Panel A: A scatter plot shows B cell development in bone marrow chimeric mice. The y-axis represents the log2 of the measured AIDR112H slash WT ratio, with values indicating the advantage or disadvantage of AIDR112H. Different B cell stages are compared, with significant differences noted in Transitional 1 and T2 MZP stages. Panel B: Flow cytometry plots and bar graphs show total plasma cell differentiation in the spleen, with no significant differences between WT and AIDR112H mice. Panel C: Flow cytometry plots and bar graph depict T follicular helper cells in the spleen, showing non-significant differences between WT and AIDR112H mice. Panel D: Flow cytometry plots illustrate the gating strategy to identify plasma cells and other B cell populations in germinal centers and GL7lo-CD95 positive cell populations from WT and AIDR112H cells. Panel E: Bar graphs quantify the data from Panel D, showing significant differences in CD138 positive cells in the dark zone and GL7lo-CD95 positive cells between WT and AIDR112H mice. Panel F: Immunohistochemistry images of germinal centers from spleen sections of WT and AIDR112H mice, stained for CD21 slash 35, GL7, and IgD, with a scale bar of 500 micrometers. Panel G: A heatmap displays the top 50 differentially expressed genes comparing GL7lo-CD95 positive vs GL7 plus CD95 positive cells.
B cell development and differentiation in bone marrow chimeric mice. (A) B cell development in the bone marrow chimeric mice. y axis: log2 of measured AIDR112H/WT-to-grafted AIDR112H/WT ratio, value 1 = twofold advantage, −1 = twofold disadvantage of AIDR112H; B220+, transitional 1: P < 0.0001; T2 MZP, P < 0.01. one-sample t test (n = 14–22, three experiments). (B) PC differentiation in the spleen. FACS plot and quantification: PB and PC, P > 0.05, unpaired t test (n = 9, two experiments). (C) Tfh cells in the spleen. FACS plot and quantification, nonsignificant. ANOVA (n = 5, two experiments). (D) Gating strategy to identify PB and PC in GC and GL7lo/−CD95+ cell population from CD45.1 WT cells and CD45.2 AIDR112H cells. PB: B220+CD138+, PC: B220−CD138+, representative FACS plots. (E) Quantification for D, CD138+ cells (PB + PC) in DZ and LZ. DZ: P = 0.0024; LZ: nonsignificant, two-way ANOVA (n = 6, two experiments); GL7lo/−CD95+ cells in CD45.1 WT cells and CD45.2 AIDR112H cells, P < 0.001, paired t test (n = 15, three experiments); CD138+ cells in different subsets from GL7lo/−CD95+ cell population: CXCR4hi: P = 0.0014; CD83/86+: P = 0.225, two-way ANOVA (n = 6, two experiments). (F) Immunohistochemistry of GC from full spleen sections of WT and AIDR112H mouse 7 days after SRBC immunization. Red: CD21/35; green: GL7; blue: IgD. Scale bar, 500 µm. (G) RNA-sequencing heatmap for the top 50 DEGs comparing GL7lo/−CD95+ versus GL7+CD95+. T2 MZP, transitional 2 marginal zone progenitor; DEGs, differentially expressed genes. *P < 0.05, **P < 0.01, ***P < 0.001.