BACKGROUND

Acute liver failure (ALF), characterized by inflammation-mediated hepatocyte dysfunction, poses a significant global health burden. Although neutrophils and macrophages are central to driving inflammatory responses in ALF, the molecular mechanisms governing IL-36-driven inflammation in toll-like receptor 3 (TLR3)-mediated NETosis and macrophage activation remain poorly understood. This study systematically delineates wherein TLR3 activation induces IL-36 hyperactivation and NETosis, which cooperatively trigger macrophage activation to accelerate ALF progression.

METHODS

First, the role of IL-36 signaling was evaluated in the TLR3-provoked mouse peritoneal macrophages (MPMs) and murine bone-derived macrophages (BMDMs) in vitro. Further, the murine model of ALF was established with polyinosinic-polycytidylic acid (Poly(I:C))/D-GalN and acute ethanol gavage in Il1rl2-deficient mice generated by lentivirus-mediated shRNA, and the released NETs were evaluated with immunoblotting and immunofluorescence staining.

RESULTS

We observed Poly(I:C), the specific ligand of TLR3, when combined with extracellular ATP, provoked the IL-36 signaling and the release of IL-1β in macrophages. In contrast, Il36g deficiency reduced the secretion of proinflammatory cytokines, such as IL-1β and HMGB1, upon macrophage activation. While Poly(I:C)/D-GalN or binge ETOH-induced ALF in mice was accompanied by the enhanced level of IL-36γ and NET formation, IL-36R knockdown reversed the liver damage by decreasing the expressions of proinflammatory cytokines and chemokines, reducing the lipid accumulation and preventing the NET formation as well.

CONCLUSION

Our study demonstrates that TLR3-mediated NET formation was enhanced in ALF, and this process was tightly regulated by IL-36 signaling. These findings suggest that targeting IL-36 represents a potential therapeutic target for modulating inflammation in ALF.