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.