Anti-apoptosis Effect Of Yi Yin Helping Yang In Cardiomyocytes Induced By Isoproterenol

Objective:To study the effect of "Yi Yin helping Yang and activating blood treatment method" on the changes of hemodynamics,microscopic index pathology and neuroendocrine factors of macroscopical indexes in rats.And detected the effect of this method on cardiac and myocardial fibrosis in rats with heart failure.In order to provide more methods and ways for the clinical treatment of chronic heart failure.Further study at the cellular level,the possible mechanism of "Yi Yin helping Yang and activating blood treatment method" against myocardial apoptosis has been explained by testing relevant indexes on Isoproterenol induced rat primary myocardial cell injury.Methods:1、Establishment of heart failure model in SD rats,and SD rats anaesthetized by intraperitoneal anesthesia(1%Pentobarbital sodium 20-25 mg/kg),hypodermic injection ISO(170 mg/kg,Interval 24h)。A heart failure model can be formed in about 4 weeks,and examined the color doppler ultrasound,make short axis views of left ventricles.Model establishment standard:EF≤45%.2、Myocardial tissue from SD rat were gained to investigate the collagen tissue and fibrosis of rat myocardium in different experimental groups by Masson staining.3、Effects of "Yi Yin helping Yang and activating blood Therapy" on ALD,BNP,ET-1 and ANGⅡ in rats of each experimental group were investigated by ELISA.4.Preparation of contral,"Yi Yin helping Yang and activating blood Therapy" or "metoprolol"containing serum clinical equivalent.Blood were gained after the end of the experiment.ISO induced primary myocardial cell injury model were established in vitro.Detection of the protective effect of "Yi Yin helping Yang and activating blood Therapy" on ISO induced primary cardiomyocytes of SD rats injury model by CCK-8.5、Detection of cell apoptosis by Annexin-V/PI double staining method.6、Observation of the effect of "Yi Yin helping Yang and activating blood Therapy" on the morphology of the nucleus of the ISO induced primary cardiomyocytes of SD rats injury model by Hochest-PI double staining method.7、Detection of the effect of " Yi Yin helping Yang and activating blood Therapy" on the expression of Bcl-2,Bax and Bak in ISO induced primary cardiomyocytes of SD rats injury model by Western Blot.Results:1、Compared with the normal group,the LVDd and LVDS of SD rats in model group had increased significantly and LVPWd,LVPW,EF,FS,LVSd and LVSs had declined significantly.And all of these indicators have been improved to varying degrees after treatment of"Yi Yin helping Yang and activating blood Therapy".2、Compared with the model groups,the content of collagen tissue and the degree of fibrosis around the myocardium were significantly lower than that of the model group in Metoprolol group,middle dose treatment group and high dose treatment group.3、Compared to the normal group,the level of ALD,BNP,ET-1 and ANGⅡ in the serum of model rats increased obviously.Compared with the model group,the levels of ALD,BNP,ET-1 and ANG Ⅱ in the treatment group were all decreasing.4、Compared with blank group(Drug containing serum 0%),primary myocardial cell viability of each drug group(drug containing serum 5-20%)were increased.5、Compared with the model group,the apoptosis rate of Metoprolol group was down regulated(P<0.01),and " Yi Tin helping Yang " group in middle and high dose were down regulated(P<0.01).6、Compared with the model group,Metoprolol group and " Yi Yin helping Yang " group in middle and high dose of apoptotic cells with bright blue were decreased.7、Compared with the normal group,the expression of Bcl-2 protein in the model group was down regulated,and the expression of Bak and B’ax protein was up-regulated.And compared with the model group,the expression of Bcl-2 protein in "Yin Yin helping Yang" high dose group showed an uptrend,and the expression of Bak and Bax protein showed a downward trend.Conclusion:"Yi Yin Helping Yang and Activating Blood Therapy" has good curative effect to treat chronic heart failure.It can reduce the production of myocardial collagen,the level of LVDd and LVDS,and improve the level of LVSd,LVSs,LVPWd,LVPWs,EF,and FS.It also can reduce the the levels of ALD,BNP,ET-1 and ANG Ⅱ in serum.Study on the primary cardiomyocytes of SD fetal rats in vitro,we can find out that" Yi Yin helping Yang and activating blood Therapy" has anti apoptosis effect on the rat primary cardiomyocytes injury model induced by Isoproterenol.