Figure 1. Metabolism of AM14 B cells stimulated in vitro.

A - B. Mitochondrial stress test performed on AM14 B cells stimulated with PL2-3, R848 or unstimulated for 16 h. Representative time-course plots and quantitation of basal OCR (A) and basal ECAR (B), which each correspond to the first three time-points on the time-course plots before the addition of oligomycin (ATP synthase inhibitor, first arrow). The subsequent addition of the OXPHOS uncoupler carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP, second arrow) result in maximal respiration and glycolysis. The final addition of rotenone (complex I inhibitor) and antimycin A (complex III inhibitor) shuts down the electron transport chain. C. Basal OCR/ECAR ratio. D. Glycolysis stress test in which glycolysis is induced by the addition of glucose (first arrow), followed by the addition of oligomycin (second arrow) that induces maximal glycolysis, then 2DG that blocks glycolysis. E. Glycolysis (increase in ECAR value after the addition of glucose relative to background), glycolytic capacity (increase in ECAR value after the addition of oligomycin relative to background), and glycolytic reserve (difference between glycolytic capacity and glycolysis) calculated from data shown in D. A - E: n = 3 – 4, with different mice used in the two assays. F - H. Effect of 2DG on AM14 B cells and 13C2 T cells co-cultured with PL2-3. F. Representative FACS plots showing CTV dilution as a measure of proliferation in AM14 B cells (top) and 13C2 T cells (bottom). G. Representative 4–44 IgM and IgG2a ELISPOTs, with each well representing a mouse with 2 × 10⁶ cells. Cells in wells in a same column (IgM and IgG2a) come from the same mouse. Corresponding quantitation on the right. H. IL-2 and IFNγ production in the culture supernatants, n = 3. Paired t tests, *P < 0.05; **P < 0.01. ***P < 0.001.