Figure 4
Role of TGF-β and STAT1 in CD73 expression. Mice were inoculated orally with 40 cysts of ME-49. (a) Spleen and LP cells were isolated from C57BL/6 naive or day 9 infected mice. Cells were restimulated in vitro with PMA and ionomycin in the presence of Brefeldin A. Cells were stained for TCRβ, CD4, T-bet, CD73, and IFN-γ and analyzed by flow cytometry. Dot plots illustrate CD73, T-bet, and IFN-γ staining profiles of TCRβ+ CD4+-cells. The results are representative of five independent experiments. (b) CD4+ GFP− T lymphocytes were isolated by cell sorting from naive (left panel) or infected (right panel) Foxp3eGFP mice and activated with anti-CD3/CD28 in the absence or presence of TGF-β and/or RA. After 24 h, CD4+ GFP− T lymphocytes were analyzed for CD73 expression by flow cytometry. The results are representative of three independent experiments. (c) Graph summarizes the percentage of TCRβ+ CD4+ cells producing IFN-γ in the spleen, MLN, LP, and IEL compartments from WT and STAT1 KO infected mice. Data are pooled from three independent experiments (n=10). (d) Dot plots illustrate CD73 and IFN-γ staining profiles of CD4+ TCRβ+ cells from WT or STAT1 KO mice. The non-parametric Mann–Whitney U-test was used to determine statistical differences (**P<0.01, ***P< 0.001). GFP, green fluorescent protein; IEL, intraepithelial lymphocyte; IFN-γ, interferon-γ; KO, knockout; LP, lamina propria; MLN, mesenteric lymph node; RA, retinoic acid; TGF-β, transforming growth factor-β; WT, wild type.
