| Objective:Mitochondrial dysfunction and oxidative stress play an important role in the pathogenesis of atrial fibrillation（AF）and diabetes mellitus（DM）.However,the mechanism is still unclear.Wenxin Keli（WXKL）is a traditional Chinese medicine and is used for the treatment of different clinical arrhythmias.Current research showed that WXKL can shorten action potential duration,prolonging atrial effective refractory period and atrial-selectively inhibiting sodium currents to prevent AF.In this study,we aim to investigate the potential therapeutic effects and mechanisms of WXKL in atrial remodeling and atrial muscle mitochondrial function by hydrogen peroxide（H2O2）-induced the oxidative stress in atrial fibroblast and type 2 diabetic rat models.Methods:Primary atrial fibroblasts of neonatal SD rats were divided into four groups:control,H2O2,H2O2+WXKL 1?g/L,and H2O2+WXKL 3?g/L groups.Intracellular mitochondrial membrane potential（MMP）,reactive oxygen species（ROS）and mitochondrial oxygen consumption were measured.SD male rats were randomly divided into three groups:control,DM,DM+WXKL groups.Rats in the DM+WXKL group were treated with daily gavage of WXKL at 3g/kg.We performed following experiments on all rats after 8 weeks of treatment.Left atrial（LA）diameter（LAD）,left ventricular end-diastolic dimension（LVEDD）,and left ventricular end-systolic dimension（LVESD）,left ventricular posterior wall thickness（LVPW）and interventricular septal thickness（IVS）were obtained in the parasternal long-axis view.Aortic systolic blood pressure（SBP）,diastolic blood pressure（DBP）,and mean blood pressure（MBP）were recorded carefully after a stabilization period.Subsequently,the cannula was inserted through the aortic valve to the left ventricle to record the left ventricular end-diastolic pressure（LVEDP）and maximum increasing or decreasing rate of left ventricular pressure（±dp/dt max）.Electrocardiogram were selected to measure heart rate,P wave duration,PR interval,QRS duration,and QT interval.Sinus cycle length（SCL）,interatrial conduction time（IACT）,atrial-ventricular Wenckebach cycle length conduction（AVWCL）,atrial effective refractory period（ERP）and AF inducibility was measured by a Langendorff perfusion system.The tissues were stained with hematoxylin and eosin（HE）to observe the morphological changes and Masson’s trichrome stain to evaluate interstitial fibrosis.Western blot was used to determine oxidative stress,inflammation,fibrosis and mitochondrial-related proteins.Results:（1）ROS was significantly higher and MMP was significantly reduced in the H2O2group compared with the control group.Basal respiration and maximal respiration were also decreased in the H2O2group compared with the control group.The above changes were improved in the H2O2+WXKL 3g/L group.（2）Fasting blood glucose levels（FBG）and serum insulin level of the DM and DM+WXKL groups were significantly higher than the control group.LAD was increased in DM group compared with control group.LAD in the DM+WXKL and DM groups were not significantly different.（3）Atrial conduction velocity was significantly lower in DM group than control group.Conduction dispersion was higher in the DM group than the control group.Conduction velocity and dispersion of the DM+WXKL group was improved compared with DM group.No significant differences of SCL,AVWCL and ERP between three groups.AF incidence was significantly higher in the DM group than control group.The AF incidence in the DM+WXKL group was considerably lower than that in the DM group.Cross-sectional area of cardiomyocytes was higher in the DM group than control group.The ratio of myocardial interstitial fibrosis was higher in DM group compared with control group.However,cardiomyocyte hypertrophy and fibrosis ratio were alleviated in the DM+WXKL group.（4）hs-CRP,TC and TG levels were higher in the DM group and DM+WXKL group than the control group.There was no difference between DM+WXKL group.Lower SOD and higher MDA levels were detected in the DM group compared to the control group.Serum levels of BUN,Cr,HDL-C and LDL-C were not significantly different amongst three groups.TGF-βand NF-κb were up-regulated in the DM group and DM+WXKL group.Compared with the control group,the expressions of fibrotic proteins,collagen I,collagen III andα-SMA were significantly increased in the DM group.These changes were significantly reversed in DM+WXKL group.The expression level of Bax was up-regulated and Bcl-2 was down-regulated in the DM group.The expression levels of Bax and Bcl-2 were improved in DM+WXKL group compared with DM group.（5）MMP was decreased in the DM group compared with the control group.The mitochondrial morphology was swollen,vacuolated,sacral rupture,and myofilament ruptured.The above changes were improved in the DM+WXKL group.The protein expression levels of TFAM,Drp1,Mfn1 and Mfn2were significantly lower in the DM group compared with control group and these mitochondria-associated proteins were significantly increased in the DM+WXKL group.Conclusion:WXKL can inhibit oxidative stress and improve mitochondrial function in vitro induced by H2O2and in vivo induced by DM,alleviate atrial remodeling and reduce AF incidence in DM rats. |
