| Background: ST-Elevation Myocardial Infarction(STEMI)throw immense health and economic burdens globally.Rapid reperfusion of the ischemic myocardium is the standard treatment for acute myocardial infarction(AMI)to rescue the jeopardized myocardium and reduce infarction size(IS).Paradoxically,however,reperfusion itself may abrogates myocardial salvage and can induce further injury to the ischemic myocardium by complex mechanisms termed as ischemia-reperfusion injury(IRI).Ischemic postconditioning(IPost C)defined by Zhao et al in 2003 conferred cardioprotection against IRI in dogs.However,ischemic postconditioning was negative in recent large randomized trials,including the largest DANAMI-3-i POST trial to date and the latest meta-analysis.In 1993,Przyklenk first proposed the concept of remote ischemic adaptation.Remote ischemic conditioning can be induced during(percondition),or follow(postcondition)the sustained coronary occlusion that causes the infarction.However,a neutral clinical study by Verouhis et al(RECOND trial)has been published recently,in which up to seven cycles of lower limb RIC with at least one cycle initiated prior to reperfusion failed to reduce MI size as a percentage of the AAR in anterior STEMI patients.The RIPOST-MI study of RIPer C and IPOS combined interventions did not lead to further reduction of infarct size.The LIPSIA CONDITIONING study in the hospital,RIPer C+IPost C significantly improved the myocardial salvage index,and reported superior to a single intervention.In view of the recent reports of IPost C-negative reports in clinical studies,we turn our attention to RIC,especially the combination of RIPer C and RIPost C to be fully elucidated.Wei M et al used a combination of RIPer C and RIPost C in an in vivo rat myocardial infarction model to provide no further protection.However,the animal experiments RIPer C and RIPost C are closely linked,and the interval between RIPer C and RIPost C is zero,similar to a cycle number Enhanced remote ischemic condition,Although the number of optimal ischemic postconditioning has not been determined,over-adaptation to "hyperconditioning" may be detrimental.RIPer C and RIPost C have a certain time interval could meet the clinical reality.Therefore,it is necessary to explore the combination ofRIPer C and RIPost C separated by a certain period of time to observe whether the combined application of two types of remote ischemic condition is more beneficial than a single RIC.Cellular autophagy plays a key role in myocardial IRI.In Wei C’s rat IR model,activated autophagy plays a protective role in the myocardium after Post C.Han Z et al studie that in normal mice but not in diabetic(DM)mice,RIPost C may partially reduce IRI by inducing autophagy in cardiomyocytes.However,studies of autophagy involving RIPer C in myocardial IRI have not been reported.Stromal derived factor 1α(SDF-1α)and nitric oxide(NO)may be protective humoral factors after ischemia.There is no research on how RIPer C affects the expression of SDF-1α and NO in STEMI patients.In view of the above research background,we sought to prove: ⑴ Can the combination of RIPer C and RIPost C attenuate IRI and have a protective effect on the heart? Is RIPer C better than RIPost C in combination with a RIC? ⑵ If there is cardioprotection with RIPer C,is this cardioprotective effect of RIPer C associated with autophagy? ⑶ Is SDF-1α and NO one of the protective mechanisms of RIPer C in patients with STEMI?Part Ⅰ Basic research of remote ischemic conditioning on myocardial ischemia reperfusion injury in rabbitsObjective: To study the effect of remote ischemic condition(RIC)on myocardial ischemia-reperfusion injury(IRI)in rabbits.The present study used an in vivo rabbit myocardial ischemia-reperfusion model to assess whether synergistic effects are