Danshensu Sodium Prevents I/R Arrhythmia By Inhibiting CaMK?-mediated Phosphorylation Of RyR2

Objective:The upregulation of cardiac ryanodine receptor phosphorylation byCa²⁺/calmodulin-dependent protein kinase?Ca MK??may contribute toarrhythmogenesis in myocardial ischemia-reperfusion injury?MIRI?.We previously demonstrated cardio-protection by Salvia miltiorrhiza Injection in rats with acutemyocardial infarction is related to ryanodine receptor.Danshensu is the main watercomponents of Danshen Injection.In this sudy we investigated the effects of Danshensu preconditioning on reperfusion arrhythmia,oxidative stress,assessed expression andphosphorylation status of calcium transport related protein expression in isolated I/R hearts,and measured spontaneous calcium waves and sarcoplasmic reticulum calcium leak in I/R cardiomyocytes.Methods:?1?The Langendorff perfusion device was used to establish I/R model and ECG records were used to monitor the occurrence of arrhythmia in isolated rat hearts during 30minutes of global ischemia followed by 30 minutes of reperfusion.The incidence of ventricular tachycardia and ventricular fibrillation,the time of occurrence and the number of occurrences were observed.The SOD activity and MDA content in collection of coronary flow and the retention of myocardial tissue for were measured.The protein expressions of Ca M,Ca MK?,Ry R2-p Ser2814 were examined by Western blot analysis.?2?In the cell experiment,the rat ventricular myocytes were isolated by enzymatic hydrolysis,and the MIRI cell model was established by perfusion of ischemic solution?sugar-free KH solution,2-Deoxy-D-glucose,Na₂S₂O₄?.Intracellular calcium imaging technique of laser confocal microscopy was used to observe spontaneous calcium waves and Calcium leakage from the sarcoplasmic reticulum.Results:?1?The results of isolated heart test showed that arrhythmia occurred in each group after ischemia-reperfusion.Danshensu reduced the incidence of arrhythmia?ventriculartachycardia and ventricular fibrillation?and shortened the occurrence time of arrhythmia caused by ischemia-reperfusion?P<0.05?in dose-dependent manner.The Danshensumiddle dose group can shorten the ventricular tachycardia and ventricular fibrillation time.Danshensu increased the activity of SOD and decreased the content of MDA in ischemia-reperfusion myocardium and coronary flow?P<0.05?.Ischemia-reperfusion activated Ca MK?and enhanced ryanodine receptor phosphorylation at Serine 2814.However,Danshensu preconditioning decreased Ca MK?dependent Ry R2phosphorylation during reperfusion.?2?Ischemia-reperfusion in cardiomyocytes produced spontaneous calcium waves and sarcoplasmic reticulum calcium leakage.Danshensu can reduce spontaneous calcium waves?P<0.01?and sarcoplasmic reticulum calcium leak?P<0.05?in ischemia-reperfusion myocardial cells.Conclusion:Danshensu reduced the occurrence of reperfusion arrhythmia in isolated heart,ameliorated oxidative stress and protected I/R myocardium.Danshensu regulated the gating of Ry R2 calcium release channel by mediating Ca MK?dependent Ry R2phosphorylation after myocardial ischemia-reperfusion,and reduced spontaneous calcium and sarcoplasmic reticulum calcium leak in I/R myocytes.