| Objective: Cardiovascular disease has always been a major health problem of humans,and ischemic heart disease is the main cause of death.However,intervention methods such as intervention and surgery don't reduce the degree of myocardial ischemia,but may increase myocardial damage,which is known as myocardial ischemia-reperfusion injury.Studies have shown that ischemic postconditioning before ischemic myocardial reperfusion can reduce injury.However,due to clinical complex factors,such as combination with hypercholesterolemia,the cardioprotective effect of ischemic posttreatment can be cancelled.Therefore,finding effective interventions to inhibit myocardial ischemia-reperfusion injury combined with complex clinical factors has become a research direction that everyone pays attention to.The occurrence of myocardial ischemia-reperfusion injury may be related to mechanisms such as excessively open mitochondrial membrane permeability transition pores,excessive activation of endoplasmic reticulum stress,reactivation injury rescue kinase pathway and excessive activation of apoptotic pathway.Lycopene is a family member of carotenoids and a natural antioxidant.Studies have shown that myocardial ischemia and reperfusion damage can be reduced by inhibiting the excessive activation of endoplasmic reticulum stress and blocking the opening of mitochondrial membrane permeability transition holes.In summary,the purpose of this study is to clarify the effect of lycopene on myocardial ischemia-reperfusion injury in ischemia postconditioning hypercholesterolemia rats,and to reveal the possible mechanism for this effect.Methods: Part ?:The research on the effect of lycopene on myocardial ischemiareperfusion injury in ischemia postconditioning hypercholesterolemia rats.1.Establishment and identification of hypercholesterolemia rat model.Ninety rats in the high cholesterol diet group(HCD group)were continuously fed with cholesterol-rich cholesterol feed.After 12 weeks,total cholesterol(TC),high density lipoprotein cholesterol(HDL-C)and The serum value of low-density lipoprotein cholesterol(LDL-C)was compared with the blood lipid results of the two groups to identify the establishment of a rat model of hypercholesterolemia.2.The Langendorff device was used to prepare experimental rat model with heart ischemia-reperfusion injury.The heart of the experimental rat was isolated,and the ischemia-reperfusion group(IR group)was subjected to total heart ischemia for 30 minutes,and then simulated ischemia-reperfusion for 60 minutes;the ischemic posttreatment group(IPoC group)was given for 2 minutes before reperfusion(10s ischemia / 10 s reperfusion × 6 times)ischemic post-treatment,the rest is the same as IR group;lycopene pretreatment group(LP5 group or LP20 group)started 5 days before heart ex vivo,and received 5mg / kg of intraperitoneal injection daily lycopene or 20mg/ kg lycopene solvent,the rest is the same as the IR group;lycopene + ischemic posttreatment group(LP5 + IPoC group)started 5 days before the heart was separated and given a daily intraperitoneal injection of 5mg / kg lycopene,the rest was same as the IPoC group;the 4-phenylbutyric acid sodium pretreatment group(the 4-PBA group)was given an intraperitoneal injection of 20 mg / kg 4-PBA solvent 1 hour before the ischemia,and the rest was the same as the IR group.3.Detection of cardiac function in isolated rat hearts.Monitor and record relevant indicators reflecting cardiac function: heart rate(HR),left ventricular development pressure(LVDevP),the maximum rate of left ventricular pressure rise(+ dp / dt)and the maximum rate of left ventricular pressure fall(-dp / dt),during the equilibrium phase before ischemia in experimental rats,at 30 min after ischemia reperfusion and at60 min after ischemia reperfusion,respectively.4.Use a test kit to evaluate the activity of lactate dehydrogenase(LDH)and creatine kinase MB(CK-MB)activity released from myocardial cells into coronary effluent during perfusion and to evaluate myocardium Damage enzymatic changes.5.Use HE staining to observe the damage of myocardial microstructure of the isolated rat heart,and to calculate the myocardial injury score.6.Observe the changes of myocardial mitochondria in isolated rat hearts by electron microscope.7.Detect myocardial infarction in isolated rat hearts by TTC staining,and calculate the percentage of myocardial infarction.Part II:The research on the mechanism of lycopene on myocardial ischemiareperfusion injury in ischemia postconditioning hypercholesterolemia rats.1.Examination the openness of myocardial MPTP in isolated rat hearts by applying the calcium induction,and the result of min / max A540 was calculated by the formula.2.Western blot was used to detect changes in endoplasmic reticulum stress(ERS)markers,reperfusion injury rescue kinase(RISK)pathway and mitochondrial apoptosisrelated proteins.3.Use TUNEL method to detect the myocardial apoptosis in isolated rat hearts,and calculate the myocardial apoptosis rate.Results:Part ?: 1.After 12 weeks,compared with the ND group,serum TC and LDL-C levels were significantly increased in the HCD group.There was no significant difference in serum TC and LDL-C levels before and after LP administration.2.Compared with the IR group,the cardiac function,the amount of CK-MB and LDH released,the degree of damage to myocardial microstructure and the percentage of myocardial infarction were not improved significantly in the IPoC group and LP-5group.3.Compared with the IR group,the cardiac function,the amount of CK-MB and LDH released,the degree of damage to myocardial microstructure and the percentage of myocardial infarction were all improved significantly in the IPoC + LP-5 group,LP-20 group and 4-PBA group.Part II:1.The sensitivity of myocardial mitochondria to calcium-induced MPTP opening and the min / max A540 value were not changed significantly,the protein expression levels of GRP78 and CHOP,the phosphorylation levels of AKT,ERK1 / 2,GSK-3?,the protein expression levels of Cytochrome C,Cleaved Caspase9,and Cleaved Caspase3 were not changed significantly,the rate of myocardial apoptosis was not changed significantly,in the IPoC group and LP-5 group compared with the IR group.2.The sensitivity of myocardial mitochondria to calcium-induced MPTP opening decreased significantly,and the min / max A540 value increased significantly,the protein expression levels of GRP78 and CHOP were significantly reduced,the phosphorylation levels of AKT,ERK1/2,GSK-3? were significantly increased,the protein expression levels of Cytochrome C,Cleaved Caspase9 and Cleaved Caspase3 were significantly reduced,the rate of myocardial apoptosis was significantly reduced,in the IPoC + LP-5 group,LP-20 group and 4-PBA group compared with the IR group.Conclusion: 1.LP can reduce the degree of myocardial ischemia-reperfusion injury in ischemia postconditioning hypercholesterolemia rats.2.LP activates the activation of RISK pathway,inhibits ERS,and inhibits the opening of MPTP and the activation of mitochondrial apoptotic pathway to reduce the degree of myocardial ischemia-reperfusion injury in ischemia postconditioning hypercholesterolemia rats. |
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