| Background The global prevalence of nonalcoholic fatty liver diseaseis have been increasing,which result in severe clinical and economic burden.It has become a global health problem.At present,there have not one drug which can be completely confirmed treatment effect and be supporedt by clinical guidelines.Development of new drugs is also a trend of the health.The gut-liver axis has become a new therapeutic method.Objective Based on gut-liver axis,the research aim to explore the Tianhuang formula effects on glucolipid metabolism disorder because of intestinal barrier function and bile acid metabolism in NAFLD mice.Metheds 1.C57/BL male 6J mice were fed high fat diet for 19 weeks to build a model of NAFLD.According to stratified random group,the mice were random Ly divided into normal control group,model group,positive drug(atorvastatin calcium),low dose and high dose of Tianhuang formula,and then by intragastric adiministrated for 11 weeks.2.Test the weight of mice,liver index,muscle weight,and HE staining in the liver and epididymal adipose tissue,oil red O in the liver.3.Fasting blood glucose,insulin,plasma TG,plasma TC,plasma HDL-C,plasma LDL-C were tested in accordance with the kit instructions.The lipid metabolism genes of perlipin2,CD36,C/EBP α,FAS,SCD1,PPARα,lipin1 in the liver were detected by RT-PCR.4.Pasma LPS was detectd by ELISA.The mRNA and protein levers of TLR4 and NF-κB were respectively detected by RT-PCR and Western Blot in the liver.The expression of inflammation related genes were detectd by RT-PCR in the liver.5.16 S r DNA sequencing were used to detect the effects on the OTUs,diversity and analysis some metabolic related microbiota abundance.6.The m RNA and protein levers of tight juncation related molecules such as Calduin-7,occludin,Calduin-1,Calduin-2,Calduin-3,Calduin-4,Calduin-18,were respectively detected by RT-PCR,western blot and immunofluorescence in the small and large intestine.7.The expression of inflammation related genes(TNFα,IL-6,IL-1β,IL-10)were detectd by RT-PCR in the small and large intestine.8.The expression of bile acide related genes(CYP7α1,CYP7β1,CYP8α1,FXR,FGFR4,SHP,FGF15)were detectd by RT-PCR in the liver and gut.Results 1.During the modeling period,the body weight of high fat diet(HFD)fed mice increased continuously.Fasting blood glucose,insulin,HOMA-IR,plasma TG and TC in 19 weeks HFD-fed mice significantly increased.2.Tian Huang formula reduced liver weight and spleen index,the liver lipid droplets decreased in HE staining test and oil red O staining test,and the adipocyte size in e WAT decreased significantly.3.Compared with HFD-fed mice,Tian Huang formula significantly reduced fasting blood glucose,plasma insulin,HOMA-IR,plasma TG,plasma TC,plasma LDL-C and increased plasma HDL-C.In the liver,Tian Huang formula significantly decreased TG and TG content and decreased m RNA level of perlipin2,C/EBPα,CD36,FAS,SCD1 and increased m RNA level of PPARα,ACOX-1 and lipin1.4.Tian Huang formula significantly decreased the levels of plasma LPS,significantly reduced the m RNA expression of TLR4,NF-κB,TNF-α,IL-6,IL-1β in the liver.5.Tian Huang formula changes in the microbial diversity and composition.This remodelling increased Firmicutes/Bacteroides ratios and A.muciniphlia.6.Tian Huang formula increased the genes expression of claudin-7,occludin,ZO-1,GLP2,GCG and increase the protein expression of claudin-7,occludin,ZO-1,Claudin-1,Claudin-2,Claudin-3,Claudin-4 in the small and larger intestine.7.Tian Huang formula significantly reduced the genes expression of TNFα,IL-6,IL-1β and increased the expression of IL-10 in the small intestine and large intestine.8.Tian Huang formula regulate bile acid metabolism markers in the liver and gut(CYP7A1,CYP7B1,CYP-8B1,FXR,FGFR4,SHP,FGF15).Conclusion 1.Tian Huang formula improved the lipid deposition,insulin resistance,lipid metabolism,endotoxemia and hepatic inflammation in NAFLD mice.2.Its mechanism may be due to the effects on improving intestinal biological barrier,mechanical barrier,immune barrier and bile acid metabolism. |
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