Protective effect of limb remote ischemic postconditioning on reperfused heart at different time points and its mechanism of mediating mitochon-drial-dependent apoptosis and necrosis pathways
AIM:To investigate the impact of transient limb remote ischemic postconditioning(RIpostC)on mice with myocardial ischemia/reperfusion(MI/R)at various time points,and to elucidate its protective mechanism associ-ated with mitochondrial-dependent apoptosis and necrosis pathways.METHODS:Male C57BL/6J wild-type mice under-went either sham operation or left coronary artery ischemia for 45 min followed by 24 h of reperfusion.The RIpostC,con-sisting of 3 cycles of ischemia(5 min)/reperfusion(5 min),was performed on the double lower limbs at 0,1,5,10,15,30 or 60 min after myocardial reperfusion.The assessment of myocardial infarction size was conducted using Evans blue and TTC staining,while serum cardiac troponin I levels were measured using an ELESA kit.Apoptosis and necrosis were evaluated through TUNEL and high mobility group box protein 1(HMGB1)immunofluorescence staining.Western blot analysis was employed to determine the expression levels of cyclophilin D(CypD),Bak,Bax,mitochondrial calcium uni-porter(MCU),voltage-dependent anion channel(VADC)and ATP synthase β subunit(ATP5B).RESULTS:In wild-type mice subjected to MI/R,significant decreases in myocardial infarct size were observed with the following treatments:PostC,RIpostC,RIpostC-1 min,RIpostC-5 min,RIpostC-10 min,and RIpostC-15 min.The RIpostC resulted in de-creased TUNEL and HMGB1 positive cardiomyocytes in reperfused myocardial paraffin slides.However,RIpostC-30 min and RIpostC-60 min treatments did not show an apparent reduction in myocardial infarct size.The RIpostC induced cardio-myocyte mitochondrial swelling and down-regulated Bax,Bak and CypD levels during MI/R.The protective mechanism was further verified using peptidylprolyl isomerase F(Ppif)gene(encoding CypD)knockout mice and mitochondrial per-meability transition pore(mPTP)inhibitors.CONCLUSION:The RIpostC bestows cardioprotection even when adminis-tered with a significant delay of 15 min after the initiation of myocardial reperfusion.It significantly reduced cardiomyocyte necrosis and apoptosis by inhibiting the expression of Bax,Bak and CypD during MI/R.
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