To further evaluate the role of hepatocyte-derived damage-associated molecules in inflammasome activation and potential cross-talk between hepatocytes and mononuclear cells, we tested whether PA-treated hepatocytes could induce inflammasome activation in inflammatory cells. Hepatocytes CP-673451 mouse were treated with PA for 6 hours and then were cultured in fresh media without PA. We found that these PA-free supernatants from PA-pretreated hepatocytes induced up-regulation of NALP3 (Fig. 7D) and IL-1β mRNA (Fig. 7E) in LMNCs; this suggests that FA-exposed hepatocytes can transfer activation to surrounding immune cells. The transmission of hepatocyte-derived
danger signals to mononuclear cells was dependent on caspase activation in hepatocytes;
this was suggested by the lack of LMNC activation with hepatocyte supernatants when ZVAD was added together with PA to hepatocytes (Fig. 7D,E). These results suggest that hepatocytes are the first target of FAs and produce inflammasome-mediated danger signals, which in turn activate macrophages in a caspase-dependent manner. The pathogenesis of steatosis and inflammation in NASH has been linked to multiple mechanisms,3 such as oxidative stress, mitochondrial damage in hepatocytes, and gut-derived LPS.7-9 Inflammation is a response to cellular injury and pathogens, and it is triggered by endogenous and exogenous danger signals, respectively. NALPs, the receptor components of the inflammasome, sense endogenous danger signals, which up-regulate the inflammasome, a multiprotein complex involved in caspase-1–mediated Roxadustat ic50 IL-1 cleavage. Inflammasome activation is typically a result of two signals via TLR activation MCE by exogenous or endogenous danger signals.10 Here we report several findings related to the novel role of inflammasome activation
in NASH. First, we describe up-regulation of the components of the NALP3 inflammasome (including NALP3, ASC, and pro–caspase-1) in mouse models of NASH and in human livers, and we demonstrate functional activation via caspase-1 activation and IL-1β production. Our data also suggest that inflammasome activation occurs in steatohepatitis and not in early steatosis in mice. Second, we report that although increased levels of circulating endotoxin likely contribute to inflammasome activation, exogenous LPS can amplify inflammasome activation and IL-1β secretion in steatohepatitis. Third, we demonstrate for the first time that inflammasome activation and IL-1β secretion occur in isolated hepatocytes in NASH. We reveal mechanistic insights into inflammasome activation and show that saturated FAs (but not unsaturated FAs) increase inflammasome expression and sensitize hepatocytes to IL-1β release by a second stimulus via TLR4 activation. Our novel data show that FAs not only up-regulate the inflammasome but also induce apoptosis and the release of danger signals in hepatocytes.