Figure 5.

In vivo proteasome inhibition abolishes the antiinflammatory actions of LX-induced SOCS2: modulation of proinflammatory response during infection. (A) WT and SOCS2-deficient mice (n = 10/group) were infected intraperitoneally with T. gondii, and treated with PBS (0.2 ml/mouse) or PR11 (0.2 mg/kg) every other day, from 11 to 27 d after infection. Serum concentrations of IL-12 p70 (A) and IFN-γ (B) were quantified by ELISA 15 d after infection. At the same time point, mice were killed, spleens were harvested, and cell suspensions were prepared and incubated with PBS or STAg (100 ng/ml) for 4 h, followed by flow cytometric quantification of intracellular IFN-γ in CD4⁺ (C) and CD8⁺ T cells (D). 30 d after infection, the number of brain cysts was determined by microscopy (E). Mortality curves were monitored (F). (G) Spleens were removed from mice 30 d after infection. Frozen sections of spleens were processed and for in situ detection of SOCS2 (I–VI), TRAF2 (I, III and V), and TRAF6 (II, IV and VI) specific primary antibodies were used, followed by a secondary antibody anti-IgG Alexa Fluor 488 (green) and anti-IgG Alexa Fluor 594 (red), and the cells were counterstained with DAPI. Images were captured using an Axiovert microscope with AxioVision software (see Materials and methods). CT, uninfected control; Tg, T. gondii-infected. Graphed values represent the means ± the SD. The experiments shown are representative of at least two independent experiments yielding similar results. Bars, 50 μm.