Maturation of Dendritic Cells by Recombinant Human CD40L-Trimer Leads to a Homogeneous Cell Population with Enhanced Surface Marker Expression and Increased Cytokine Production (original) (raw)
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Blood, 1998
Recently it has been shown that dendritic cells (DC) can develop from peripheral blood monocytes when grown in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). However, it is unclear whether DC can also develop from monocytes in absence of these cytokines. We therefore analyzed the effect of Flt-3 ligand (Flt3L) and of CD40 ligand on the development of human DC from blood monocytes in the absence of GM-CSF. Adherent peripheral blood mononuclear cells (PBMNC) were cultured in the presence of different cytokine combinations and analyzed for the expression of surface molecules and antigen presenting capacity. For functional analyses, cells were tested for their ability to stimulate allogeneic T lymphocytes in a mixed lymphocyte reaction (MLR), to present soluble antigens, and to induce primary HIV-peptide–specific cytotoxic T-cell (CTL) responses in vitro. Furthermore, expression of DC-CK1, a recently identified chemokine with specific...
Vojnosanitetski pregled, 2011
Background/Aim. 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-polycytidylic acid [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-? (IFN-?), 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...
Immunology, 2005
There is growing interest in the in vitro generation of dendritic cells (DC) from peripheral blood monocytes, but the effect of the method chosen to isolate CD14 + monocytes for subsequent DC generation is poorly documented. The method used to isolate monocytes may have an impact on the subsequent function of DC by affecting their ability to express costimulatory molecules (CD80/86), maturation marker (CD83) and/or to produce important immunomodulatory cytokines. In this study, we show that the positive selection of monocytes by anti-CD14-coated microbeads inhibits the lipopolysaccharide (LPS)-induced production of interleukin (IL)-12, IL-10 and tumour necrosis factor-a (TNF-a) from human DC. However, when DC were grown from monocytes isolated by plastic adherence, LPS induced the production of much higher levels of these cytokines. DC derived from adherence-isolated monocytes induced the development of potent cytotoxic T lymphocytes of the Tc1 subset specific for influenza matrix protein, as confirmed by interferon-c (IFN-c) enzyme-linked immunosorbent spot-forming cell assay (ELISPOT), cytotoxicity assay, major histocompatibility complex (MHC)-peptide tetrameric complexes and T helper 1/T helper 2 (Th1/Th2) cytokine production assays.
Human Dendritic Cells Activated by TSLP and CD40L Induce Proallergic Cytotoxic T Cells
Journal of Experimental Medicine, 2003
Human thymic stromal lymphopoietin (TSLP) is a novel epithelial cell-derived cytokine, which induces dendritic cell (DC)-mediated CD4 ϩ T cell responses with a proallergic phenotype. Although the participation of CD8 ϩ T cells in allergic inflammation is well documented, their functional properties as well as the pathways leading to their generation remain poorly understood. Here, we show that TSLP-activated CD11c ϩ DCs potently activate and expand naive CD8 ϩ T cells, and induce their differentiation into interleukin (IL)-5 and IL-13producing effectors exhibiting poor cytolytic activity. Additional CD40L triggering of TSLP-activated DCs induced CD8 ϩ T cells with potent cytolytic activity, producing large amounts of interferon (IFN)-␥ , while retaining their capacity to produce IL-5 and IL-13. These data further support the role of TSLP as initial trigger of allergic T cell responses and suggest that CD40L-expressing cells may act in combination with TSLP to amplify and sustain pro-allergic responses and cause tissue damage by promoting the generation of IFN-␥producing cytotoxic effectors.
Journal of Leukocyte Biology
That monocytes can differentiate into macrophages or dendritic cells (DCs) makes them an essential link between innate and adaptive immunity. However, little is known about how interactions with pathogens or T cells influence monocyte engagement toward DCs. We approached this point in cultures where granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin (IL)-4 induced monocytes to differentiate into immature DCs. Activating monocytes with soluble CD40 ligand (CD40L) led to accelerated differentiation toward mature CD83 ؉ DCs with up-regulated human leukocyte antigen-DR, costimulatory molecules and CD116 (GM-CSF receptor), and down-regulation of molecules involved in antigen capture. Monocytes primed by phagocytosis of antibody-opsonized, killed Escherichia coli differentiated into DCs with an immature phenotype, whereas Zymosan priming yielded active DCs with an intermediate phenotype. Accordingly, DCs obtained from cultures with CD40L or after Zymosan priming had a decreased capacity to endocytose dextran, but only DCs cultured with CD40L had increased capacity to stimulate allogeneic T cells. DCs obtained after E. coli or Zymosan priming of monocytes produced high levels of proinflammatory tumor necrosis factor ␣ and IL-6 as well as of regulatory IL-10, but they produced IL-12p70 only after secondary CD40 ligation. Thus, CD40 ligation on monocytes accelerates the maturation of DCs in the presence of GM-CSF/IL-4, whereas phagocytosis of different microorganisms does not alter and even facilitates their potential to differentiate into immature or active DCs, the maturation of which can be completed upon CD40 ligation. In vivo, such differences may correspond to DCs with different trafficking and T helper cell-stimulating capacities that could differently affect induction of adaptive immune responses to infections.
