Study On The Mechanism Of Jianpi Tiaozhi Decoction In The Treatment Of NAFLD Based On FXR

Objective:In this study,we first obtained the core prescription(JPTZY)of the tutor in the treatment of non-alcoholic fatty liver disease(NAFLD)through data mining.On this basis,the core target and mechanism of the core prescription(JPTZY)in the treatment of NAFLD were predicted through the study of network pharmacology.Then through animal experiments to clarify the molecular mechanism of the core prescription(JPTZY)in the treatment of NAFLD.Methods:1.Data mining: 146 effective cases of NAFLD treated by mentors were collected and input into the cloud platform of ancient and modern medical records.Common syndrome,treatment and core prescription(JPTZY)were obtained through correlation and complex network analysis.2.Network pharmacological research: JPTZY and NAFLD targets were obtained through online databases such as TCMSP,OMIM and Gene Cards,the intersection of them was obtained by Venn.the core targets were obtained by STRING database and Cytoscape3.7.2 software,and the main pathways and targets were obtained by DAVID and KEGG.3.Experimental study: The NAFLD rat model was induced by high-fat diet for 12 weeks.41 rats were randomly divided into control group(n = 8)and model group(n = 33).At the end of the 12 th week,ONE rat were randomly selected from the model group and liver tissue pathological staining was performed to determine whether the model was successful or not.After success,the rats fed with high-fat diet were randomly divided into model group,lowdose traditional Chinese medicine group(5.84g/kg/d),medium-dose traditional Chinese medicine group(11.67g/kg/d)and high-dose traditional Chinese medicine group(23.34g/kg/d).Normal saline was given to normal group and model group according to the above-mentioned dose.The general condition and body weight of rats were recorded.After8 weeks of treatment,liver index was calculated according to the body weight of rats.The liver tissue HE staining and Oil Red O staining were observed.The levels of ALT,AST,TG,TC in serum and TC,TG,FFA in liver tissue were detected.The expression of FXR,SHP,SREBP-1c,PPAR?,CPT1 A,ACCm RNA was detected by q-PCR method,and the expression of FXR,SREBP-1c,PPAR? protein was detected by Western Blot method.Result:1.Data mining: In this study,146 patients were collected,and the results showed that most of the patients were turbid phlegm syndrome(44.52%).The drugs were mainly flat,slightly cold and warm,and the five flavors were sweet,bitter and pungent.The main meridian is spleen meridian.The main high-frequency drugs are invigorating the spleen,resolving phlegm and removing dampness,and it is concluded that the core pathogenesis of NAFLD is "spleen deficiency and phlegm deficiency".The core prescription of Jianpi Tiaozhi decoction was obtained by complex network analysis: Codonopsis pilosula,Coix seed,Atractylodes macrocephala,Poria,tangerine peel,Trichosanthes,alisma,Yinchen,lotus leaf,Patrinia,Polygonum cuspidatum,turmeric,licorice.2.Network pharmacology research: It is predicted by network pharmacology that 37 signaling pathways are mediated by 37 targets of Jianpi Tiaozhi decoction in the treatment of NAFLD.The core targets include: FXR,SREBP-1,HMOX1,AHR,FAS,IL6,IFNG,PPAR?,PPAR?,APOE,MTOR,VEGFA,GSTP1,HGF,IL1?,CTNNB1,APP,MMP1,CAT,MAPK8,LDLR,CCL2,mainly converge on NAFLD pathway,PPAR signaling pathway and FXR pathway.Finally,three targets closely related to lipid metabolism were screened: FXR,SREBP-1 and PPAR?.3.Experimental study: The NAFLD rat model was successfully established after 12 weeks of high-fat diet.(1)The body weight,liver index,serum ALT,AST,TC,TG and liver tissue TC,TG,FFA: in the model group were significantly higher than those in the blank control group(P< 0.01),and the above indexes in each treatment group were significantly lower than those in the model group(P < 0.05,P < 0.01).Compared with the low dose group,the middle and high dose groups decreased significantly and improved significantly(P < 0.05),and the high dose group decreased significantly and improved more significantly than the middle dose group(P < 0.05),and the improvement in the high dose group was significantly lower than that in the middle dose group(P < 0.05).(2)The color,shape,texture and touch of the liver were observed with naked eyes,and the liver of the model group was yellowish brown and obviously swollen.The blank control group was better than the model group and each drug group,and each drug group was better than the model group,especially the liver morphology of the high dose group was close to the blank control group.HE staining and oil red staining of liver tissue were observed under400 x light microscope.In the blank control group,the structure of hepatic lobule was intact,the hepatocytes were cord-shaped,radially arranged,the cell structure was clear,the cytoplasm was uniform,and the ratio of nucleus to cytoplasm was normal.In the model group,the structure of hepatic lobule was destroyed,cells swelled and deformed,showing diffuse steatosis,hepatocyte volume was enlarged,cytoplasm was loose,the boundary was unclear,balloon-like degeneration,punctate necrosis and inflammatory cell infiltration could be seen.Compared with the model group,after treatment with different doses of Jianpi Tiaozhi decoction,the steatosis and inflammation of liver in each group were improved to a certain extent,in which the number of lipid droplets in the high dose group was less and the cell structure was more regular.the liver tissue structure of some rats tends to be normal.(3)Compared with the blank control group,the FXR,SHP,SREBP-1c,PPAR?,CPT1 A and ACCm RNA: of the liver tissue in the model group were significantly abnormal(P <0.01),and compared with the model group,FXR,SHP,PPAR? and CPT1 Am RNA in each treatment group increased in varying degrees.SREBP-1c and ACCm RNA decreased in varying degrees,and the high-dose group was more statistically significant than the model group;the middle and high-dose groups were significantly different from the low-dose group;the improvement of FXR,SHP,ACC and CPT1 Am RNA in the high-dose group was significantly higher than that in the middle-dose group(P<0.05).(4)The expression of FXR,SREBP-1c and PPAR? protein in liver tissue: compared with the blank control group,the relative expression of FXR and PPAR? protein decreased significantly(P < 0.01),while the relative expression of SREBP-1c protein increased significantly(P < 0.01).Compared with the model group,the relative expression of FXR protein in the middle dose group increased significantly(P < 0.01),especially in the high dose group(P < 0.01).The relative expression of PPAR? protein did not increase significantly(P > 0.05),while the relative expression of SREBP-1c protein decreased significantly(P < 0.01).In the high dose treatment group,the relative expression of FXR and PPAR? protein increased significantly(P < 0.01),while the relative expression of SREBP-1c protein decreased significantly(P < 0.01).Compared with each treatment group,the expression of PPAR? and FXR protein in high dose group was significantly higher than that in low dose group(P < 0.05),and the relative expression of SREBP-1c protein in middle and high dose group was significantly higher than that in low dose group(P < 0.01).Conclusion:1."Spleen deficiency and insidious phlegm" is the important pathogenesis of NAFLD,and invigorating the spleen and resolving phlegm is the core treatment.The core prescription is Jianpi Tiaozhi decoction: Codonopsis pilosula,Coix seed,Atractylodes macrocephala,Poria,tangerine peel,Trichosanthes,alisma,Yinchen,lotus leaf,Patrinia,Polygonum cuspidatum,turmeric,licorice.2.Jianpi Tiaozhi decoction may reduce fat synthesis by activating FXR,inhibiting the expression of SREBP-1c and downstream target gene ACC,activating PPAR?,regulating CPT1 A gene transcription,promoting fatty acid ? oxidation,jointly regulating lipid metabolism,reducing liver lipid deposition and improving liver histopathology.