The Research On Huatan Qushi Huoxue Recipe Intervention Nonalcoholic Steatohepatitis Rat Model Of ADPN/ACC/AMPK Pathway

Objective：This topic based on previous experimental studies,by feeding with highlipid food and celio-injecting tetracycline to make rat nonalcoholic steatohepatitis model,discuss the effect of Huatan Qushi Huoxue Recipe （HQHR） of model ratADPN/AMPK/ACC signal pathway of protein and gene, reveal ADPN/AMPK/ACCsignaling mechanism in the developing of NASH,interpretation of HQHR prevention andtreatment of NASH, the protein and gene level to provide theoretical basis for the TCMprevention and treatment of NASH.Methods：Using polyene phosphatidylcholine capsule （Yi Shan Fu） as positivecontrol,72SPF SD male rats were subdivided randomly into normal control group（NC）,model group（MX）, positive controls group（PC）, Huata Qushi Huoxue Recipe high dosegroup（HD）、middle dose group（MD）、low dose group（LD）.Every group were12rats. Eatand fed in coops in IVC animal experimental lab. The control group fed with normal food,the other groups were fed with fat food （84%basic food+10%pig fat+5%yolk+1%cholesterol） everyday and were injected tetracycline （First dose of10mg/100g·rat weight,second to the last dose of7mg/100g rat weight.） every six day. From the second week,excepting the control group, every group were fed physiological saline and the othergroups were fed corresponding medicines once a day. The dose of polyenephosphatidylcholine capsule for PC group was0.0285g/100g·d^-1.The dose of HQHR forHD、MD、LD group were respectively5.04g/100g·d^-1、2.52/100g·d^-1、1.26/100g·d^-1.Therats were weighed every week and the dose adjusted according to the weight.7weeks later,all the groups were killed.Observation of serum ALT, AST, TG, TC, GLU level;The liverwere picked off and observed to wet weight and liver index, pathologic changes, liverhomogenate FFA content, immunohistochemical localization ADPN and AdipoR2proteindistribution; Liver tissue ADPN ELISA and real-time fluorescence quantitative PCR test, AdipoR2, AMPK, ACC, CPT-1protein gene expression.Results：1. Compared with the normal control group, model group and liver wet weight, liverindex, serum ALT, AST, TC, TG, GLU levels, liver homogenate FFA levels weresignificantly higher （P＜0.01）; ADPN/AMPK/ACC signal pathways in hepatic tissueADPN, AdipoR2, AMPK, CPT1protein and gene expression were significantly lower （P＜0.01）, the ACC protein and gene was significantly increased （P＜0.01）; Pathologicalchanges are moderately severe fatty change.2. Compared with model group: the liver index, serum ALT, AST, TC, TG,liverhomogenate FFA of the four treated groups were all decreased obviously the curativeeffects of HD groups are bes（tP＜0.01）.In ADPN/AMPK/ACC signaling pathway, the HDgroup can effectively increase ADPN, AdipoR2, AMPK, CPT-1protein and geneexpression, cut the ACC protein and gene, curative effect is better than the other treatmentgroup （P＜0.01）.3. Compared with PC group,the curative effect of HG group was better than PC group;MD Group decreased liver index, serum ALT, AST, TG, TC, GLU, liver homogenate FFA,curative effect was same to PC group （P＞0.05）,and generally better than LD group InADPN/AMPK/ACC signaling pathways, raised the ADPN, AdipoR2, AMPK, CPT1protein and gene expression, cut the ACC protein and gene. In pathways, MD group andPC group’s protein gene expression regulation is quite （P＞0.05）, and is better than LDgroup.4. ADPN, AdipoR2, AMPK, CPT1and rat weight, liver wet weight, liver index, ALT,AST, TG, TC, GLU, FFA, ACC present negative correlation. HQHR stand against NASHwas positively associated with its dose.Conclusion:1. This study adopts feeding with high lipid food and celio-injecting tetracycline,make NASH model in rats successfully.2. In the formation and development process of NASH, as the ADPN, AdipoR2,AMPK, CPT-1between group protein content decreased, rat weight, liver wet weight, liverindex, ALT, AST, TG, TC, GLU, FFA, ACC presents the rising trend.3. HQHR can effectively activate gene and protein expression of ADPN/AMPK/ACCpathway, make it recover to near normal levels. Reduce cholesterol and fatty acid synthesisraw material, increase the beta oxidation of fatty acid, reduce liver cell fatty deposits andinflammation of the liver. 4. From curative effect index or gene and protein expression of ADPN/AMPK/ACCpathway, the high dose of HQHR’s curative effect is better than that of polyenephosphatidylcholine（Yi Shan Fu）capsules.5. With the dose increasing, which indicate the curative effect relate to the dose inHQHR, Therapeutic effect of order is HD＞MD＞LD. The choice of clinical dosage isprovided for us by the important theory basis.