IL-2Rα and IL-2Rβ expressions on CD4⁺ and CD8⁺ T cells following 4-1BB triggering.

<p>(A) CD4^<b>+</b> T and CD8^<b>+</b> T cells isolated from C57BL/6 mice were labeled with CFSE, activated with 0.1 μg/ml of anti-CD3 mAb for 16 h, and further treated with anti-rat IgG or 5 μg/ml of anti-4-1BB mAb for another 48 h. The cells were stained with the indicated antibodies and subsequently analyzed by FACSCalibur (BD Bioscience). CFSE dilution of CD4^<b>+</b> T and CD8^<b>+</b> T cells 48 h after the treatment of anti-rat IgG or anti-4-1BB mAb. Values represent the mean number of divisions (A). Percentages of each viable dividing cells were calculated using BD CellQuest software. The absolute numbers of dividing T cells were calculated by multiplying the total number of viable cells by the calculated percentages (B). (C-D) C57BL/6 or RAG2^<b>-/-</b>mice were injected i.v. with CFSE-labeled OT-1 CD8^<b>+</b> T cells (2 × 10^<b>6</b> cells/mice), immunized with the emulsified OVA into the flank sites via s.c. route, and further injected i.p. with anti-rat IgG or anti-4-1BB mAb at days 0 and 2. Inguinal LN cells were prepared from each group of mice at day 5, stained with anti-CD8 and H-2K^<b>b</b>/OVA257-564 pentamer, and subsequently analyzed by FACSCalibur (BD Bioscience) (C). Percentages and absolute numbers of each dividing T cells were assessed as described above (D). (E-G) CD4^<b>+</b> T and CD8^<b>+</b> T cells were cultured as described in (A), CD4- or CD8-gated cells were plotted as CFSE vs. CD25 (E). Anti-rat IgG- or anti-4-1BB-treated CD4^<b>+</b> or CD8^<b>+</b> T cells for 48 h were stained with anti-CD25 (IL-2Rα) or anti-CD122 (IL-2Rβ) along with anti-CD4 or anti-CD8 mAb. Percentages of CD25-expressing CD4^<b>+</b> or CD8^<b>+</b> T cells were calculated (F). MFIs of CD25 and CD122 were calculated using BD Cell Quest software (BD Bioscience) (G). Data are representative of at least three independent experiments. Results in B, D, F, and G are mean ± SD (n = 3; *, p< 0.05; **, p< 0.01;***, p< 0.001). MFI: mean fluorescence intensities.