Toll/IL-1 receptor domain-containing adaptor protein plays a crucial role in macrophage-mediated hepatic stellate cell activation in alcoholic liver disease.

BACKGROUND: Alcoholic liver disease (ALD) remains a significant global health issue, marked by chronic liver injury and progressive accumulation of extracellular matrix (ECM), ultimately leading to fibrosis. A key driver of this fibrotic process is the activation of hepatic stellate cells (HSCs), largely influenced by inflammatory signals from immune cells-particularly macrophages. This study explores the specific signalling mechanisms in macrophages that contribute to HSC activation in the context of alcohol exposure, aiming to identify new therapeutic targets. METHODS: To mimic the disease condition, THP-1-derived macrophages were stimulated with ethanol and lipopolysaccharide (LPS). Cytokine expression was quantified by RT-qPCR, while the activation of TIRAP, MAPKs, and fibrotic markers in THP-1-derived macrophages and LX-2 cells was evaluated by western blotting and immunofluorescence staining. RESULT: Activation was indicated by increased expression of pro-inflammatory and fibrogenic factors, including IL-1β, TNF-α, IL-6, TGF-β, and PDGF-α. Signalling analysis revealed heightened phosphorylation of TIRAP (Toll/IL-1 receptor domain-containing adaptor protein), along with activation of downstream MAPK pathways (p38, ERK, JNK) and NF-κB. The conditioned media from these activated macrophages was applied to HSC cultures, resulting in elevated levels of fibrotic markers such as α-smooth muscle actin (α-SMA) and collagen, confirming HSC activation. Importantly, silencing TIRAP in macrophages significantly reduced the expression of these markers in HSCs, suggesting a key role for TIRAP in driving fibrogenesis. CONCLUSION: Overall, these findings underscore the importance of TIRAP- mediated signalling in macrophages as a central mechanism in alcohol- induced liver fibrosis, and propose macrophage TIRAP as a promising target for therapeutic intervention in ALD.