Cryptotanshinone And Salvianolic Acids Prevent Myocardial Ischemia-reperfusion Injury Through ENOS/GRK2 Pathway

Myocardial ischemia reperfusion injury can accelerate myocardial damage and seriously threaten human health.Salvia miltiorrhiza is a traditional Chinese medicine for cardiovascular diseases.Its effective ingredients are mainly classified into tanshinone(Cryptotanshinone,CT)and salvianolic acid(Danshensu,DSS and Protocatechuic acid,PCA),which have certain protective effects on cardiovascular system.In this study,myocardial ischemia reperfusion model and Western Blot were used to investigate the myocardial protective effect of cryptotanshinone and salvianolic acids(SAs)and its regulatory effects on eNOS/GRK2Methods: Hypoxia-reoxygenation(HR)model was established in vitro.Verapamil at 10 ?M as positive control,the cytoprotective effect of CT at 1,3,and 10 ?M,DSS at 3,10,and 30 ?M,PCA at 3,10 and 30 ?M,CT in combination with DSS and PCA were detected.And the concentrations of LDH,MDA and SOD in the culture medium were determined.The rat ischemia-reperfusion injury(Ischemia-reperfusion injury)model was established in vivo.The rats were randomly divided into normal group,sham group,model group,isosorbide mononitrate group,CT groups at 10,20 and 40 mg/kg,DSS groups at 30,60 and 120 mg/kg,PCA groups at 30,60 and 120 mg/kg,CT in combination with DSS groups,and CT in combination with PCA groups.Then,the myocardial infarction rate and the concentrations of CK-MB,LDH,MDA and SOD in plasma were measured.Western Blot was used to detect the levels of eNOS and GRK2 in vivo and vitro.Results: in vitro,HR could decrease the viabilities of myocardial cells(0.627 ± 0.021).CT,PCA and DSS at 10,30 and 30 ?M,respectively,can significantly increase the cell viabilities(0.778 ± 0.035,0.713 ± 0.036 and 0.886 ± 0.046,respectively)reduced by HR injury.The cell viabilities of CT in combined with PCA and DSS were significantly higher than that of alone(0.866 ± 0.027 and 0.902 ± 0.023,respectively).The concentrations of LDH,MDA and SOD were consistent with cell viabilities.In vivo,IR increased the rate of infarction(54.0% ± 3.4%).CT,PCA and DSS at 10,30 and 120 mg/kg,respectively,can significantly reduce the rate of infarction(20.0% ± 2.1%,17.4% ± 3.3% and 19.2% ± 5.7%,respectively).CT in combined with PCA and DSS can remarkable protect the myocardium(11.5% ± 1.4% and 14.4% ± 3.5%,respectively),which were significantly different from that of alone.Western Blot experiments in vivo and in vitro showed that,compared with the model group,CT can play a role in protecting myocardium by increasing the phosphorylation level of eNOS.Compared with CT,PCA and DSS can also increase the level of p-eNOS/GRK2 by reduce the level of GRK2,while have little influence on the phosphorylation of eNOS.CT combined with PCA and DSS can remarkable increase p-eNOS/GRK2 through aggrandizing p-eNOS,meanwhile,lower the expression of GRK2.Conclusion: CT combined with PCA and DSS can significantly increase the level of p-eNOS /GRK2 and protect myocardium.This study lays a foundation for further research on the mechanism of cardiovascular protection and the development and utilization of salvia miltiorrhiza,and provides a reference for the study on the effect of myocardial protection and mechanism of traditional Chinese medicine.