| Purpose:By the measurement of left ventricular fractional shortening(FS)and ejection fraction(EF)of classical prescription Zhenwu Decoction on heart kidney Yang deficiency model rats with heart failure through the cardiac color doppler ultrasound,by the influence of brain natriuretic peptide(BNP),tumor necrosis factor alpha(TNF-?),interleukin-6(IL-6),galectin-3(Galectin-3)and heat shock protein 70(HSP70)in rat serum,B-cell lymphoma-2(Bcl-2),Bcl-2 associated X protein(Bax),caspase-3(Caspase-3)and silent information regulator 1(Sirt-1)protein expression in myocardial tissue,and by the observation of Zhenwu Decoction on myocardial cell ultrastructure changes to explore the possible mechanism of classical prescription Zhenwu Decoction on treating heart failure.Provide the scientific and objective experimental data and theoretical basis for Zhenwu Decoction on the treatment of heart failure.Material and method:80 healthy male Wistar rats were randomly divided into the normal group-the blank group(n=10),and the operation group(n=70).Replicate the heart kidney Yang deficiency rat model with heart failure after myocardial infarction by ligation the anterior descending branch of coronary artery and exhaustive swimming with cold water in operation group of 70 rats,the 50 rats of successful model were randomly divided into low-dose group of Chinese medicine,medium-dose group of Chinese medicine,high-dose group of Chinese medicine,western medicine group and model group,each group of 10 arts.Normal group and model group were given saline to fill the stomach,western medicine group was given captopril 10mg/(kg · d)to fill the stomach,low-dose group of Chinese medicine was given low-dose Zhenwu Decoction equivalent to the amount of crude drug9.45g/(kg · d),medium-dose equivalent to the amount of crude drug 18.9g/(kg · d)and high-dose equivalent to the amount of crude drug 37.8 g/(kg·d)to fill the stomach.Liquid of six groups were made into 6ml,two times a day orally,3ml each time,lasting 15 days.Experiment one:By color doppler ultrasound for the measurement of left ventricular fractional shortening(FS)and ejection fraction(EF).By enzyme-linked immunosorbent assay method(ELISA)for the determination of brain natriuretic peptide(BNP),tumor necrosis factor alpha(TNF-alpha)and interleukin-6(IL-6),galectin-3(Galectin-3),heat shock protein70(HSP70)level in the rat serum.Experiment two:By using immunoblotting method(Western blotting)for the determination of B-cell lymphoma-2(Bcl-2),Bcl-2 associated X protein(Bax),caspase-3(Caspase-3)and silent information regulator 1(Sirt-1)protein expression level in the rat myocardial tissue.Experiment three:By H-7650 electron microscope for the observation of the ultrastructural changes of the rat myocardial cells.Statistical method used SPSS17.0 software for statistical analysis.The measurement data resulted with mean +/-standard deviation(±s)expression.Use the single factor variance analysis for comparison among groups.If it had homogeneity of variance,the result of P<0.05 was significant difference index.Results:1.The general condition of rats in each group:Rats in the normal group had eyes with god,sensitive response,flexible movement,shiny and dense fur which was close to the body,normal appetite,normal feces and urine;Rats in the model group had dull eyes,listlessness,slow activity,decreased endurance,down back temperature,cold and dark tail,dry and erect fur which was easy to fall off,huddle body and shrink shoulder,flock together,foot dorsum edema,weight decreased,loss of appetite,the amount of urine reduced,loose stools and other features of heart kidney Yang deficiency.The general conditions of rats in the western medicine group and Chinese medicine high-dose group were close to the normal group;The general conditions of rats in the Chinese medicine low-dose and medium-dose groups were between Chinese medicine high-dose group and model group;During the feeding period,rats in each Chinese medicine group had different degrees of urine volume increased,and the general condition of rat spirit,fur,back temperature,diet and so on was not significantly different with the western medicine group.It indicated Zhenwu Decoction could obviously improve the general condition of heart kidney Yang deficiency syndrome rats with heart failure.2.FS and EF value of rat heart:Compared with the normal group,FS value and EF value were significantly decreased in the model group(P<0.01)with statistical significance,it indicatedthat the heart function of rats in model group was damaged,and the heart failure model was successfully constructed,FS and EF value in the western medicine group and Chinese medicine high-dose group were no significant difference compared with the normal group(P>0.05)without statistical significance;Compared with the model group,FS and EF value in the western medicine group and Chinese medicine high-dose group were significantly higher(P<0.01)with statistical significance,FS and EF value in Chinese medicine low-dose and medium-dose groups were no significant difference(P>0.05)without statistical significance;Compared with the western medicine group,FS and EF value in each Chinese medicine group had no obvious difference(P>0.05)without