Human CD4⁺ CD25^hi Foxp3⁺ regulatory T cells are derived by rapid turnover of memory populations in vivo
J. Clin. Invest. Milica Vukmanovic-Stejic, et al. 116:2423 doi:10.1172/JCI28941 [Go to this article.]

Figure 3
Constraints on CD4⁺ CD25^hi T cell maintenance. (A) Based on CD25 expression, the CD4 population was subdivided into CD25^–, CD25^int, and CD25^hi populations. Bcl-2 expression in each subset was determined by intracellular staining and expressed as median mean fluorescent intensity (MFI). Statistical significance was determined using a 2-tailed, paired Student’s t test. (B) CD4⁺CD25^hi T cells have significantly shorter telomeres than total CD4 T cells in both younger and older donors. Telomere length was measured using a 3-color flow-FISH technique. P values were determined by 2-tailed, paired Student’s t test. (C) CD4⁺CD25^hi T cells cannot upregulate telomerase. FACS-sorted CD4⁺CD45RO⁺CD25^– and CD4⁺CD45RO⁺CD25^hi cells from younger and older donors were stimulated with anti-CD3/anti-CD28 beads for 4 days. Telomerase activity was measured by a TRAP assay. An equivalent number of proliferating (Ki67⁺) cells were used in each reaction. The negative control (– cnt) contains the PCR mix without cell extract and the positive control (+ cnt) contains an extract of a telomerase-positive tumor cell line. TSR8 is a telomeric template, used as PCR control.