SUMMARY Increased translocation of intestinal bacteria is a hallmark of chronic liver disease and contributes to hepatic inflammation and fibrosis. Here we tested the hypothesis that the intestinal microbiota and Toll-like receptors (TLRs) promote hepatocellular carcinoma (HCC), a long-term consequence of chronic liver injury, inflammation and fibrosis. Hepatocarcinogenesis in chronically injured livers depended on the intestinal microbiota, and TLR4 activation in non-bone marrow-derived resident liver cells. TLR4 and the intestinal microbiota were not required for HCC initiation but for HCC promotion, mediating increased proliferation, expression of the hepatomitogen epiregulin, and prevention of apoptosis. Gut sterilization restricted to late stages of hepatocarcinogenesis reduced HCC suggesting that the intestinal microbiota and TLR4 represent therapeutic targets for HCC prevention in advanced liver disease.

Background & Aims Activated hepatic stellate cells (HSCs), the main fibrogenic cell type of the liver, undergo apoptosis after cessation of liver injury, thereby contributing to the resolution of liver fibrosis. In this study, we investigated whether HSC deactivation constitutes an additional mechanism of liver fibrosis resolution. Methods HSC activation and deactivation were investigated by single cell PCR and genetic tracking in transgenic mice expressing tamoxifen-inducible CreER under control of the endogenous vimentin promoter (VimCreER). Results Single cell quantitative PCR demonstrated activation of virtually the entire HSC population in fibrotic livers, and a gradual decrease of HSC activation during fibrosis resolution, indicating deactivation of HSCs. VimCreER marked activated HSCs, demonstrated by a 6- to 16-fold induction of a membrane-bound green fluorescent protein (mGFP) Cre-reporter following injection of carbontetrachloride (CCl4) in both liver and isolated HSCs, and a shift in localization of mGFP-marked HSCs from perisinusoidal to fibrotic septa. Tracking of mGFP-positive HSCs revealed the persistence of 40–45% of mGFP expression in livers and isolated HSCs 30–45 days after cessation of CCl4, despite normalization of fibrogenesis parameters, thereby confirming reversal of HSC activation. After fibrosis resolution, mGFP expression was observed again in desmin-positive perisinusoidal HSCs; no mGFP expression was detected in hepatocytes or cholangiocytes, thereby excluding mesenchymal-epithelial transition. Notably, reverted HSCs remained in a primed state, with higher responsiveness to profibrogenic stimuli. Conclusion In mice, reversal of HSC activation contributes to the termination of fibrogenesis during fibrosis resolution but results in higher responsiveness of reverted HSCs to recurring fibrogenic stimulation.

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