title: The influence of CD40 ligation and interferon-gamma on functional properties of human monocyte-derived dendritic cells activated with polyinosinic-polycytidylic acid.
abstract: OBJECTIVE
  Ligation of a Toll-like receptor (TLR) by specific TLR agonists is a powerful tool for maturation induction of monocyte-derived dendritic cells (MoDCs). Studies so far have shown that the treatment of dendritic cells (DCs) with a TLR3 ligand, polyinosinic-polycytidylicacid [Poly(I:C)], may be an appropriate activation agent for obtaining mature MoDCs, competent to prime effective immune responses. However, little is known about how subsequent interaction of MoDCs with T cell-derived stimuli, such as CD40 or interferon-gamma (IFN-gamma), modulates MoDC functions. Therefore, this problem was the main objective of this study.
  
  
  METHODS
  Immature MoDCs were prepared by cultivation of monocytes from peripheral blood mononuclear cells with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin (IL)-4 for 5 days. After that, maturation was induced by the treatment of these cells with Poly(I:C) for 2 days. At day 6, immature MoDCs and Poly(I:C)-activated MoDCs were incubated either with CD40 ligand (L)-transfected J558 cells or IFN-gamma for additional 24 hours. Cytokine production was measured by ELISA and FlowCytomix Human T helper Th1/Th2 11plex. Allostimulatory capability of MoDCs was tested using an allogeneic mixed leukocyte reaction (MLR) assay.
  
  
  RESULTS
  Immature MoDCs showed a moderate potential for stimulation of proliferation of CD4+ T cells, which was enhanced by the treatment with Poly(I:C). Ligation of CD40 or treatment with IFN-gamma of immature or Poly(I:C)-treated MoDCs significantly up-regulated their allostimulatory activity. MoDCs matured in the presence of Poly(I:C) up-regulated the production of IL-12 and IL-10, which was followed by increased levels of IFN-gamma and decreased levels of IL-5 in co-cultures with allogeneic CD4+ T cells. Ligation of CD40 on immature MoDCs upregulated the production of IL-12 and IL-23 which was accompanied by increased secretion of IFN-gamma in co-culture. Stimulation of CD40 on Poly(I:C)-treated MoDCs significantly enhanced the production of IL-12, IL-23 and IL-10. However, such treated MoDCs decreased the production of IFN-gamma and IL-10 and up-regulated the secretion of IL-17. Immature MoDCs treated with IFN-gamma up-regulated IL-12, but lowered the production of IL-5 and IL-17 by CD4+ T cells. Treatment of Poly(I:C)-activated MoDCs with IFN-gamma down-regulated the production of IL-12 and up-regulated IL-10 by these cells and increased/decreased the levels of IL-10/ IFN-gamma, respectively, in co-culture with CD4+ T cells.
  
  
  CONCLUSIONS
  Treatment with Poly(I:C) or ligation of CD40 on immature MoDCs induces maturation of these cells into a phenotype that supports Th1 response. Activation of CD40 on Poly(I:C)-treated MoDCs shifts the immune response towards Th17. Treatment of immature MoDCs with IFN-gamma down-regulated Th2 and Th17 responses. However, addition of IFN-gamma to Poly(I:C)-activated MoDCs down-regulated Th1 response and promote T regulatory mechanisms. Each of these results may have functional and therapeutic implications.
authors.affiliation: Military Medical Academy, Institute for Medical Research, Belgrade, Serbia.
authors.forename: Ana
authors.initials: A
authors.lastname: Dragicević
authors.forename: Tanja
authors.initials: T
authors.lastname: Dzopalić
authors.forename: Sasa
authors.initials: S
authors.lastname: Vasilijić
authors.forename: Dragana
authors.initials: D
authors.lastname: Vucević
authors.forename: Biljana
authors.initials: B
authors.lastname: Bozić
authors.forename: Ivana
authors.initials: I
authors.lastname: Majstorović
authors.forename: Bela
authors.initials: B
authors.lastname: Balint
authors.forename: Miodrag
authors.initials: M
authors.lastname: Colić
mesh_terms: D023201:CD40 Ligand / Q000494:pharmacology*; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism*; D003713:Dendritic Cells / Q000166:cytology*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D009000:Monocytes / Q000276:immunology / Q000378:metabolism*; D011070:Poly I-C / Q000494:pharmacology*; D013601:T-Lymphocytes / Q000276:immunology; D051196:Toll-Like Receptor 3 / Q000378:metabolism; D051193:Toll-Like Receptors / Q000819:agonists*; D015854:Up-Regulation
chemical_list: D016207:Cytokines; D051196:Toll-Like Receptor 3; D051193:Toll-Like Receptors; D023201:CD40 Ligand; D007371:Interferon-gamma; D011070:Poly I-C