Ammonium tetrathiomolybdate attenuates acetaminophen-induced acute liver failure by inhibiting the TRPV4/Calcium/NF-κB signaling pathway

Acute liver failure (ALF), characterized by fulminant hepatic necrosis and excessive inflammatory-oxidative cascades, remains a critical clinical challenge with limited therapeutic options. This study investigates the therapeutic potential of ammonium tetrathiomolybdate (ATTM)-a copper-chelating agent with multimodal anti-inflammatory and antioxidant properties-in acetaminophen (APAP)-induced ALF. Utilizing APAP-challenged C57BL/6J mice, we demonstrated that ATTM administration, whether prophylactic or delayed by 2 h post-exposure, significantly attenuated hepatotoxicity, as evidenced by reduced histopathological damage and improved survival rates. These therapeutic effects were further confirmed in AML12 hepatocytes, thereby reinforcing the observed in vivo findings. RNA sequencing revealed that calcium signaling is the predominant pathway modulated by ATTM. Subsequent mechanistic validation identified Transient Receptor Potential Cation Channel Subfamily V Member 4 (TRPV4)-mediated calcium influx as the critical therapeutic target. ATTM suppressed TRPV4-dependent calcium mobilization, thereby inhibiting the sequential phosphorylation of NF-κB pathway components in both murine liver tissue and AML12 cells. Crucially, TRPV4 agonism via RN-1747 reversed the hepatoprotective effects of ATTM, thereby confirming the centrality of this axis in mediating ATTM's therapeutic actions. These findings establish ATTM as a novel modulator of the TRPV4/calcium/NF-κB signaling cascade, capable of interrupting inflammatory-oxidative loops at multiple nodes. Our work not only elucidates a previously unrecognized mechanism for copper chelators in ALF management but also positions ATTM as a promising therapeutic candidate warranting clinical translation.