statistical significance.By color doppler ultrasound in detecting rat heart FS and EF value,it indicated Zhenwu Decoction could significantly improve cardiac function in heart failure rats,and Zhenwu Decoction effect had no significant difference with Kato Pury effect in western medicine group.3.Serum brain natriuretic peptide BNP levels:Compared with the normal group,the level of BNP in the model group was significantly higher(P<0.01)with statistical significance,it indicated the success of rat model with heart failure,compared with the normal group,western medicine group and low-dose Chinese medicine group BNP levels were obviously increased(P<0.05)with statistical significance,compared with the normal group,Chinese medicine medium-dose and high-dose groups had no obvious difference(P>0.05);Compared with the model group,western medicine group and Chinese medicine groups BNP levels were decreased,the western medicine group and low-dose group BNP levels were obviously lower(P<0.05)with statistical significance,Chinese medicine medium-dose and high-dose groups BNP levels were significantly decreased(P<0.01)with statistical significance;Compared with the western medicine group,Chinese medicine high-dose group BNP level was obviously lower(P<0.05)with statistical significance,Chinese medicine low-dose and medium-dose groups BNP levels were no significant difference(P>0.05)without statistical significance.It indicated Zhenwu Decoction could improve heart function of rats with heart failure in a certain extent,and curative effect of Zhenwu Decoction high-dose group was obviously better than captopril group.4.Serum TNF-??IL-6 levels:Compared with the normal group,TNF-? levels of each group except high-dose Chinese medicine group and IL-6 levels of each group were increased(P<0.05,P<0.01)with statistical significance;Compared with the model group,western medicine group and Chinese medicine group TNF-? levels were lower,the western medicine group,low-dose Chinese medicine group and medium-dose Chinese medicine group TNF-?levels were obviously decreased(P<0.05)with statistical significance,high-dose Chinese medicine group TNF-? level was significantly decreased(P<0.01)with statistical significance,compared with the model group,medium-dose Chinese medicine group IL-6 level was obviously decreased(P<0.05)with statistical significance,high-dose Chinese medicine group IL-6 level was significantly decreased(P<0.01)with statistical significance;Compared with the western medicine group,medium-dose Chinese medicine group and high-dose Chinese medicine group TNF-? levels were obviously significant(P<0.05)with statistical significance,compared with the western medicine group,Chinese medicine groups IL-6 levels were no obvious difference(P>0.05)without statistical significance.It Indicated TNF-? and IL-6 levels were correlated with the degree of heart failure,Zhenwu Decoction could improve heart function of rats with heart failure in a certain extent by regulating the levels of TNF-and IL-6.Its curative effect was better than or equal to the captopril group.5.Serum Galectin-3?HSP70 levels:Compared with the normal group,Galectin-3 levels of each group were increased(P<0.05,P<0.01),model group HSP70 level was significantly increased(P<0.01)with statistical significance,low-dose Chinese medicine group and medium-dose Chinese medicine group HSP70 levels were obviously increased(P<0.05)with statistical significance;Compared with the model group,western medicine group,medium-dose Chinese medicine group and high-dose Chinese medicine group Galectin-3 levels were significantly decreased(P<0.01)with statistical significance,low-dose Chinese medicine group Galectin-3level was obviously decreased(P<0.05)with statistical significance,western medicine group and high-dose Chinese medicine group HSP70 levels were significantly decreased(P<0.01)with statistical significance,low-dose Chinese medicine group and medium-dose Chinese medicine group HSP70 levels were obviously decreased(P<0.05)with statistical significance;compared with western medicine group,Each of Chinese medicine group Galectin-3 and HSP70 levels had no significant difference(P>0.05)without statistical significance.It Indicated Galectin-3 and HSP70 levels were associated with the degree of heart failure,Zhenwu Decoction had good curative effect on the treatment of heart failure byregulating Galectin-3 and HSP70 levels.Its curative effect had no obvious difference with the captopril western medicine group.6.Bax and Bcl-2 protein expression in myocardial tissue:Compared with the normal group,model group Bax protein expression level was obviously increased(P<0.05)with statistical significance,model group Bcl-2 protein expression level was obviously decreased(P<0.05)with statistical significance;Compared with the model group,Zhenwu Decoction medium-dose group,Zhenwu Decoction high-dose group and western medicine group Bax protein expression levels in rat myocardial tissue were obviously decreased(P<0.05)with statistical significance,Zhenwu Decoction low-dose group Bax protein expression level had no significant difference(P>0.05)without statistical significance compared with the model group.Compared with model group,Zhenwu Decoction