Fig. 5.

Zbtb24-deficiency specifically inhibits LPS-induced differentiation of peritoneal B1 cells toward PCs in vitro. CD19⁺B220^lowCD23⁻ B1 and CD19⁺B220^highCD23⁺ B2 cells were FACS-sorted from peritoneal cavities of Cd19^Cre/+ and Zbtb24^B−CKO mice, and subsequently cultured (2–5 × 10⁴ cells/well in 96-U bottom plate) in medium (M), 0.1/1 μg/ml LPS (L-0.1/L-1, respectively) for 3 days. A Representative contour-plots showing the percentages of CD19^lowCD138⁺ PCs in differently cultured B1 or B2 cells on day 3. B, C Bar graphs showing the percentages/absolute numbers of PCs (B) or total living cells (C) 3 days post stimulation. D Bar graphs showing the IgM and IgG3 levels in culture supernatants of LPS-stimulated B1 or B2 cells on day 3. Antibody levels in supernatants of B cells cultured in medium were too low to be detected after 5 × dilution. Each dot represents a single mouse of the indicated genotype (male, 10 weeks of age). E Representative overlayed histograms showing the comparable division profiles (top panel) or contour-plots showing the proliferation (CFSE) versus differentiation (CD138, middle and bottom panels) of cultured peritoneal B1 cells isolated from Cd19^Cre/+ versus Zbtb24^B−CKO mice (male, 14 weeks old, n = 3 and 4 for Cd19^Cre/+ and Zbtb24^B−CKO, respectively). B1 cells were labeled with the cell division tracker CFSE (10 μM) and subsequently cultured in medium (M), LPS (0.1/0.5 μg/ml) or anti-IgM (αCD40, 1/5 μg/ml) for 3 days before flow cytometry analysis. Numbers below horizontal lines in B–D indicate P values. Data are representative of 2–4 independent experiments