Fig 2: Vitamin C (VC) promotes the generation of bona fide plasma cells.(A, B) Expression analysis of lineage transcription factors (TFs). B cells were cultured as indicated for 7 days, and the expression of transcription factors was analyzed by intracellular staining and fluorescence-activated cell sorting (FACS) (n = 6). Representative FACS plots (A) and summaries (B) are shown. Note that BLIMP1 and IRF4 are important TF plasma cell differentiation. (C) Antibody secretion by VC-induced plasma cells. Culture supernatants were collected after day 7, and the secreted antibodies with indicated isotypes (IgM, IgG1, IgE) were analyzed by ELISA (n = 6). All data are from at least two independent experiments. Mean ± SEM is shown for bar charts, and the statistical significance was determined by unpaired Student’s t-test. ***p<0.001.

Fig 3: NP-specific B cells from aged mice are phenotypically similar to those from young adult mice. (A) Gating strategy for NP-specific IgD+ B cells, follicular B cells (CD23hiCD21int), marginal zone B cells (CD23−CD21+), CD23−CD21− cells, and atypical B cells (CD23−CD21−CD11c+) in young (6 wo) and aged (99 wo) B1-8i Tg mice. Numbers adjacent to gates indicate percentage of parent population. Cells were pregated for live cells and single cells. (B) Graphs depicting the percentage of IgD+ cells, follicular cells (CD23hiCD21int), marginal zone cells (CD23−CD21+), CD23−CD21− cells, and CD11c+CD23−CD21− cells among NP+B220+CD19+ cells in young (6–12 wo) and aged (90–100 wo) B1-8i Tg mice. Bar height corresponds to the mean, error bars indicate SD, and each symbol represents values from individual mice. Statistics were calculated using unpaired Mann–Whitney U test. Data were pooled from five independent repeat experiments. (C–N) Representative flow cytometric histograms and graphs showing the mean fluorescence intensities (MFIs) of CXCR4 (C), CD62L (D), CXCR5 (E), CD38 (F), GL7 (G), IRF4 (H), CXCR3(I), CD11b (J), CD11c (K), CD86 (L), CD40 (M), and MHC class II (N) of NP+B220+CD19+ cells from young (6 wo) or aged (99 wo) B1-8i Tg mice. Each symbol represents values from individual mice, with young and aged donors from the same experiment shown as paired values. Statistics were calculated using Wilcoxon matched pairs signed rank test. Data are pooled from at least four independent repeat experiments. wo, wk old.