Figure 6.

CYBB-deficiency in cDCs limits endogenous myelin peptide presentation following phagocytosis of injured oligodendroglial cells. (A) Representative histograms depicting proliferation of 2D2/TCR^MOG-derived myelin-specific CD4⁺ T cells upon coculture with MOG_35-55-pulsed FACS-purified ITGAX^hi MHCII^hi splenic DCs (Cybb^fl/fl: gray; cybb^fl/fl-Itgax-Cre: red). 2D2/TCR^MOG-derived CD4⁺ T cells alone served as a control (light gray). Quantification of 2D2/TCR^MOG-derived CD4⁺ T cell proliferation (CellTrace Violet^dim) (B) or IFNG and IL17 secretion (C) upon coculture with either MOG_35-55- (M_35-55) or MOG_1-125 (M_1-125)-pulsed FACS-purified ITGAX^hi MHCII^hi splenic DCs. One representative of 2 independent experiments is shown. Statistical analysis: Mean ± SEM is depicted. Unpaired two-tailed Student t-test was applied. ns, not significant: P > 0.05. (D) Set-up for sequential coculture assay using damaged (UV-irradiated) or untreated MOG-expressing oligodendroglial cell line MO3.13 (MOG⁺ ODCs). (E) Surface expression levels of CD80, CD86, CD40 and MHCII on ITGAX^hi MHCII^hi are similar upon coculture with iODCs when comparing Cybb^fl/fl and cybb^fl/fl-Itgax-Cre genotypes. Quantification of IFNG and IL17 secretion (F) or 2D2/TCR^MOG-derived CD4⁺ T cell proliferation (CellTrace Violet^dim) (G, left panel) upon coculture with FACS-purified ITGAX^hi MHCII^hi splenic DCs previously pulsed with either UV-irradiated (iODC) or untreated (ODC) MOG-expressing oligodendroglial cells. One representative of 2 independent experiments is shown. Statistical analysis: Mean ± SEM is depicted. Unpaired two-tailed Student t-test was applied. *P < 0.05; **P < 0.005. (G, right panel) Representative histograms depicting proliferation of 2D2/TCR^MOG-derived myelin-specific CD4⁺ T cells upon sequential coculture with ODC-pulsed FACS-purified ITGAX^hi MHCII^hi splenic DCs (Cybb^fl/fl: gray; cybb^fl/fl-Itgax-Cre: red). 2D2/TCR^MOG-derived CD4⁺ T cells alone served as a control (light gray)