1A-D) after acetaminophen challenge. Moreover, few splenic γδ T cells expressed IL-23 receptor but most of them expressed CD27, which were prone to producing IFN-γ (Supporting Fig. 1E,F).20 Together, our results demonstrate that hepatic γδ T cells are the major Lumacaftor solubility dmso producers of IL-17A during acetaminophen-induced liver inflammation. Meanwhile, after depletion of γδ T cells, liver injury

was attenuated (Fig. 4; Supporting Fig. 2). ALT and bilirubin levels were reduced (Fig. 4A,B). The necrotic hepatic areas were also reduced (Fig. 4C). The survival rate of γδ T cell-depleted mice was markedly improved (Fig. 4D), with a decreased content and total number of CD11bhiLy-6G+ neutrophils in the liver (Fig. 4E,F). The attenuated liver injury, decreased neutrophils

in the liver, and improved survival ratio were also observed in TCRδ−/− mice compared to that of age-matched control mice (Supporting Fig. 2). Thus, hepatic γδ T cells are critical during acetaminophen-induced, damage-associated IL-17A-mediated liver inflammation. INK 128 cell line To investigate the role of IL-23 in the production of IL-17A by hepatic γδ T cells, IL-23 in the sera and liver was measured. Serum IL-23 significantly increased and peaked at 12 hours after acetaminophen challenge (Fig. 5A), and p40, one subunit of IL-23, also increased and peaked at 12 hours (Fig. 5A). In the liver, p19 and p40, two subunits of IL-23, also increased (Fig. 5B). To further determine whether IL-23 is required for the production of IL-17A, we neutralized its function using an anti-IL-23p19 antibody or p40-deficient mice (Fig. 5C). Serum IL-17A significantly decreased after neutralizing IL-23 or using p40-deficient mice. Moreover, infiltration of neutrophils into the liver was significantly ameliorated (Fig. 5D-F). Meanwhile, the liver injury was also reduced in the p40-deficient mice compared to the aged-matched control mice (Supporting Fig. 3). Taken together, these results

show that IL-23 is important for the production IL-17A by γδ T cells after acetaminophen challenge. To confirm the role of IL-23 in the generation MCE of IL-17A-producing γδ T cells, we stimulated hepatic γδ T cells with exogenous IL-23 in vitro. After stimulation for 48 hours, the percentage of IL-17A-producing γδ T cells was significantly increased (Fig. 6A,B). After a second cycle of stimulation with PMA for 5 hours, the percentage of IL-17A-producing γδ T cells further increased to 32.3% (Fig. 6A). Supernatant IL-17A from total hepatic lymphocytes or purified γδ T cells was increased after stimulation with IL-23, which was further enhanced by IL-23+IL-1β stimulation (Fig. 6C,D). Therefore, the in vitro experimental data demonstrate that IL-23 is required for the production of IL-17A from γδ T cells. To understand whether macrophages mediate the production of IL-23, we inhibited macrophages, including Kupffer cells, with GdCl3.