4-1BB signaling induces selective expression of effector molecules.

<p><b>A</b> Purified splenic PDCA-1⁺ B cells from naïve B6 mice were stimulated with soluble anti-4-1BB (5 µg/ml) for 3d, cell-free culture supernatants were collected, and levels of IFN-α were analyzed by ELISA. These values are presented as bar graphs (mean ± SD). The results of three pooled independent experiments are shown (<i>n</i> = 2). <b>C</b> Total bone marrow-derived cells were stimulated with anti-4-1BB (5 µg/ml). After 6d, cells were collected, washed, stained as indicated, and analyzed. One of three independent experiments is shown (<i>n</i> = 3). <b>D</b> Purified splenic PDCA-1⁺ B cells were stained for the indicated markers either immediately or 3d after stimulation with anti-4-1BB (5 µg/ml). One of two independent experiments is shown (<i>n</i> = 3). <b>C</b> FACS-sorted PDCA-1⁺ B cells from spleens of naïve mice were stained as indicated and flow cytometry was performed. One of three independent experiments is shown (<i>n</i> = 3). <b>E</b> Sorted PDCA-1⁺ B cells were stimulated for 3d with rat IgG or soluble anti-4-1BB (both 5 µg/ml), and cell-free supernatants were collected and analyzed for cytokines using commercially available array screens. The results of three pooled independent experiments are shown (<i>n</i> = 3). <b>F</b> Table showing the position of individual cytokines/chemokines used in the assay. Upregulated molecules in the cells stimulated with anti-4-1BB and their position in the array are highlighted. Pos, positive; Neg, negative.