produced when RIPost C is combined with RIPer C.We also explored whether autophagy is involved in the cardioprotection effect of remote limb ischemic perconditioning on myocardial ischemia reperfusion injury in rabbit.Methods: Male New Zealand white rabbits were randomly divided into 8 groups.1: sham group(Sham group).2: Ischemia-reperfusion model group(IR group)(Model group),myocardial ischemia for 40 minutes,and reperfusion for 3 hours.3: Remote ischemic adaptation group(RIPer C).4: Remote ischemic postcondition group(RIPost C).5: U1 group-RIPer C + RIPost C groups,two groups at intervals of 0 minutes.6: U2 groupRIPer C + RIPost C groups,two groups at intervals of 5 minutes.7: U3 group-RIPer C +RIPost C groups,two groups were separated by 10 minutes.8:3 MA group-3 MA+ RIPer C.RIC’s plan is to apply microvascular blocking to the left femoral artery for 5 minutes,release it for five minutes,and repeat three cycles.After 3 hours of reperfusion,serum troponin I(c Tn I)was measured.The morphological and autophagic activity of the myocardium were observed by electron microscopy.The autophagy inhibitor(3-MA)was used to study the role of autophagy in RIPer C.The expression of six genes,Beclin-1,LC3,P62,Bcl2,Bax,and Caspase 3 in rabbit myocardium,was detected by-time PCR.After four weeks of feeding,echocardiography was performed to evaluate cardiac function,and the left ventricular end-diastolic diameter(LVEDd),left ventricular end-systolic diameter(LVIDs),and left ventricular ejection fraction were measured.LVEF),fraction shortening(FS).Left ventricular global longitudinal systolic peak strain(GLPS)was assessed by speckle tracking strain and myocardial infarct size was determined by staining with triphenyltetrazolium chloride(TTC).Results: 1.There was no difference in body weight between rabbits in each group(P>0.05).The seven groups of vascular ligation had ST elevation in the lead of ECG II after ligating the coronary arteries;all groups showed ST segment fall at 10 minutes after releasing the ligation line,and there was no difference between groups in ST segment fallback(P>0.05).)2.Serum troponin I,compared with Sham group,c Tn I in IR,RIPer C,RIPost C,U1,U2,U3,3-MA+RPer C groups were all increased(P<0.05),compared with IR group,RIPer C,The c Tn I was decreased in RIPost C,U1,U2,and U3 groups(P<0.05).There was no statistical difference in c Tn I between RIPer C,RIPost C,U1,U2,and U3 groups.The 3-MA+RPer C group had higher c Tn I than the RIPer C,RIPost C,U1,U2,and U3 groups.3.Effect of RIC on myocardial infarct size Myocardium was stained with TTC staining.The Sham group had no infarction due to TTC staining and no statistics were obtained.Infarct size(IS/LV)was 33.72±9.96% in IR group,21.37±11.11% in RIPer C group,24.75±6.50% in RIPost C group,20.75±6.63% in U1 group,24.00±9.31 in U2 group,19.58±7.23% in the U3 group and 38.86±8.66 in the 3-MA+RPer C group(P<0.05).Compared with IR group,the infarct size of RIPer C,RIPost C,U1,U2,and U3 groups were all decreased(P<0.05).There was no statistical difference in myocardial infarct size in 3-MA+RPer C group.There was no statistical difference in infarct size between RIPer C,RIPost C,U1,U2,and U3 groups.4.Effect of RIC on Left Ventricular Function The left ventricular overall longitudinal systolic peak strain(GLPS)results(negative values)in each group were: Sham group,21.5125±2.7378;IR group,10.4667±1.5338;RIPer C group,10.6222±3.9493;RIPost C group,11.1444±3.2269;U1 group,13.9889±2.4882;U2 Group 13.6567±3.1008;U3 group 12.6667±2.0304;3-MA+RPer C group 12.4444±2.5274(P<0.05).Compared with Sham group,the absolute values of GLPS in IR,RIPer C,RIPost C,U1,U2,U3,and 3-MA+RPer C groups all decreased(P<0.05);compared with IR group,the values of GLPS in U1 and U2 groups increased.There was no statistical difference between