| Crataegus pinnatifida Bunge(Botan-rosaceae) is a traditional Chinese medicine, which has digestive effect.It was found that the flavones,were extracted from Crataegus pinnatifida Bunge,could lower blood fat,have hypotensive effect,increase coronary flow and have protective effects on myocardial ischemic injury,etc.Vitexin,a flavone glycoside(8-C-β-D-glucopyranosyl-apigenin),is isolated from the leaf of Crataegus pinnatifida Bunge which is attributed with varied medicinal properties. Vitexin has been demonstrated to have hypotensive effect and anti-inflammatory action. However,there's been no study about vitexin on myocardial ischemic and reperfusion injury.This research therefore designed to observe the protective effect of vitexin on myocardial ischemic and reperfusion injury and the possible underlying mechanism.Partâ… The protective effects of vitexin on experimental myocardial ischemia model1.Effect of vitexin on ECG time in trachea clamping miceOn the myocardial anoxia model induced by clamping trachea in mice,it is found that the ECG survival time was much longer in the groups pretreated with vitexin(12, 6,3 mg·kg-1) than that in NS control group.The maximal elongation rate of ECG survival time reached a height of 32.94%.2.Protective effect of vitexin on acute myocardial ischemia injury induced by isoproterenol in ratsAcute myocardial ischemia episodes and typical isehemia ECG developed after subcutaneous injection(sc) of Iso.The results showed that vitexin(6,3,1.5 mg·kg-1) could ameliorate the changes of segment S-T of ECG(P＜0.05 or P＜0.01 compared with model).The myocardial water content(MWC) in model group was 1.89%higher than NS control.Vitexin(6,3 mg·kg-1) can significantly inhibit the increase of MWC and the inhibition rates were 52.78%and 38.19%,respectively(P＜0.05 or P＜0.01 compared with model).3.Protective effects of vitexin on experimental myocardial ischemia induced by pituitrin in ratsTo explore the protective effects of vitexin on experimental acute myocardial ischemia induced by pituitrin(Pit) in rats.The ischemia model was established induced by 1.5 IU·kg-1 Pit through intravenous injection.It was showed that the S-T segment of ECG in model group rosed significantly at 30 second,1min,5min,10min,20min of ischemia induced by Pit(P＜0.01 compared with control).The administration of vitexin (6,3,1.5 mg·kg-1) markedly decreased the elevation of segment S-T of ECG(P＜0.05 or P＜0.01 compared with model),reduced the activity of lactate dehydrogenase(LDH) and creatine phosphokinase(CPK) and increased the activity of superoxide dismutase (SOD) and glutathione peroxidase(GSH-PX) in the serum of rats(P＜0.05 or P＜0.01 compared with model),vitexin(6,3,1.5 mg·kg-1) significantly increased the activity of ATPase in the myocardium of rats(P＜0.05 or P＜0.01 compared with model).The administration of vitexin(6,3 mg·kg-1) improved myocardial pathologic alternation.4.Protective effects of vitexin on experimental myocardial infarction by ligating left anterior descending coronary artery in ratsTo study the protective effect of vitexin on experimental acute myocardial infarction(AMI) by ligating left anterior descending coronary artery(LAD) in rats.The results showed that vitexin(6,3,1.5 mg·kg-1) could decrease the infarction range marked by N-BT staining,and vitexin(6,3mg·kg-1) could reduce the activities of lactate dehydrogenase(LDH) and creatine phosphokinase(CPK) in the serum of rats(P＜0.05 or P＜0.01 compared with model).5.Protective effects of vitexin on experimental myocardial infarction by ligating left anterior descending coronary artery in dogsTo observe the protective effect of vitexin on experimental acute myocardial infarction(AMI) by ligating left anterior descending coronary artery(LAD) in dogs.36 hybrid dogs were divided into 6groups at random.Vitexin(6,3,1.5 mg·kg-1) could decrease the myocardial ischemic risk and the infarction size,and vitexin(6,3 mg·kg-1) could decrease the myocardial index(MI)(P＜0.05 or P＜0.01 compared with model) in dogs as well.it was seen that vitexin