Figure 2.

Signaling via TLR1 and TLR2 drives T_reg cell glycolysis and proliferation, yet reduces suppressive capacity. (a) Glut1 expression (as in Fig. 1) of Ki67^high and Ki67^low tT_reg cells derived from the spleens of wild-type mice an of iT_reg cells derived from CD4⁺CD25⁻ T cells isolated from the spleens of wild-type mice and polarized for 5 d under T_reg cell–skewing conditions and treated with H₂O vehicle (Veh) or Pam₃CSK₄ (PAM; 5 μg/ml) for the final 24 h. (b) Immunoblot analysis of HK2, Glut1 and actin (loading control) in iT_reg cells as in a. (c,d) ECAR of vehicle or Pam₃CSK₄ treated iT_reg (c), and Ki67 expression and phosphorylated S6, determined by flow cytometry (d), in iT_reg cells as in a. (e) Inhibition of the proliferation of T_eff cells by T_reg cells treated for 24 h with DMSO (vehicle (−)) or 20 nM rapamycin (Rap) along with Pam₃CSK₄ (5 μg/ml) (left margin), then repurified by magnetic selection and functionally assessed an in vitro suppression assay with various ratios (above plots) of T_reg cells to T_eff cells. *P < 0.05 (Two-tailed Student’s t-test). Data are representative of three independent experiments (a,b,d,e) or two independent experiments with four technical replicates per group (c; mean + s.d.).