Toxicology in Vitro, 2001
One of the most promising alternatives to identify the sensitizing potency of new products is the in vitro culture of human dendritic cells (DC). In vivo, dendritic cells present in the skin are highly specialized antigen-presenting cells (APC) of the immune system, which play a crucial role in the induction of allergic reactions. The DC produce specific cytokines and upregulate specific co-stimulatory molecules in addition to the antigen-MHC complex in order to promote an optimal T-cell activation. The aim of our study is to assess the phenotype, cytokine production and autologous T-cell stimulatory capacity of the in vitro CD34+derived dendritic cells after 24 hours of incubation with the metal allergen NiCl 2 (100-300 mm) and the irritant sodium dodecyl sulfate (SDS; 0.01%), in order to find a sensitive endpoint to discriminate between sensitizers and irritants. After exposure to Ni, a significant increase in the number of CD83+ and CD86+ cells was noticed. The intensity of CD86 as well as the intensity of the HLA-DR molecule on the DC also showed a significant increase. The expression of the co-stimulatory molecule CD80 was not changed after exposure. SDS was not able to increase the expression of any of the analysed markers. The production of IL-6 increased significantly after exposure of dendritic cells to Ni, but not after SDS exposure. Results on tumor necrosis factor-a (TNFa) production are somewhat equivocal. Although not statistically significant, TNF-a was upregulated in one out of three experiments after 48 hours of exposure to the Ni allergen, but increases were also noticed after exposure to SDS (two out of three experiments). Both exposure to Ni and SDS caused an upregulation (not significant) in the IL-12 production by DC, but the production was higher in Ni-exposed DC compared to SDS-exposed cells. In none of the exposed DC cultures was it possible to detect IL-1b. The antigen-presenting capacity of the DC in autologous MLR could not be demonstrated. Nevertheless, T-cell proliferation after DC stimulation was noticed in allogenic MLR. #
Cancer Immunology, Immunotherapy, 2004
Interactions between dendritic cells (DCs) and activated T cells are critically important for the establishment of an effective immune response. To develop the basis for a new DC-based cancer vaccine, we investigated cell-to-cell interactions between human monocyte-derived DCs and autologous T cells that are activated to express the CD40 ligand (CD40L). Peripheral blood monocytes were cultured in the presence of granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin 4 (IL-4) to induce differentiation of DCs. Activated T cells (ATs) consisted of autologous peripheral blood lymphocytes that had been activated with phytohemagglutinin (PHA) and then stimulated with calcium ionophore to up-regulate expression of CD40L. Coculture of these DCs and ATs induced significant production of interleukin 12 (IL-12) and also enhanced the production of interferon c (IFNc). The production of IL-12 was blocked by an anti-CD40L antibody or by separation of the DC and AT fractions by a permeable membrane. Furthermore, coculture of DCs and ATs induced DCs to upregulate CD83 expression and stimulated migration of DCs toward the macrophage inflammatory protein 3-b (MIP-3b). ATs also migrated toward the MIP-3b. These results suggest a combination of DCs and ATs as a potentially effective therapeutic strategy.
British Journal of Haematology, 2001
Using cDNA arrays, we characterized patterns of gene expression in populations of human dendritic cells (DCs) produced for clinical use. Culture and maturation induction of myeloid adherent cells under serum-free conditions yielded DCs with phenotypes similar to those described in serum-based systems. Analysis of gene expression in DCs treated with tumour necrosis factor alpha, soluble CD40L trimer or interferon gamma, however, showed specific patterns for each factor examined. Our studies document the expression of several transcripts that have not hitherto been described in DCs and/or differentially regulated according to the differentiation state of the DCs, and suggest important functional differences among the DC populations examined. In addition, DC maturation directs changes in the levels of mRNA specific for transcriptional regulators that effect the production of cytokines (e.g. BCL-6, c-rel). Other changes observed, including alteration in the gene expression profile of adhesion molecules and chemokine receptors such as CD44H, CD 49B, Rantes R, CXCR5 and CD37, suggest differences in trafficking potential between the populations studied. This broad-based description of DC populations, produced under serum-free conditions, has enabled us to better define intermediate stages of DC maturation as well as the differentiation-inducing effects of cytokines on these cells.