New Ginsenoside AD2 Inhibits GPCR/cAMP/PKA Pathway And Inflammatory Factor Expression To Regulate Tachyarrhythmia

Objective: The purpose of this experiment is to investigate the regulatory effect of ginsenoside AD2 on isoproterenol-induced cardiac arrhythmia in rats.Methods: Primary neonatal rat ventricular myocytes were isolated,the effects of AD2 on the cell survival of primary neonatal rat ventricular myocytes were assessed by the CCK8 assay in vitro.Male Wistar rats were randomly allocated to four groups: the CON group,the ISO model group,the AD2+ISO group,and the AD2 group.The Langendorff-perfused rat hearts were subjected to different drugs,myocardial electrical activity was performed with a multi-electrode array in isolated rat hearts.Then,high-throughput gene sequencing,and biological information analysis were performed on the ventricular muscles of the CON group and the ISO model group to speculate the possible pathogenic genes of the arrhythmia model group.The Real-Time PCR technology was used to detect the levels of GPR35,GPR65 and GPR132 m RNA in the ventricular muscles of rats in each group to determine the myocardial cell membrane targets of AD2 in inhibiting tachyarrhythmia.The activity of cAMP and PKA activity was measured with targeted fluorescence resonance energy transfer probes in primary neonatal rat ventricular myocytes.Inflammatory factor levels in isolated rat hearts were measured by ELISA.Fluorescence mapping technology was used to detect the changes of AD2 on ISO-induced APD90,potential activation time,and heart rate of guinea pig cardiomyocytes.C57BL/6 mice were randomly divided into four groups,CON group,ISO group,AD2+ISO group,and AD2 group.The changes in heart rate and electrocardiogram of mice in different groups were detected by BL-422 I.Myocardial fibrosis in mouse heart tissue was detected by Masson staining.Results: AD2(0～50μM)had no significant differences in primary neonatal rat ventricular myocytes survival.Compared to the CON group,rat hearts perfused with ISO showed changes in the direction of conduction,increased left ventricular conduction velocity,APD90,and heart rate(P<0.05).shortened the potential activation time.GPR35 m RNA level,cAMP,and PKA levels in primary neonatal rat ventricular myocytes,as well as the expression of inflammatory cytokines,including CXCL1,CXCL2,CXCL3,and TNF-α,were also significantly increased compared with the CON group(P<0.05).Besides,compared to the CON group,the PR interval was shortened(P<0.05),and the QRS duration and QT interval were prolonged(P<0.05),the severity of myocardial fibrosis was increased in mouse hearts with ISO.However,compared with the ISO group,the AD2+ISO group showed a significant decrease in heart rate(P<0.05),APD90,GPR35 m RNA level,cAMP,and PKA levels,the expression of inflammatory cytokines(P<0.05),QRS duration,QT interval(P<0.05),and the severity of myocardial fibrosis.In addition,the action potential activation time and PR interval were prolonged in the AD2+ISO group.Conclusion: AD2 alleviates ISO-induced tachyarrhythmia by inhibiting GPR35/cAMP/PKA pathway and inflammatory factor levels(CXCL1,CXCL2,CXCL3,TNF-ɑ).