AIMS: Acute liver failure (ALF) is marked by extensive inflammation and immune dysregulation, which are closely associated with neutrophil infiltration and NETosis. However, the specific mechanisms that drive NETosis in ALF remain poorly understood. MATERIALS AND METHODS: We employed flow cytometry, western blot, qRT-PCR, and cf-DNA assay to investigate the link between NETosis and ALF. The role of HDAC6-mediated deacetylation of histidine triad nucleotide-binding protein 2 (Hint2) was assessed, along with the effects of lentiviral vector-based overexpression and knockdown of Hint2 on mitochondrial function and NETosis. Additionally, CO-IP, IF, protein docking analysis, mCa KEY FINDINGS: Our study demonstrated that Hint2 undergoes HDAC6-mediated deacetylation, disrupting mitochondrial dynamics and triggering NETosis during ALF. Furthermore, MICU1 bridges Hint2 and NETosis by regulating mCa SIGNIFICANCE: Our findings recognized the HDAC6/Hint2/MICU1 axis as a novel pathway in neutrophils, the inhibition of which intercepts mCa