Role Of The TLR4-TRIF-IFN? Pathway In Kupffer Cell Induced Nonalcoholic Fatty Liver Disease

Aim:To investigate the role of TLR4-TRIF-IFN? pathway and IL-1RA in nonalcoholic steatohepatitis(NASH)by observing the expression of TLR4-TRIF-IFN? and interleukin1 receptor antagonist(IL-1RA)at different stages of nonalcoholic fatty liver disease(NAFLD)induced by a methionine /choline-deficient(MCD)diet in mice.Methods:10-week-old male C57BL/6J mice were randomly divided into two groups for standard chow(SC)or MCD diet.At the end of experimental 1week,4weeks and 8weeks,measurements were taken for the NASH-related parameters of hepatosomatic index(HIS),ALT,AST,IL-1RA.Histology of liver were evaluated after HE staining,oil red O staining.Immunohistochemical staining were used to detect Kupffer cells(KCs)in liver.Expression of TLR4,TRIF,IFN?,F4/80,TNF-?,and MCP-1 m RNA levels from liver tissue(RT-PCR)were also detected.Expression of IL-1RA m RNA levels from liver tissue and KCs were also detected.Results:(1)The mice in the MCD group showed significantly higher HSI,Which were compared to the mice in the SC group at the Weeks 4 and 8(P <0.05).The HSI in mcd group was increased gradually until Week 8.As compared to the mice in the SC group at the Weeks 1,4 and 8,the mice fed with MCD showed significantly higher serum levels of ALT,AST and IL-1RA(P <0.01,P <0.05,P <0.05).IL-1RA levels in the MCD group decreased gradually until Week 8.(2)The liver tissues of mcd fed mice exhibited microvesicular steatosis as early as Week 1.Furthermore,macro-and microvesicular steatosis,a little of hepatocellular ballooning,and Inflammatory cells can be observed after 4 weeks feeding of the MCD diet.When mcd diet feeding was extended to 8 weeks,the pathological features,including hepatocellular steatosis,hepatocellular ballooning and lobular inflammation,became Obvious.Notable liver fibrosis was also observed with the excessive accumulation of collagen,as shown in liver sections by haematoxylin and eosin(H&E)staining.(3)As compared to the mice in the SC group at the Weeks 1,4 and 8,the mice in the MCD group were significantly change in hepatic m RNA expression of F4/80,MCP-1 and TNF-?(p <0.05 for all).Hepatic m RNA expression of MCP-1 and TNF-? increased gradually until Week 8.(4)Hepatic TLR4 m RNA expression in the mcd group was significantly higher compared with the SC group at the Weeks 1,4 and 8(p <0.05),and increased gradually until Week 8.Hepatic TRIF m RNA expression from mcd group increased at week 1 and decreased at week 8 compared with SC group(P <0.05).As compared to the mice in the SC group at the Weeks 1 and 4 the mice fed with MCD showed significantly higher hepatic m RNA expression of IFN? and IL-1RA.Hepatic IFN? m RNA expression was markedly upregulated at Week 4 and then declined at Week 8.Hepatic IL-1RA m RNA expression decreased gradually until Week 8.Expression of IL-1RA m RNA from Kupffer cells peaked at Week 4 and then declined at Week 8.Conclusions: Insufficient activation in TLR4-TRIF-IFN? pathway and insufficient production of anti-inflammatory cytokine IL-1RA may be a key factors that progress NAFL to NASH in mice.