| Cardiovascular disease has become the leading cause of death in urban and rural residents in China.Heart failure(HF)is almost the end stage of all cardiovascular diseases.The important progress has been made in the management of HF,but its morbidity has remained high in recent years.At present,domestic and foreign studies suggest that the basic mechanism of HF is myocardial remodeling,and apoptosis is a turning point from HF to decompensation.A large number of clinical trials show that Chinese medicine has a significant therapeutic effect on HF.The Chinese medicine pathogenesis of chronic heart failure(CHF)is mainly the virtual standard,and the virtual deficiency is yang deficiency,qi and blood deficiency,the standard is mostly blood stasis,sipping.Sini Decoction(SND)is from ShangHanLun,which is the one of the yang-warming therapies.Based on the above,we observed the effect of SND on cardiac function and myocardial apoptosis in mice with CHF after Transverse Aortic Constriction(TAC).ObjectiveTo establish a CHF model by TAC,evaluate the cardiac function in CHF mice after the therapeutic effect of SND,detect the expression level and molecular expression level of apoptosis-related protein,and discuss the related mechanism.MethodsEighty male C57BL/6J mice were randomly divided into sham group(n=10)and model group(n=70).The CHF mice model was established by TAC.The sham group is threaded and not narrowed.Four weeks after TAC,the mice that were successfully modeled were randomly divided into model group,SND high dose group,SND medium dose group,SND low dose group and benazepril group.The medicines were administered continuously for 8 weeks,and the dosage are 16.8g/(Kg·d)in the SND high dose group,8.4g/(Kg·d)in the SND medium dose group,4.2 g/(Kg·d)in the SND low dose group and 1.3 mg/(Kg·d)in the benazepril group,the model group and the sham group were given an equal volume of deionized water,the gastric volume was 8.4mL/(Kg·d).Eight weeks later,small animal echocardiography was used to detect the corresponding indicators of cardiac function in each group.Left ventricular short-axis papillary muscles were recorded with M-mode ultrasound to record left ventricular motion curve,measured left ventricular end-systolic diameter(LVIDs),inter-ventricular septum end-stolic thickness(IVSs),left ventricular posterior wall end-diastole thickness(LVPWs),left ventricular end-diastole diameter(LVIDd),inter-ventricular septum end-diastole thickness(IVSd),left ventricular posterior wall end-diastole thickness(LVPWd).After two-dimensional ultrasound was taken from the apical four-chamber view,the PW mode was switched to detect the peak flow velocity(E)of the early mitral valvular E wave and the peak flow velocity(A)of the late diastolic A wave.The parameters were continuously measured for three cardiac cycles.Left ventricular ejection fraction(LVEF),left ventricular short-axis contraction rate(LVFS)and E/A values were calculated.HE staining was used to observe the pathological changes of myocardial tissue in each group.TUNEL pod method was used to detect myocardial cells in myocardial tissues.The apoptotic index was calculated and the protein expression level of Bax and Bcl-2 in myocardial tissue was detected by Western blot.Protein expression levels of Cytochrome C,Caspase9 and Caspase3 were detected by ELISA.RT-PCR for detection of Caspase9 and Caspase3 mRNA transcription levels.Results1 General situation:In the model preparation process and during the 4 weeks of routine feeding,9 mice died.The causes of death may be pneumothorax,vascular rupture and excessive ligation.The remaining 61 mice underwent ultrasound examination 4 weeks after surgery,and the rate of formation was 75%(46/61).Forty-six successful mice were randomly divided into 5 groups:model group(n=10),benazepril group(n=9),and SND high(n=9),medium(n=9)and low dose(n=9)groups.One mouse died in each the sham group and the model group during the intragastric administration period.Therefore,the total number of mice observed was 45,9 in each group.Throughout the whole process,it was observed that the mice in the model group had poor appetite and liked agglomeration,and the activity was significantly reduced compared with the sham group.2 Echocardiographic results:Compared with the sham group,the LVEF,LVFS,and IVSs in the model group were significantly lower,and IVSd,LVIDd,LVPWd,LVPWs,E/A did not change obvious.Compared with the model group,the LVEF and LVFS in the SND medium dose group and the benazepril group were significantly increased(P<0.05 or P<0.01),IVSs in the SND high dose and low dose group were lower(P<0.05),and there were not obvious changes in other index(P>0.05).3 HE staining showed that the sham group had tightly arranged myocardial fibers,clear nuclei and uniform cytoplasmic staining;in the model group,the number of myocardial fibers was significantly reduced in the same multiple unit,the transverse diameter of myocardial fibers was widened,the coloration of myocardial cytoplasm was uneven,and the color was shallow.In addition,the myocardial cells were loosely connected.In the benazepril group,the myocardial fibers in SND high dose,medium dose and low dose group showed different degrees of uneven staining.Among them,the SND high dose and low dose group had myocardial fiber rupture.The boundary between myocardial fibers was unclear and the gap was increased.The integrity and continuity of myocardial fibers were worse than those in the benazepril group and SND medium dose group.4 TUNEL staining showed that the nucleus of sham group showed more blue zone,less brown particles than other group,and the brown nucleus of the model group increased significantly.The medication groups had different degrees of brown nucleus.After calculation,it was found that the model group was significantly higher than the sham group(P<0.01).Compared with the model group,the benazepril group and the SND medium dose group were lower(P<0.05),and there was no significant change in the SND high dose and the low dose group(P>0.05).5 Western Blot results:Compared with the sham group,Bcl-2 was significantly decreased in the model group and each medication group(P<0.05 or P<0.01),while Bax were not significant change;there was no significant change of Bax in each medication group compared with the model group(P>0.05).6 ELISA was used to detect the cytochrome C protein:the model group and the SND high dose,medium dose and low dose group were significantly elevated than sham group;there were significant decrease in the SND high dose,medium dose group and benazepril group than model group(P<0.05 or P<0.01),while SND low dose group was not obvious change(P>0.05).7 RT-PCR results:compared with the sham group,the Caspase9 and Caspase3 mRNA in the model group increased significantly;and Caspase3 mRNA in the SND high dose,medium dose group and benazepril group decreased significantly(P<0.05),and there was no significant change in SND low dose group(P>0.05).8 ELISA was used to detect the expression of Caspase9 and Caspase3:the expression of Caspase9 and Caspase3 in the model group were higher than the sham group,and SND high dose,medium dose and low dose group and benazepril group were significantly reduced than model group(P<0.01),Caspase9 in the SND low dose group reduced as well(P<0.01),while Caspase3 was not change(P>0.05).Conclusion1 TAC can successfully establish a model of CHF in mice.2 SND can improve the cardiac function in mice with CHF,improve the LVEF during compensatory period,and reduce the apoptosis of cardiomyocytes.The mechanism may be related to cytochrome C,Caspase9 and Caspase3 protein level. |
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