<i>Cblb</i>−/− BMDCs show increased allogeneic T cell proliferation but are not more effective in the induction of antigen-specific T cell responses and cross-presentation.

<p>(A) Increased allogeneic T cell proliferation induced by <i>cblb</i>−/− BMDCs. LPS matured wildtype or <i>cblb</i>−/− BMDCs were added in increasing numbers to 2×10⁵ allogeneic T cells for two days. Incorporation of ³H-Thymidine radioactivity during the last 16–18 hours of culture was measured. Data represent counts per minute (mean value ± SD, n = 4), representative of 4 independent experiments is shown, *p<0.05 wildtype BMDC <i>versus cblb</i>−/− BMDCs (Student t-test). (B/C) No difference in antigen specific T cell proliferation between wildtype and <i>cblb</i>−/− BMDCs. Wildtype and <i>cblb</i>−/− BMDCs were matured with 100 ng/ml LPS and primed with (B) 10 µM SIINFEKL-peptide (OT-I) or (C) 10 µM ISQAVHAAHAEINEAGR–peptide (OT-II) on day seven of culture overnight. Graded doses of BMDCs were then co-cultured with 2×10⁵ antigen-specific CD8⁺ (OT-I) or CD4⁺ (OT-II) transgenic T cells for 48 hours. Incorporation of ³H-Thymidine radioactivity during the last 16–18 hours of culture was measured. Data represent counts per minute (mean value ± SD, n = 3), representative of at least 3 independent experiments is shown. (D) Unaltered antigen specific T cell priming of <i>cblb</i>−/− BMDCs <i>in vivo</i>. C57BL/6 recipient mice were injected intravenously with 5×10⁶ CFSE-labeled transgenic CD8⁺ T cells. 24 hours thereafter 5×10⁶ wildtype <i>versus cblb</i>−/− BMDCs matured with 100 ng/ml LPS and pulsed <i>in vitro</i> with 10 µM SIINFKEL peptide or solvent as control were injected subcutanously in the flank of the mouse. Representative FACS histogram of CFSE dilution in OT-I transgenic T cells in the draining lymph node after injection of SIINFEKL peptide loaded wildtype <i>versus cblb</i>−/− BMDCs in wildtype recipients is shown. Data represent % proliferated OT-I⁺ T cells (mean value ± SEM, one representative experiment is shown with n = 3 mice per group), G = Generation. (E/F) No difference between wildtype and <i>cblb</i>−/− BMDCs in cross-presentation of protein antigen. Wildtype and <i>cblb</i>−/− BMDCs were cultured from day seven to day eight with 1 mg/ml OVA-protein and 100 ng/mL LPS. Graded doses of BMDCs were then co-cultured with 2×10⁵ antigen-specific (E) CD8⁺ (OT-I) or (F) CD4⁺ (OT-II) T cells for 48 hours. Incorporation of ³H-Thymidine radioactivity during the last 16 to 18 hours of culture was measured. Data are counts per minute (mean value ± SD, n = 2), representative of 2 independent experiments is shown.