BACKGROUND & AIMS: Acetaminophen (APAP) overdose is the leading cause of acute liver failure, with oxidative stress being a critical factor in this process. Glutathione (GSH) plays a vital defensive role. Activation of nuclear factor erythroid 2 like 2 (Nrf2) pathway mitigates APAP-induced liver damage by promoting GSH biosynthesis and enhancing drug detoxification. Although the role of B cell leukemia/lymphoma 3 (Bcl3) in regulating inflammatory responses, cellular oncogenesis, and immune balance is well-documented, its function in APAP-induced liver injury remains unclear. METHODS: We employed liver-specific Bcl3 knockout (Bcl3 RESULTS: Our study reveals a significant upregulation of Bcl3 expression in the livers of male mice following APAP administration, suggesting Bcl3's potential involvement in this pathological process. In Bcl3 CONCLUSION: We uncovered a previously uncharacterized role of Bcl3 in APAP-induced liver injury, emphasizing the role of the Bcl3-Nrf2 axis in oxidative stress-related liver damage.