could reduce the degree of myocardial infarction (Σ-ST) and the myocardial infarction range(N-ST) in dogs(P＜0.05 or P＜0.01 compared with model).At the same time,vitexin could reduce the activities of lactate dehydrogenase(LDH) and ereatine phosphokinase(CPK) in the serum of dogs(P＜0.05 or P＜0.01 compared with model).6.Effect of vitexin on the formation of throbosis on the A-V thrombosis pass-by model in ratsTo observe the effect of vitexin on the formation of throbosis on the A-V thrombosis pass-by model in rats.The results showed that vitexin(6,3 mg·kg-1) could obviously decrease the dry weight of thrombosis on the A-V thrombosis pass-by model.Partâ…¡The effects and mechanism of vitexin on myocardial ischemia/reperfusion injury in rats in vitro To study the effect and mechanism of vitexin on myocardium ischemia/reperfusion in rats in vitro.To set rats model of MI/R in vitro,and observe the coronary flow and pathologic changes of myocardium with HE staining,give a morphological observation with TEM,and detect tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β) with RIA, apoptosis rate of myocardium with TUNEL,protein expression of Bax,Bcl-2,ICAM-1 and NF-κB p65 with immunohistochemical method.The results showed that vitexin could significantly inhibit the reductions of coronary flow,and improve myocardial pathologic and ultrastructure changes in MI/R,reduce the contents of TNF-α,IL-1βin myocardial homogenate,inhibit apoptosis of cardiac muscle cell,down regulate the expression of Bax gene and up regulate the expression of Bcl-2 gene,and inhibit the protein expression of NF-κB p65.Partâ…¢Mechanisms of vitexin preconditioning effects on cultured neonatal rat cardiomyocytes with anoxia and reoxygenationTo observe the protective effects and its mechanism of vitexin preconditioning (VPC) on cultured neonatal rat cardiomyocytes after anoxia and reoxygenation(A/R). An A/R model was established using cultured neonatal rat cardiomyocytes.Cellular injury was evaluated by measuring cell viability,the releases of creatine kinase(CPK), and lactate dehydrogenase(LDH).We measured the apoptosis rate of cardiomyocytes after Anoxia/reoxygenation,activities of extracellular signal-regulated protein kinases (ERKs).The intracellular calcium indicated by the fluorescence in cardiomyocytes was measured by the laser confocal microscope.The results showed that 10,30 and 100μmol/L Vitexin preconditioning significantly enhanced the cell viability from 82.47±0.97(A/R group) to 91.58±1.32%,93.44±0.89%and 95.87±0.73%(p＜0.01), respectively.Vitexin preconditioning(10,30 and 100μmol/L) markedly inhibited A/R-induced increases of LDH and CPK release(p＜0.05 or p＜0.01).By using Hoechst33258 method,it was found that vitexin preconditioning 10,30,100μmol/L obviously decreased the number of apoptotic cardiomyocytes(p＜0.01).The examination of flow cytometer(FCM) method also showed in range of 10～100μmol/L, vitexin preconditioning had significant inhibitory effect on A/R-induced cardiomyocytes apoptosis.Vitexin preconditioning 10,30 and 100μmol/L markedly decreased the fluorescence intensity value of[Ca2+]i in cardiomyocytes from 37.78±4.90(A/R group) to 28.65±6.12,26.55±4.53 and 23.15±3.85((p＜0.05 or p＜0.01),respectively.The band for phospho-ERK was observed in cardiomyocytes of each group.Exposure to anoxia or vitexin preconditioning significantly increased the phospho-ERK level,and the increase was markedly inhibited by PD98059,an inhibitor of the upstream kinase of ERK.Conclusions:The protective effects and mechanism in anti-myocardial ischemia and reperfusion of vitexin were studied.It was found that vitexin could protect heart against myocardial ischemia and reperfusion injury,the protective mechanism may related with anti-oxidation,inhibiting inflammatory factor releasing and the cardiomyocytes apoptosis,lowing the cardiomyocytes calcium overload and inhibiting nuclear ectopy of NF-κB p65 and increasing the abundance of phosphor-ERK1/2 of the cardiomyocytes.
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