Figure 5. Splenic FO and recirculating FO-like bone marrow B cells produce IL-10 which prevents ABC and TNFα-mediated inhibition of B cell precursor growth.

Panel A: Bone marrow cells (1 × 10⁶) from young mice as a source of B cell precursors were co-cultured, separated in transwells, with old splenic ABC (2 × 10⁵) or old splenic FO B cells or old bone marrow FO-like B cells as shown. In addition, as indicated, certain cultures of old splenic ABC also were mixed with either young splenic FO B cells (2 × 10⁵), old splenic FO B cells (2 × 10⁵). Particular cultures also contained anti-IL-10 neutralizing antibody (1μg/ml) or isotype control (cAb) antibody (data not shown; cAb had no effects on IL-7 CFU values). Cultures also contained 5ng/ml rmIL-7. After 3 days of co-culture, the bone marrow cells were harvested from the transwell compartment and seeded into triplicate IL-7 CFU assays. Data is cumulative for 3-11 experiments with data normalized to their respective control cultures. Panel B: FO splenic B cells and FO-like recirculating bone marrow B cells from old mice or splenic FO B cells from young adult IL-10 gene knockout mice, were co-cultured with bone marrow cells (1 × 10⁶) with or without rmTNFα (10ng/ml) for 3 days and bone marrow cell IL-7 CFU were determined in triplicate cultures. Data are cumulative for 2-3 experiments, with the exception of TNFα alone for 5 experiments, with data normalized to their respective control cultures. Panel C: Isolated CD19⁺ B lineage cells from young bone marrow were cultured with 10ng/ml rmTNFα and with graded concentrations of rmIL-10 as shown for 3 days together with 5ng/ml rmIL-7. B lineage cells were then harvested and seeded into triplicate IL-7 CFU assays. Data are cumulative for 3 experiments with data normalized to their respective controls. Control (Ctrl) cultures had B lineage cells with no added cytokines other than rmIL-7. Statistical significance as shown: *, p<0.04; **, p<0.003; ***, p<0.0006; ****, p<0.0001.