Characterization of Foxp3-expressing CD4⁺CD25⁺ T cells after sham or CLP surgery. (A-C) Spleens from sham and CLP mice at days 1, 3, and 15 after surgery were stained for CD4, CD25, and Foxp3. The percentage of Foxp3-expressing CD4 T cells with various CD25 expression is shown. (D) CD4⁺CD25⁻ T cells were sorted from spleens of sham and CLP mice. Foxp3 expression was assayed by quantitative PCR, normalized to GAPDH levels, and compared with naive CD4⁺CD25⁻-sorted cells. (E) Absolute numbers of CD4⁺CD25⁺Foxp3⁺ cells in spleens from sham and CLP mice. Combined data of 2 to 3 independent experiments and means ± SEM are shown. *P ≤ .05; **P ≤ .01; ***P ≤ .001 when CLP mice were compared with sham mice; #P ≤ .05 when CLP mice at day 3 were compared with CLP mice at day 15 after surgery. The levels (MFI) of (F) CTLA-4, (G) CD103, (H) GITR were analyzed in the population of CD4⁺CD25^highFoxp3⁺ cells from spleens of sham and CLP mice at days 1, 3, and 15 after surgery. Data are representative of 2 independent experiments (n = 4-5 per group). *P ≤ .05; **P ≤ .01; ***P ≤ .001 when post septic mice were compared with sham mice.