6, 7), suggesting that the accumulation of neutrophils contributes to the exacerbated liver injury observed in α-Galcer-treated IFN-γ−/− and STAT1−/− mice. IFN-γ−/− mice had lower levels of hepatic expression of IL-4 compared to WT mice after α-Galcer injection (Supporting Fig. 7), suggesting that IFN-γ is required for the production of IL-4. However, this unlikely attributes to IFN-γ prevention
of hepatic neutrophil infiltration because IL-4 promotes hepatic neutrophil accumulation (see above). Our further findings indicate that IFN-γ attenuates hepatic neutrophil accumulation by inducing neutrophil apoptosis after α-Galcer injection, as neutrophil apoptosis was suppressed in IFN-γ−/− mice (Fig. 6).
Mechanistic studies suggest that the proapoptotic effect of IFN-γ is mediated by the induction of several proapoptotic genes by way of a STAT1-dependent mechanism 3-deazaneplanocin A order (Fig. 7F). Collectively, these findings suggest that IFN-γ stimulates the expression of proapoptotic genes in neutrophils by Mdm2 antagonist way of a STAT1-dependent mechanism, thereby playing an important role in preventing hepatic neutrophil accumulation in α-Galcer-induced liver injury. In addition to their opposing roles in the control of hepatic neutrophil accumulation, IL-4 and IFN-γ have been shown to inversely control NKT cell proliferation in vitro. During the course of our studies, we observed the percentage and total number of liver iNKT cells in WT, IL-4−/−, IFN-γ−/−, and IL-4−/−IFN-γ−/− dKO mice MCE公司 were comparable before α-Galcer injection. After α-Galcer injection, liver iNKT cells rapidly disappeared within 24 hours. This disappearance was similar among these four strains of mice (data not shown). These findings suggest that the differences in hepatic neutrophil accumulation 3 hours post-α-Galcer injection
among WT, IL-4−/−, IFN-γ−/−, and IL-4−/−IFN-γ−/− dKO mice were not caused by the changes in iNKT cells at the early timepoints after α-Galcer injection. Additionally, expression of activation markers (CD11b and CD62L) and production of reactive oxygen species (ROS) were comparable in neutrophils from α-Galcer-treated WT, IL-4−/−, and IFN-γ−/− mice (Supporting Fig. 8), suggesting IL-4 and IFN-γ regulate hepatic neutrophil accumulation but not activation. Although IL-4 and IFN-γ mediate many crucial functions of iNKT cells in the liver,[6-8] IL-4−/−IFN-γ−/− dKO mice still had significant liver injury after α-Galcer injection, suggesting that mechanisms other than IL-4 and IFN-γ are involved. It was previously reported that α-Galcer treatment induces TNF-α production by iNKT cells and that inhibition of TNF-α ameliorated α-Galcer-induced liver injury and diminished the aggravating effects of IFN-γ neutralization in this liver injury. These findings suggest that TNF-α likely contributes to the α-Galcer-induced liver injury in IL-4−/−IFN-γ−/− dKO mice.