RIPer C,RIPost C,U3,3-MA+RPer C groups.Left ventricular ejection fraction(EF)results in each group were 63.5000±4.3753 in Sham group,44.5556±3.5746 in IR group,49.1111±7.3220 in RIPer C group,49.7778±4.2361 in RIPost C group,50.3333±2.9580 in U1 group,53.1111±5.1343 in U2 group,and U3 group 49.6667±4.0927,3-MA+RPer C group 48.6667±3.6742.Compared with Sham group,the EF values of IR,RIPer C,RIPost C,U1,U2,U3,3-MA+RPer C groups decreased(P<0.05);compared with IR group,The EF value in RIPer C,RIPost C,U1,U2,U3 groups increased(P<0.05).There was no statistical difference between the IR group and the 3-MA+RPer C group.Left ventricular short axis shortening rate(FS)results in each group: Compared with IR group,the FS values of RIPer C,RIPost C,U1,U2,and U3 groups increased(P<0.05),no statistical difference between IR group and 3-MA+RPer C group.Left ventricular end-diastolic diameter(LVEDd)results in each group: Compared with Sham group,there was no statistical difference between IR,RIPer C,RIPost C,U1,U2,U3,and 3-MA+RPer C groups;End of ventricular end-systolic diameter(LVIDs)results: compared with Sham group,RIPer C,RIPost C,U2,3-MA+RPer C group LVIDs value increased(P <0.05);Compared with IR group,U3 group LVIDs value increases,there are statistics Differences in learning(P<0.05).5.The relative contents of six genes,Beclin-1,LC3,P62,Bcl2,Bax,and Caspase 3,were detected by Real-time PCR.LC3 m RNA Compared with Sham group,RIPer C,RIPost C,U1,U2,U3,and 3-MA+RPer C groups all increased(P<0.05),but IR group had no significant difference(P<0.05).Compared with IR group,RIPer C,RIPost C,U1,U2,U3 and 3-MA+RPer C groups all increased(P<0.05).Compared with 3-MA+RPer C group,RIPer C,RIPost C,U1,U2,andU3 were all higher(P<0.05).Beclin-1 m RNA Compared with Sham group,the RIPer C,RIPost C,U1,U2,and U3 groups all increased(P<0.05).There was no statistical difference between IR and 3-MA+RPer C groups(p>0.05).Compared with IR group,RIPer C,RIPost C,U1,U2,and U3 groups all increased(P<0.05),and 3-MA+RPer C group had no statistical difference(p>0.05).There was no statistical difference between the RIPer C,RIPost C,U1,U2,and U3 groups(p>0.05).P62 m RNA Compared with Sham group,IR,RIPer C,RIPost C,U1,U2,U3,3-MA+RPer C groups were all decreased(P<0.05);Compared with IR group,there was no statistical difference between RIPost C,U1,U2,and U3 groups.(p>0.05).Bcl2/Bax Compared with Sham group,RIPer C,RIPost C,U1,U2,and U3 groups all increased(P<0.05);IR,3-MA+RPer C group had no statistical difference(p>0.05).Caspase3 m RNA Compared with Sham group,IR,RIPer C,RIPost C,U1,U2,U3,3-MA+RPer C groups all increased(P<0.05);compared with IR group,RIPer C,RIPost C,U1,U2,3-MA +RPer C group decreased(P<0.05),U3 had no statistical difference(p>0.05).There was no statistical difference between the RIPer C,RIPost C,U1,U2,and U3 groups(p>0.05).6.Transmission electron microscopy to detect autophagic body results In IR group,myocardial fibers were arranged neatly,mitochondria showed edema,mitochondrial membranes were intact,and the number of autophagosomes was significantly increased compared with Sham group.The number of autophagosomes per 1000 μm 2 was 39.67±4.93;there were multiple in RIPer C group.Double-layered membranes encapsulated the phagosomes of shrinking mitochondria,and the number of autophagosomes per 1000 μm 2 was 90.67±12.06;in the RIPost C,U1,U2,and U3 groups,mitochondria showed edema,autophagic bodies The number of autophagosomes per 1000 μm 2 were 106.7±16.26,94±17.09,106.3±26.5,and 97.67±7.02,respectively.There was no significant difference between the groups,and the statistical results showed that The number of autophagosomes was significantly different between RIPer C,RIPost C,U1,U2,and U3 groups compared with IR(P<0.01).In the 3MA group,mitochondria were swollenand mitochondria were