Mechanisms Of Fibroblast Growth Factor 19 Regulation Of PPARα Transcriptional Expression

Non-alcoholic fatty liver disease(NAFLD)is a major global public health problem.Its pathogenesis is not yet fully understood.Excessive accumulation of fat in hepatocytes,leading to hepatocellular damage and triggering an inflammatory response in the liver,are key features of NAFLD.Peroxisome proliferation activated receptor alpha(PPARα),a key regulator of lipid oxidative catabolism promotion in the liver,has great potential in the treatment of NAFLD.Fibroblast Growth Factor 19(FGF19)is mainly expressed in the human intestine and is a major regulatory protein regulated by Farnesoid X Receptor(FXR)and involved in bile acid homeostasis.FGF15 is its mouse homologue.Previous studies showed that the levels of PPARα and its downstream target genes were significantly upregulated in the liver of FGF15 transgenic mice,and that FGF15 may regulate the transcriptional expression of PPARα through its downstream ERK1/2 and JNK1/2 signaling pathways,or affect the level and composition of bile acids in the body,and indirectly regulate the transcriptional expression of PPARα through bile acid signaling molecules.FGF19,as a human homolog of FGF15,may also regulate h PPARα in the human body.On this basis,this study investigated the regulatory role of FGF19 and bile acids on the transcriptional expression of human PPARα gene.As a novel discovery,this mechanism can provide different ideas for the prevention and treatment of NAFLD.In this study,we investigated the role of FGF19 or bile acids in regulating the transcriptional expression of h PPARα gene through its 5’ end promoter region by dual luciferase reporter gene assay,bioinformatics analysis,recombinant PCR targeted mutagenesis,and immunoblotting in human hepatocellular carcinoma cells(Hep G2),as well as the signaling pathways and cis-acting elements involved.The main findings are as follows:1.PETDute-FGF19 recombinant protein expression vector was constructed,successfully induced in E.coli SHffle-T7-B and isolated and purified to obtain FGF19 recombinant protein,which was verified to activate ERK1/2 and JNK1/2 signaling pathways in Hep G2 cells.2.FGF19 recombinant protein failed to induce transcriptional activity of the h PPARα promoter fragment in Hep G2 cells,but SP600125,a selective small molecule inhibitor of the JNK1/2 signaling pathway,inhibited the transcriptional activity of the h PPARα promoter fragment,and this inhibition was partially mediated by a predicted c Jun binding site in the promoter fragment,suggesting that the JNK1/2 signaling pathway is positively associated with h PPARα gene transcription,and FGF19 may regulate h PPARα transcriptional expression in vivo through activation of the JNK1/2signaling pathway.3.Farnesoid X receptor(FXR),a physiological receptor for bile acids,significantly induced the transcriptional activity of the h PPARα gene promoter fragment after overexpression of h FXR in Hep G2 cells and activation by GW4064,indicating that h FXR regulates the transcriptional expression of h PPARα.further studies in the h PPARα promoter fragment validated an FXR binding site.4.The regulatory effects of bile acids on ERK1/2 and JNK1/2 signaling pathways were investigated in Hep G2 cells.Deoxycholic acid(DCA)activated ERK1/2 signaling pathway and inhibited JNK1/2 signaling pathway,lithodeoxycholic acid(LCA)activated ERK1/2 signaling pathway and ursodeoxycholic acid(UDCA)inhibited ERK1/2 signaling pathway.In contrast,chenodeoxycholic acid(CDCA)and glycodeoxycholic acid(GDCA)did not regulate ERK1/2 and JNK1/2 signaling pathways.5.In the dual luciferase reporter gene assay,LCA inhibited the transcriptional activity of the promoter fragment of the h PPARα gene,and the sites of action were mainly located in the fragment from-744 bp to-128 bp,and the specific mechanism needs further study.CDCA,DCA,GDCA,and UDCA did not regulate the transcriptional activity of the promoter fragment of the h PPARα gene.In conclusion,JNK1/2 signaling pathway,FXR,and LCA regulate the transcriptional expression of h PPARα through the 5’ promoter region of h PPARα.The mechanism by which FGF19 regulates h PPARα transcriptional expression in vivo may act by activating the JNK1/2 signaling pathway on the one hand,and by affecting the level and composition of bile acids on the other.