Unveiling a novel macrophage-driven mechanism for obeticholic acid-induced liver injury in cholestatic: targeting NLRP12Arg185 fuels Relb-TNFSF14-mediated ductular reaction

For individuals with Primary Biliary Cholangitis (PBC) who are either intolerant or nonresponsive to ursodeoxycholic acid (UDCA), obeticholic acid (OCA) is authorized as a therapeutic treatment. Nevertheless, its clinical application has been limited due to the occurrence of several adverse effects. We employed the bile duct ligation (BDL) model to simulate cholestatic conditions observed in patients, revealing that high doses of OCA exacerbated hepatic inflammatory infiltration and fibrosis, accompanied by a marked augmentation in the extent of ductular reaction (DR) within BDL. Our findings mentioned that macrophages played a critical part in mediating OCA-induced aggravation of DR. OCA significantly upregulated the non-canonical NF-κB Relb signaling in macrophages, thereby contributing to proliferation of cholangiocytes. Cellular thermal shift assay (CETSA), surface plasma resonance (SPR) and liquid chromatography mass spectrometer (LC-MS) analysis showed that OCA directly bond to Nucleotide-binding oligomerization domain-like receptor (NLR) pyrin domain (PYD)-containing protein 12 (NLRP12), leading to a reduction in its protein level. Further, activity-based protein profiling (ABPP) method identified the Arg185 residue of NLRP12 as the binding site for OCA, elucidating the mechanism by which OCA downregulates NLRP12 protein expression. This interaction diminished the inhibitory effect of NLRP12 on the Relb pathway. Collectively, our study delineated the NLRP12^Arg185-Relb-TNFSF14 axis in macrophages as a key mechanism underlying OCA-induced hepatotoxicity.