The Protective Mechanism Of Electroacupuncture Pretreatment On Myocardial Ischemia Reperfusion Injury Based On Inflammatory Response Dynamic Changes

Objectives:The project aimed to observe the myocardial protective effect of electroacupuncture(EA)pretreatment on myocardial ischemia reperfusion injury(MIRI)in rats,and explore the relevant mechanism from the perspective of inflammatory response dynamic changes.Methods:1.Male SD rats were randomly divided into Normal group,Sham group,myocardial ischemia reperfusion injury(MIRI)group,EA pretreatment+myocardial ischemia reperfusion injury(EMIRI)group.In the Sham,MIRI and EMIRI group,rats were further divided into three subgroups:6h,24h and 3d after reperfusion.The model of myocardial ischemia-reperfusion was established by ligating the left anterior descending coronary artery(LAD)for 30min,followed by reperfusion.In the EMIRI group,the rats were received EA on Neiguan before modeling,and the parameters of EA were 2/100Hz,2mA,20min/time,once a day,for consecutive 4 days.The myocardial protective effect of EA preconditioning on MIRI was evaluated by echocardiography,cardiac mass index,myocardial enzymes,myocardial infarct area and HE staining of myocardial tissue.2.To observe the effect of electroacupuncture pretreatment on the mitochondria,plasma free mitochondrial DNA(mtDNA)was detected by real-time quantitative PCR,myocardial mitochondrial cytochrome C(mit-Cyto C)and cytosolic cytochrome C(cyt-Cyto)was detected by Western-blot.3.To observe the effect of electroacupuncture pretreatment on the activation of NLRP3 inflammasome in myocardial tissue,toll-like receptor 9(TLR9),NOD receptor family protein 3(NLPR3),apoptosis-related speckle-like protein(ASC),pro-caspase 1 and caspase 1 was dectected by Western-blot.4.To observe the effect of electroacupuncture pretreatment on the transformation from acute proinflammatory phase to anti-inflammatory repair phase in myocardial tissue,myeloperoxidase(MPO)was detected by real-time quantitative PCR.Immunohistochemistry was used to dectect the expression of interleukin-1?(IL-1?),induced nitric oxide synthase(iNOS),interleukin-10(IL-10),transforming growth factor-?(TGF-?),CD86 and CD206.Results:1.EA pretreatment at Neiguan improved the left ventricular(LV)function(P<0.001),decreased cardiac mass index at 3 days of reperfusion(P<0.05),reduced plasma CK-MB and cTnT levels at each time point of reperfusion(P<0.05),decreased the size of myocardial infarction at each time point after reperfusion(P<0.05),improved the changed myocardial histomorphology caused by MIRI.2.Mitochondrial injury was aggravated and plasma free mtDNA was increased after MIRI(P<0.05).With EA pretreatment,the mitochondrial injury was improved at each time point after reperfusion(P<0.05),and free mtDNA was reduced at 6h and 24h after reperfusion(P<0.05).3.The activation of NLRP3 inflammasome was significantly increased after MIRI(P<0.01).EA pretreatment reduced the activation of NLRP3 inflammasome at each time point after reperfusion(P<0.05).4.After MIRI,the expression of M1 macrophages increased at each time point of reperfusion(P<0.05),the expression of M2 macrophages decreased significantly at 6h and 24h after reperfusion,and the infiltration of neutrophils increased at 24h after reperfusion;In the MIRI group,increased M1 macrophage(P<0.01),decreased M2 macrophage(P<0.001)and increased neutrophils infiltration(P=0.05)appeared at 24h after reperfusion compared with 6h after reperfusion,and increased M2 macrophage(P<0.001)and decreased the neutrophils infiltration(P<0.05)appeared at 3d after reperfusion compared with 24h after reperfusion.EA pretreatment reduced the expression of M1 macrophages at 24h and 3d after reperfusion of MIRI(P<0.05),increased the expression of M2 macrophages at 24h after reperfusion(P<0.001)and reduced the neutrophils infiltration at each time point of reperfusion(P<0.05);In the EMIRI group,the expression of M1 and M2 macrophages increased significantly at 24h of reperfusion compared with 6h of reperfusion(P<0.01).Conclusions:1.EA pretreatment at Neiguan could exert the cardioprotective effect after MIRI.2.EA pretreatment could reduce mitochondrial damage,inhibit the excessive activation of NLRP3 inflammasome,and promote the balance between the acute proinflammatory phase and the anti-inflammatory repair phase after MIRI,and finally exerted the cardioprotective effect.