Fig. 5. Signals and transcription factors that triggered B-cell glycolysis and inflammatory B_reg cell differentiation.

a Total B cells purified from healthy donors were cultured in medium or treated with CPG-DNA. c-Myc expression was examined by immunoblotting (n = 3). b–d Total B cells purified from healthy donors were cultured in medium or pretreated with DMSO, or inhibitor against c-Myc signals. Thereafter, the cells were cultured in the presence or absence of CPG-DNA. Basal extracellular acidification (b), expression of glycolytic enzymes (c), and cytokine secretion (d) were detected by seahorse analyzer, immunoblotting, and ELISA, respectively (n = 3 for b and c; n = 7 for d). e Purified total B cells from healthy donors were cultured in medium or stimulated with CPG-DNA. Activation of MAP kinase and NF-κB signals were determined by immunoblotting (n = 3). f–i Total B cells purified from healthy donors were cultured in medium or pretreated with DMSO, or inhibitor against MAP kinase (f–i) or NF-κB signals (f). Thereafter, the cells were cultured in the presence or absence of CPG-DNA. Cytokine production (f), expression of c-Myc (g) and glycolytic enzymes (h), and extracellular acidification (i) were detected by immunoblotting, seahorse analyzer, and ELISA, respectively (n = 10 for f; n = 3 for g–i). Data were acquired from more than four independent experiments and shown as means ± SEM. Significance was determined with one-way ANOVA test (d, f). ***p < 0.001