Figure 5.

BAFF costimulates B cell proliferation. (A) Surface expression of BAFF in stably transfected 293 cells. 293-BAFF and 293 wt cells were stained with anti-BAFF mAb 43.9 and analyzed by flow cytometry. (B) Costimulation of PBLs by 293-BAFF cells. PBLs (10⁵/well) were incubated with 15,000 paraformaldehyde-fixed 293 cells (293 wt or 293-BAFF) in the presence or absence of anti-B cell receptor antibody (anti-μ). Fixed 293 cells alone incorporated 100 cpm. (C) Dose-dependent costimulation of PBL proliferation by sBAFF in the presence of anti-μ. Proliferation was determined after 72 h incubation by [³H]thymidine incorporation. Controls include cells treated with BAFF alone, with heat-denatured BAFF, or with an irrelevant isotype-matched antibody in place of anti-μ. (D) Comparison of (co)stimulatory effects of sCD40L and sBAFF on PBL proliferation. Experiment was performed as described in panel C. (E) BAFF costimulates Ig secretion of preactivated human B cells. Purified CD19⁺ B cells were activated by coculture with EL-4 T cells and activated T cell supernatants for 5–6 d, then reisolated and cultured for another 7 d in the presence of medium only (−) or containing 5% activated T cell supernatants (T-SUP) or a blend of cytokines (IL-2, IL-4, IL-10). The columns represent means of Ig concentrations for cultures with or without 1 μg/ml BAFF. Means of fold increase ± SD were 1.23 ± 0.11 for medium only, 2.06 ± 0.18 with T cell supernatants (four experiments), and 1.45 ± 0.06 with IL-2, IL-4, and IL-10 (two experiments). These were performed with peripheral blood (three experiments) or cord blood B cells (one experiment; 2.3-fold increase with T cell supernatants, 1.5-fold increase with IL-2, IL-4, and IL-10). (F) Dose–response curve for the effect of BAFF in cultures with T cell supernatants, as shown in panel D. Mean ± SD of three experiments.