medium-dose group,Zhenwu Decoction high-dose group and western medicine group Bcl-2 protein expression levels were obviously increased(P<0.05)with statistical significance,Zhenwu Decoction low-dose group Bcl-2 protein expression level had no significant difference(P>0.05)without statistical significance compared with the model group;Compared with western medicine group,Bax and Bcl-2 protein expression levels of Zhenwu Decoction low-dose,middle-dose and high-dose groups had no significant difference(P>0.05)without statistical significance.It showed Bax protein expression was high,Bcl-2 protein expression was low on the heart failure rats.Bax and Bcl-2 were closely related with the degree of heart failure.Zhenwu Decoction might improve rat heart function to treat heart failure by regulating Bax and Bcl-2 protein expression.7.Caspase-3 and Sirt-1 protein expression in myocardial tissue:Compared with the normal group,model group Caspase-3 protein expression level was obviously increased(P<0.05)with statistical significance,model group Sirt-1 protein expression level was obviously decreased(P<0.05)with statistical significance;Compared with the model group,Zhenwu Decoction medium-dose group and western medicine group Caspase-3 protein expression level were obviously decreased in myocardial tissue(P<0.05)with statistical significance,Zhenwu Decoction low-dose group and high-dose group Caspase-3 protein expression level had no significant difference compared with the model group(P>0.05)without statistical significance.Compared with the normal group,Zhenwu Decoction medium-dose group andwestern medicine group Sirt-1 protein expression level were obviously increased(P<0.05)with statistical significance.Zhenwu Decoction low-dose group and high-dose group Sirt-1protein expression level had no significant difference compared with the model group(P>0.05)without statistical significance;Compared with western medicine group,Caspase-3 and Sirt-1protein expression levels of Zhenwu Decoction low-dose,middle-dose and high-dose groups had no significant difference(P>0.05)without statistical significance.It showed Caspase-3and Sirt-1 were related with the degree of heart failure.Zhenwu Decoction might improve ventricular remodeling to improve heart function by regulating Caspase-3 and Sirt-1 protein expression.8.The changes of myocardial cell ultrastructure:Compared with the blank group,cell nucleus became expanding,mitochondrion became swelling obviously part of which was vacuolar,cristae became thinner and shorter,the direction became irregular,myofilament became fracture,dissolution and disorder,each band was difficult to distinguish,intercalated disc disappeared in the model group.Compared with the model group,myocardial cell ultrastructure of Zhenwu Decoction low-dose,middle-dose,high-dose groups and western medicine group had different degree improvement,mitochondrial arrangement ruled which was circular,elliptical or rod shape,cristae was clearly visible,myofilament orderly arranged,each band was obviouser,disc was more complete,Zhenwu Decoction high-dose group was the most significant.It indicated Zhenwu Decoction could obviously improve myocardial cell ultrastructure,and Zhenwu Decoction high-dose group improved most obviously.Conclusion:1.Replicate the rat model with heart failure(heart kidney Yang deficiency syndrome)after myocardial infarction successfully by ligation the anterior descending branch of left coronary artery and exhaustive swimming with cold water.2.Zhenwu Decoction could obviously improve the general condition of heart kidney Yang deficiency syndrome rats with heart failure.3.Zhenwu Decoction could significantly improve the cardiac FS and EF value of heart failure rats,reduce the serum BNP level in rats with heart failure and improve heart function.4.Zhenwu Decoction could obviously reduce TNF-? and IL-6 level of rat serum with heartfailure,it indicated Zhenwu decoction might improve ventricular remodeling and heart function by inhibiting the inflammatory factor TNF-? and IL-6 release through neuroendocrine system from the neuroendocrine point of view.5.Zhenwu Decoction could obviously reduce Galectin-3and HSP70 level of rat serum with heart failure,it indicated that Zhenwu Decoction could regulate Galectin-3and HSP70 level,inhibit myocardial fibrosis,protect myocardial cells and improve ventricular remodeling to treat the heart failure from the myocardial fibrosis point of view.6.Zhenwu Decoction could reduce Bax protein expression,increase Bcl-2 protein expression,reduce Caspase-3 protein expression and increase Sirt-1 protein expression obviously in rat myocardial tissue.From the apoptosis point of view it indicated that Zhenwu Decoction could regulate apoptosis related gene protein,regulate the apoptosis signaling pathway and inhibit oxidation stress to inhibit the apoptosis of myocardial cells and improve ventricular remodeling so as to play the role of heart failure treatment.7.From the myocardial ultrastructure point of view it confirmed that Zhenwu Decoction could obviously improve myocardial ultrastructure of rats with heart failure and improve heart function so as to treat heart failure. |
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