Abstract Primary biliary cirrhosis (PBC) is an organ-specific autoimmune liver disease characterized by the presence of anti-mitochondrial antibodies (AMA) and the destruction of small intrahepatic bile ducts with portal inflammation. In previous studies, we reported that both CD1d expression and the frequency of invariant natural killer T (iNKT) cells were increased in the liver of patients with PBC. To define a specific role of iNKT cells in the pathogenesis of PBC, particularly early events, we investigated the function of hepatic iNKT cells in our TGF-β receptor II dominant-negative (dnTGFβRII) mouse model of PBC. We report here that these dnTGFβRII mice demonstrate a massive increase of hyperactive iNKT cells within the hepatic tissues. We generated CD1d−/−and CD1d+/− dnTGFβRII mice and performed a comparative study of liver immunopathology, demonstrating that the CD1d−/−dnTGFβRII mice, lacking iNKT cells, exhibit significantly decreased hepatic lymphoid cell infiltrates and milder cholangitis compared to CD1d+/−dnTGFβRII mice. Interestingly, there was a significant increase in the production of IFN-γ in hepatic iNKT cells activated by α-galactosylceramide (α-GalCer) in young but not older dnTGFβRII mice, suggested an age dependent role of iNKT cells. These data demonstrate that iNKT cells in dnTGFβRII mice are a critical factor in liver injury.