Figure 4. Protective Tregs are generated by TGF-β–dependent peripheral conversion from naive CD4⁺CD25^– T cells.

(A) Experimental procedure for assessing in vivo conversion of naive MBP-specific T cells to Tregs. Naive splenic T cells from Tg4 mice were labeled with CFSE and transferred to nontransgenic or CRP-MBP mice; after 7 days, they were recovered again for transfer to nontransgenic mice, in which EAE was induced. (B) Analysis of transferred CFSE⁺ and residual CFSE^– cells retrieved from CRP-MBP or nontransgenic mice. The CD4⁺CFSE⁺ cells in the respective upper right gates were selected for transfer into wild-type mice. (C) CD4⁺CFSE⁺ cells retrieved from nontransgenic (n = 10) or CRP-MBP (n = 11) mice were transferred to nontransgenic recipient mice (2 × 10⁴ cells per mouse), in which EAE was induced. Mice without cell transfer (n = 8) served as a control. Data are mean ± SEM. (D–F) Analysis of CFSE-labeled T cells (D and F) and CFSE-labeled TGF-β–insensitive T cells (E). Cells were retrieved from spleen and liver 7 days after transfer to CRP-MBP (D and E), K5-MBP (F), or nontransgenic mice. Percent Foxp3⁺CFSE⁺ T cells is indicated.