Selective LXR α intestinal activation reduces hepatic inflammation and fibrosis during the development of chronic liver injury

Background and Aims: The inflammasomes are cytoplasmic multiprotein complexes that are responsible for the activation of inflammatory reactions. They mediate the cleavage and activation of caspase-1 and IL-1b that in turn leads to a complex network of cellular reactions that at the end initiate local and systemic inflammation [1]. We have shown that the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) is virtually absent in untreated cultured hepatocytes while stimulation with lipopolysaccharides (LPS) results in strong activation of NLRP3 expression in these cells [2]. In the present study we investigated the impact of NF-uB signalling on NLRP3 activation in primary hepatocytes. Methods: Murine primary hepatocytes were isolated according to the collagenase method [3]. The NF-uB activation inhibitor QNZ and the adenoviral expression vector Ad5-CMV-IuB(S32A/S36A) expressing a hemagglutinin-tagged human superrepressor of NF-uB [4] were used to block NF-uB signalling. After LPS challenge, the expression of TNF-a, IL-1b and NLRP3 was analysed by qRTPCR, Western blot and immunohistochemistry. The influence of NF-uB signalling on NLRP3 expression was further investigated in a novel murine hepatocytic cell system that can be Cre-dependently depleted for the NF-uB essential modulator (NEMO). Results: We found that QNZ blocks LPS-induced expression of TNF-a, IL-1b and NLRP3 as demonstrated by qRT-PCR, Western blot and immunohistochemistry. Even more, the basal expression that was observed in uninfected control cells or in cells that were infected with a control virus (Ad5-CMV-Luc) was abrogated. Likewise, the superrepressor of NF-uB prevents expression of NLRP3. In cells that were depleted for NEMO (and Caspase 8), we could confirm that there is a close link of NF-uB activity and NLRP3 expression. As a consequence, the expression of Lipocalin 2 representing a biomarker of hepatic inflammation was blunted suggesting that NF-uB activity plays an important role in mediating inflammasome activity. Conclusions: We conclude that NF-uB signalling is a necessary prerequisite for proper activation of the NLRP3 inflammasome in primary hepatocytes. Our data further suggests that targeting the NF-uB pathway will be potentially therapeutic useful to block hepatic inflammasome activity.