Figure 2. Loss of AMPKα1 has no defect on plasma cell differentiation but leads to increased antibody synthesis per cell.

(A) Representative flow plots of CD138 vs. B220 expression during plasma cell differentiation after B cells from tamoxifen-treated Rosa26-ER^T2Cre mice (Prkaa1 +/+ or Prkaa1 f/f ) were co-cultured on NB-21.2D9 feeder cells in the presence of 4-OHT and IL-4 (left panel). Total number (mean ± SEM) of plasma cells (CD138⁺B220^lo) throughout co-culture (right panel). (B) Mean (± SEM) concentrations of IgG1 detected in supernatants collected from days 3, 6, and 9 of co-culture. (C) Representative wells and quantification of ELISpot analysis depicting numbers of IgG1 secreting cells per 500 plated cells. (D) Mean spot size of IgG1 ASCs after nine days of co-culture. (E) Relative levels of IgG1 detected in the supernatant eight hours after plating 5 × 10⁴ / 100 μL of day 9 co-cultured cells. Data normalized to wildtype controls. Data represent mean ± SE from at least two independent experiments with n = 6 Prkaa1 +/+ vs. n = 4 or 5 Prkaa1 Δ/Δ mice. (F) Protein G precipitation of intracellular and secreted IgG1 from 7 day-LPS cultures after one hour of labeling with [³H]-leucine and the indicated chase times. (G) [³H]-leucine incorporation into IgG1 collected from lysates and supernatants at indicated chase times. (H) Ratio of [³H]-leucine incorporation in the supernatant to incorporation in the lysate, derived from (G), as an indicator of secretion efficiency. Data are representative of three independent experiments using n = 3 Prkaa1 +/+ vs. n = 3 Prkaa1 Δ/Δ mice. (C-H) Each circle (controls, filled ●; Prkaa1 Δ/Δ, open ○) represents sample from one mouse with mean ± SEM also displayed. P values determined by Mann-Whitney U non-parametric t-test or ANOVA where appropriate. ** indicates p < 0.01, *** indicates p < 0.001, **** indicates p < 0.0001.