The CXC chemokine receptor 3 (CXCR3) regulates migration and function of T cells during inflammation. Resting T cells express low levels of CXCR3 but they up-regulate CXCR3 upon activation. Several studies suggest that T helper 1 (Th1)/Th2-associated cytokines regulate levels of CXCR3 on T cells. 1-3 Interferon-(IFN-), which signals via STAT1, enhances CXCR3 expression on T cells, 2 but its role in regulating CXCR3 on CD4 versus CD8 T cells is not clear. We therefore examined the role of IFN-and STAT1 in regulating CXCR3 levels on CD4 and CD8 T cells. Naive CD4 and CD8 T cells from C57BL/6 mice were stimulated as described previously2 with anti-CD3e (3 g/mL; Biolegend, San Diego, CA) and anti-CD28 (4 g/mL; Biolegend) in the presence of anti–IFN-neutralizing or control antibody, and CXCR3 expression was analyzed by flow cytometry. CD4 T cells activated in the presence of anti–IFN-Ab (clone no. XMG1. 2; Pharmingen, San Diego, CA) expressed less CXCR3 than controls (Figure 1A). However, IFN-blockade had no effect on CXCR3 levels in CD8 T cells (Figure 1B). Control CD4 and CD8 T cells up-regulated CXCR3 (Figure 1A, B) and produced comparable IFN-(data not shown). Next, we compared CXCR3 surface expression and mRNA levels in stimulated CD4 and CD8 T cells from wild-type (WT) and STAT1/C57BL/6 mice. Activated STAT1/CD4 T cells failed to up-regulate CXCR3 as efficiently as WT CD4 T cells (Figure 1C). In contrast, STAT1/CD8 T cells showed a significant induction of CXCR3 similar to WT CD8 T cells (Figure 1D). Low CXCR3 expression on STAT1/CD4 T cells also correlated with low CXCR3 mRNA levels (Figure 1E, F).

Activated WT and STAT1/CD4 as well as CD8 T cells produced significant IFN-, but levels were lower in STAT1/T cells (WT CD4, 19908.17 [2582.868] pg/mL; WT CD8, 27361.05 [3484.116] pg/mL; STAT1/CD4, 8012.928 [1627.75] pg/mL; STAT1/CD8, 17152.14 [3680.819] pg/mL). Blockade of IFN-did not inhibit CXCR3 expression on STAT1/CD8 T cells, suggesting that IFN-was not inducing CXCR3 via a STAT1-independent pathway (data not shown). We also determined whether the transcription factors T-bet and eomesodermin (Eomes) are involved in CXCR3 induction on STAT1/CD8 T cells by measuring mRNA levels by real-time reverse transcription–polymerase chain reaction (RT-PCR). Both these factors control CD4 and CD8 T-cell activity by regulating expression of many genes, including Cxcr3, 4 and their expression in T cells can be induced via STAT1-depdendent and-independent mechanims. 5, 6 STAT1/CD4 T cells showed less induction of T-bet mRNA than WT CD4 T cells (Figure 1G). In contrast, STAT1/CD8 T cells contained more T-bet mRNA than WT CD8 T cells, but the difference was not significant (Figure 1H). STAT1/CD4 and CD8 T cells also contained less Eomes mRNA than WT T cells, but these differences were not significant (Figure 1I, J).