Gut Microbiota Mediates the Protective Effects of Andrographolide Inhibits Inflammation and Nonalcoholic Fatty Liver Disease(NAFLD) in High-Fat Diet induced ApoE(-/-) Mice

Mechanisms relating the gut bacteria to Nonalcoholic Fatty Liver Disease (NAFLD) have been proposed containing the dysbiosis-induced dysregulation of hepatic lipid metabolism that allows for the translocation of microbial components and leads to hepatic inflammation and steatosis. Andrographolide (AG) regulates inflammation mediated by NF-κB pathway which also play a key role in reduction of inflammation and fibrosis in experimental nonalcoholic steatohepatitis (NASH), yet the mechanisms linking this effect to gut microbiota remain obscure. Here we show that ApoE knockout (Apoe -/-) mice fed a high-fat diet (HFD) supplemented with AG regulates levels of biochemical index and inflammatory cytokines associated with gut microbe. Moreover, HEPG2 cells induced by ox-LDL were used as validation in vitro. H&E staining and Oil-Red staining were respectively used for tissue and cells morphology. Gut microbiota were examined by 16S rRNA sequencing. Expression of NF-κB, C/EBPβ and PPAR-γ in liver and HEPG2 cells were detected by western blot and qRT-PCR. The results showed, among others, that AG alleviate hepatic steatosis and fat content in HEPG2 cells, while it induced decreased levels of Bacteroides, and increased levels of Faecalibaculum, Akkermansia. We further identified that inhibition of NF-κB/C/EBPβ/PPAR-γ pathway of hepatic steatosis model in vivo and vitro by AG also contributes to prevention of HFD-induced inflammation and dislipidemia. Importantly, as result of pearson correlation, Bacteroides may be the most relevant one fundamentally involved in the mechanism of AG attenuates NAFLD. Together, our findings uncover an interaction between AG and gut microbiota as a novel mechanism for the anti-NAFLD effect of AG acting through prevention of microbial dysbiosis, dislipidemia and inflammation. Importance HFD due to gut microbial dysbiosis is a major contributor to the pathogenesis of dislipidemia and inflammation, which primarily mediates the development of NAFLD. A treatment strategy to reduce both dislipidemia and inflammation appears to be an effective approach for addressing the issue of NAFLD. Andrographolide (AG) is the major effect component in traditional Chinese medicine Chuan-xin-lian (Andrographis). Little is known about the role of gut microbiota in the anti-NAFLD effect of AG. 16S rRNA gene sequencing revealed that AG significantly decreased Bacteroides and increased Faecalibaculum, Akkermansia. By using vivo and vitro experiment, we prove that gut microbiota plays a key role in AG-induced protective against high-fat-diet-induced dislipidemia and inflammation. Moreover, NF-κB/C/EBPβ/PPAR-γ pathway inhibition was partially involved in the beneficial effect of AG. Together, these data suggest that the gut microbiome is a critical factor for the anti-NAFLD effects of AG.