Fig 2: Reconstitution of B cell population after depletion. (A) Flow cytometry analysis of total number of B (CD19+) and T cells (CD3+) in single cell spleen preparations from C57Bl/6 mice colonized with S. pneumoniae 3 days after B cell depletion treatment. B cell reconstitution has been assessed 26 days after aCD20 antibody IP administration (3 mice/group). Undepleted mice are shown in black, depleted in white. (B, C) Reconstituted B cell subset populations in splenocytes after one (B) or two (C) depletion treatments were analyzed by flow cytometry 31 and 26 days after depletion, respectively. Marginal zone (CD5-CD21+CD23-IgM+), follicular (CD5-CD23+IgM-), and regulatory (CD5+CD21+IgM+) B cells populations are shown. Depleted/vaccinated (white bars), undepleted/vaccinated (black bars) mice were analyzed (n=6/group). Bars represent mean values for each group, error bars indicate SDs and Mann Whitney U test was used for statistical analysis (*P < 0.05, ns, not significant).

Fig 3: ABCB7 is required for pro-B cell development but not peripheral B cell homeostasis.(A) Flow cytometry analysis of B cell development in bone marrow from wild-type (WT), Mb1-cre ABCB7 conditional knockout (cKO), and CD23-cre ABCB7 cKO mice. Pro-B cells were divided into Hardy fractions as follows: Fr. B (B220+ CD19+ CD43+ BP-1-), Fr. C (B220+ CD19+ CD43+ CD24lo BP-1+), and Fr. C’ (B220+ CD19+ CD43+ CD24hi BP-1+), Fr. D (B220+ CD19+ CD43-/low sIgM-), Fr. E (B220+ CD19+ CD43-/low sIgM+), and Fr. F (B220hi CD19+ CD43-/low sIgM+). Contour plots are representative of six independent experiments (total of 6–11 mice/group). (B) Flow cytometry analysis of splenic B cell populations in WT, Mb1-cre ABCB7 cKO, and CD23-cre ABCB7 cKO mice. Populations were identified by gating on CD19+ splenocytes: transitional type 1 (T1; AA4.1+CD21/35- IgM+ CD23-), transitional type 2 (T2; AA4.1+ CD21/35- IgM+ CD23+), transitional type 3 (T3; AA4.1+CD21/35+IgM+), follicular (FO; AA4.1- CD21/35+ IgM+), and marginal zone (MZ; AA4.1- CD21/35hi IgMhi). Contour plots are representative of seven independent experiments (total of 7–12 mice/group). (C) Graph showing the percentage of total live bone marrow cells for each Hardy fraction in (A). (D) Graph showing absolute cell numbers of splenic B cell populations in (B). (C, D) Lines represent the mean ± SEM. Statistics were obtained by using a one-way ANOVA with Dunnett’s test for multiple comparisons.