FIGURE 2.

Upon rechallenge with signals 1 plus 2, anergic T cells fail to not only produce IL-2 and proliferate but up-regulate metabolic machinery. A, Mock-treated and anergic (signal 1) A.E7 T cells were rechallenged with APCs plus PCC peptide for 48 h. Cells were then interrogated for proliferation by thymidine uptake. B, Mock-treated and anergic (signal 1) T cells were rechallenged with anti-CD3 and anti-CD28 for 16 h and interrogated for IL-2 by ELISA. C, Supernatants from rechallenged T cells were tested for L-lactate production. For these experiments some cells were left unstimulated, and some anergic T cells were rescued with high-dose IL-2. D, CD98 cell surface expression of cells treated as in B by FACS. The shaded tracing indicates the isotype control for the stimulated T cells that were initially mock treated. E, As in D, but for CD71. F, As in D and E, but some anergic cells were rescued with IL-2, G, Mock-treated or anergic A.E7 T cells were left unstimulated or were stimulated for 3 h with anti-CD3 and anti-CD28; lysates were made and probed for phospho-S6K1 by immunoblot. H, As in G, but for phospho-Akt (S473). Total enzyme (pan) probes are included as loading controls. Data are representative of three independent experiments. Statistics were performed using the Student’s t test.**, p < 0.01; ***, p < 0.001; NS, p < 0.05.