Suppression of autophagy sensitizes Kupffer cells to endotoxin

Aim: Recent evidence suggests that protein degradation system autophagy is implicated in a component of innate immunity. We report here that suppression of autophagy in Kupffer cells due to hepatic steatosis enhances an inflammatory response to endotoxin. Methods: Kupffer cells were isolated from C57BL/6J mice fed chow diet (control) or high-fat diet (HFD) for 12 weeks, liver-specific autophagy-deficient mice (Atg7F/F:Mx1-Cre) and wild-type mice (Atg7F/F). Kupffer cells were incubated with 100 ng/mL lipopolysaccharide (LPS). The concentration of tumor necrosis factor (TNF)-a in media was measured by enzyme-linked immunoassay. Expression of Toll-like receptor (TLR)4, I?B kinase (IKK)-a/beta, p38, p62 and LC3 in Kupffer cells was evaluated by western blot analysis. Results: Incubation with LPS increased LC3-II expression of Kupffer cells from control mice; however, an increase in LC3-II expression due to LPS was suppressed in Kupffer cells from HFD mice. Moreover, both p62 expression and TNF-a production in Kupffer cells from HFD mice was higher than control mice. On the other hand, LPS exposure increased TNF-a production from autophagy-deficient Kupffer cells more than wild type. There was no significant difference in expression of TLR4 between wild and autophagy-deficient Kupffer cells. Nevertheless, activation of p38 or IKK in Kupffer cells due to LPS was augmented by autophagy deficiency. The addition of the p38 inhibitor SB203580 attenuated TNF-a production in both wild and autophagy-deficient Kupffer cells. Conclusion: These results suggest that suppression of autophagy observed in Kupffer cells from steatotic liver sensitizes to endotoxin. In conclusion, suppression of autophagy may play a pivotal role on progression of NAFLD.