Immunomodulatory Effects of 1,25-Dihydroxyvitamin D₃ on Dendritic Cells Promote Induction of T Cell Hyporesponsiveness to Myelin-Derived Antigens

Figure 3

Immunophenotypic analysis and cytokine secretion profile of in vitro differentiated DC from healthy controls and MS patients upon maturation with proinflammatory stimuli. CD14+ monocytes were cultured for 6 days in the presence of IL-4 and GM-CSF or in the presence of IL-4, GM-CSF, and 1,25(OH)₂D₃ to obtain immature conventional DC (iDC) or 1,25(OH)₂D₃-treated iDC, respectively. On day 6, DC were stimulated with a cocktail of proinflammatory cytokines (i.e., cc-mDC) or with LPS and IFN-γ (i.e., LPS-mDC) or left untreated (i.e., iDC). (a) Representative example showing immunophenotypic analysis of DC. The expression of CD86, CD80, HLA-DR, and CD83 by conventional DC and 1,25(OH)₂D₃-treated DC is determined by flow cytometry. Immature DC are represented by a solid line, cc-mDC are represented by a dashed line, and LPS-mDC are represented by a dark grey filled histogram. Isotype-matched controls are represented by the light grey filled histograms. For analysis, DC were gated on light scatter properties as depicted in the forward scatter (FSC) versus side scatter (SSC) dot plot. (b) Cytokine secretion of (A) IL-1β, (B) IL-6, (C) TNF-α, (D) IL-12p70, (E) IL-10, and (F) TGF-β by conventional DC (open bars) and 1,25(OH)₂D₃-treated DC (black bars) is determined by a multiplex immunoassay or ELISA. Results are expressed as mean ± SEM (healthy controls: ; MS patients: ).