of LAMP-2 was down-regulated while the expression of LC3 II/I and P62 were up-regulated in the I/R + ADT + CQ groups(P < 0.05). ConclusionSlow-releasing H2 S donor ADT postcondition protects rat hearts against myocardial I/R injury in vivo, which may be through increasing blood H2 S concentrations, activating AMP-activated protein kinase, thereby restoring impaired autophagic flux and attenuating ROS generation during reperfusion.