Figure 3

Contribution of IgA and B cells to the protective effect of cholera toxin B (CTB). (a) Mice were treated as described in the legend of Figure 1. Total IgA and ovalbumin (OVA)-specific IgA in BAL fluid were determined by enzyme-linked immunosorbent assay (ELISA). (b) CD19⁺ lung B cells were sorted. B cells were stimulated by LPS (10 mg/ml) for 7 days. Total IgA was determined by ELISA. (c) Bone marrow-derived dendritic cells (DCs) were cultured and pulsed as described in the legend of Figure 1. One million DCs were co-cultured with 1 × 10⁶ CD19⁺ spleen B cells, anti-IgM Fab-fragments (10 μg/ml), and in some conditions SB431542 (5 μM) or a DMSO (dimethyl sulfoxide) control was added. After 7 days, supernatants were harvested and total IgA was measured by ELISA. (d) Mediastinal lymph node (MLN) B cells were isolated by CD19-microbeads and injected intravenously (5 × 10⁶) in previously OVA-sensitized mice at day 10. After two days, the mice were challenged and BAL cell differential counts were determined by flowcytometry. (e) Cells were treated as described in c. After 7 days, the B cells were harvested and injected intravenously (5 × 10⁶) as described in d. Mice were treated as described in d. BAL cell numbers were determined by flowcytometry. (f) MLN from the experiment described under e were taken and cells were stimulated by OVA (10 mg/ml). After 4 days, supernatants were taken and cytokines determined by ELISA. Data are mean±s.e.m, n=4 to 6 mice in each group. *P<0.05, **P<0.01, ***P<0.001. Data from one representative experiment out of two or three are shown.

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