Figure 6. TGF-β restricts IL-2 responsiveness and insulates early Tfh progenitor cells from mTOR signaling.
(A) Stg CD4 T cells were labelled with Cell Trace dye and cultured in vitro with 0.1 μM GP66 peptide ± 10 ng/ml TGF-β and stained for surface expression of CD25 (top) or restimulated with IL-2 to assess pSTAT5 (bottom). Data are representative of four independent experiments. (B) Stg chimeric mice (1 × 106 CD44lo TGF-βRII+/+ CD4-cre+ (WT) or TGF-βRIIf/f CD4-cre+ (KO)) were infected with LCMV Armstrong and 3 days p.i., spleens were fixed immediately in 2% PFA and stained with antibodies to measure surface expression of CD25, Ly6C, and intracellular phosphorylation of STAT5 (pSTAT5) and pS6 directly ex vivo. Data are representative of three independent experiments including 3–5 total mice/group. (C–D) 2 × 105 CD44lo TGF-βRII+/+ CD4-cre+ (WT) or TGF-βRIIf/f CD4-cre+ (KO) Stg cells were adoptively transferred into congenic C57BL/6 recipients infected with WSN-GP33/66 the following day. Mice were treated with PBS or 75 mg/kg rapamycin i.p. daily. On day 8 p.i, Stg cells in the MLN were assessed for expression of PSGL1 and Ly6C (C) or T-bet or GzmB (D). Data are representative of three independent experiments encompassing a total of 9–15 mice/group. *p < 0.05, **p < 0.005 and colored asterisks correspond to the color in the stacked graphs.