clearly visible and the number of autophagosomes per 1000μm2 was 57±7.The number of autophagosomes in the 3MA group was significantly higher than those of RIPer C,RIPost C,U1,U2,and U3 groups.Conclusion 1.In rabbit models of myocardial ischemia and reperfusion,RIPer C and RIPost C could all reduce the infarct size,reduce IRI,and protect the heart.Whereas RIPer C and RIPost C were equally effective in providing protection against myocardial IR injury,the combination of RIPer C and RIPost C failed to provide further protection than treatment with either alone.2.In myocardial IR model in rabbits,RIPer C induced autophagy,inhibited apoptosis,and activated autophagy protected the myocardium during adaptation to remote ischemia.Part Ⅱ The effect of remote ischemic perconditioning induced by limb in patients with ST-Segment Elevation Myocardial InfarctionObjective: To explore the effect of remote ischemic perconditioning(RIPer C)on ischemic-reperfusion injury(IRI)in patients with ST-segment elevation myocardial infarction(STEMI).Methods: From January,2016 to June,2017,80 consecutive patients with STEMI were randomly to receive primary percutaneous coronary intervention(PPCI)without RIPer C(PPCI group,n=38)or PPCI plus RIPer C(PPCI+ RIPer C group,n=40).RIPer C consisted of 4 cycles of 5 min/5 min ischemia/reperfusion by cuff inflation/deflation of the left low limb.Protocol was started fefore the first balloon inflation during PPCI operation.Peripheral venous blood sample in all patients were collected before PPCI operation and 0.5、8、24、48、72 hours after PPCI to detect creatine kinase-MB(CK-MB),stromal cell-derived factor-1alpha(SDF-1α)and nitric oxide(NO)concentration in serum.The transthoracic echocardiography was performed on all patients 7 days after operation to assess left ventricular ejection fraction(LVEF).Results: 1.There were no significant differences in baseline characteristics: sex,age,risk factors,vital signs on admission,ischemic time,coronary lesions,contrast agent dosage,stent implantation,between the two groups.There were no statistically significant differences in risk factors,vital signs on admission,ischemic time,coronary lesions,contrast agent dosage,stent implantation,etc.2.The peak CK-MB serum concentration in PPCI+RIPer C was significantly lower compare to PPCI group(334.13±42.87 ng/ml VS 387.21±64.12 ng/ml,P<0.01)The median CK-MB area under the curve(AUC)over 72 h of PPCI+RIPer C group was significantly less then PPCI group(728.36 [365.63-931.98] VS 817.00[683.30-1126.00],P=0.02).3.The median serum concentration of SDF-1α AUC was more markble increased in PPCI+RIPer C group compare to PPCI group(421.800 [304.200-619.000] VS 357.000 [258.600-479.800],P=0.044).4.The median serum concentration of NO AUC was more markble increased in PPCI+RIPer C group compare to PPCI group(5.726 [3.924-13.173] VS 4.836 [3.297-6.142],P=0.035).5.LVEF assessed by transthoracic echocardiography.The LVEF in PPCI+RIPer C and PPCI were 51.56±6.57% and 49.50±6.38%,respectively(P=0.164).There was no significant difference between the two groups.Conclusion:: RIPer C on left low limb in patients with STEMI could attenuates CK-MB release and reduce myocardial enzymatic infarct size,in the meantime increase serum concentration of NO and SDF-1α,suggesting that RIPer C may have protective role on myocardial ischemic-reperfusion injury in patients with STEMI.SDF-1a and NO may be protective factors involved in RIPer C.Animal experiments and clinical studies concluded: The combination of RIPer C and RIPost C did not produce a synergistic effect,RIPer C induced autophagy,and activated autophagy protected the myocardium during RIPer C.Elevated SDF-1a and NO may be involved in the protection of RIPer C in patients with STEMI. |
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