CpG-oligodeoxynucleotides enhance T-cell receptor-triggered interferon-gamma production and up-regulation of CD69 via induction of antigen-presenting cell-derived interferon type I and interleukin-12 (original) (raw)
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Systemic application of CpG-rich DNA suppresses adaptive T cell immunity via induction of IDO
European Journal of Immunology, 2006
CpG-rich oligonucleotides (CpG-ODN) bind to Toll-like receptor 9 (TLR9) and are used as powerful adjuvants for vaccination. Here we report that CpG-ODN not only act as immune stimulatory agents but can also induce strong immune suppression depending on the anatomical location of application. In agreement with the adjuvant effect, subcutaneous application of antigen plus CpG-ODN resulted in antigen-specific T cell activation in local lymph nodes. In contrast, systemic application of CpG-ODN resulted in suppression of T cell expansion and CTL activity in the spleen. The suppressive effect was mediated by indoleamine 2,3-dioxygenase (IDO) as indicated by the observation that CpG-ODN induced IDO in the spleen and that T cell suppression could be abrogated by 1-methyl-tryptophan (1-MT), an inhibitor of IDO. No expression of IDO was observed in lymph nodes after injection of CpG-ODN, explaining why suppression was restricted to the spleen. Studies with a set of knockout mice demonstrated that the CpG-ODN-induced immune suppression is dependent on TLR9 stimulation and independent of type I and type II interferons. The present study shows that for the use of CpG-ODN as an adjuvant in vaccines, the route of application is crucial and needs to be considered. In addition, the results indicate that down-modulation of immune responses by CpG-ODN may be possible in certain pathological conditions. See accompanying commentary: http://dx.
CpG-oligodeoxynucleotides co-stimulate primary T cells in the absence of antigen-presenting cells
European Journal of Immunology, 1999
CpG-containing oligodeoxynucleotides (CpG-ODN) act as powerful adjuvant during in vivo induction of T cell responses. While CpG-ODN directly activate antigen-presenting cells (APC) and thus exert an extrinsic activity on T cells, it is unclear whether they directly affect T cells (intrinsic activity). Here we analyze the effects of CpG-ODN on T cells in an APC-free cell culture. We report that CpG-ODN co-stimulate T cells provided they were triggered via their TCR. CpG-ODN induced IL-2 production, IL-2 receptor expression and thus proliferation. Proliferation was blocked by cyclosporin A or anti-IL-2 monoclonal antibodies (mAb) but not by anti-IL-4 mAb. Moreover, CpG-co-stimulated T cells differentiated into cytolytic T lymphocytes in vitro. Of note, IL-2-driven growth of primed T cells was not affected by CpG-ODN. Co-stimulation was also operative in T cells from CD28 −/− mice and in TCR-transgenic T cells stimulated with peptide. CpG-ODN-mediated co-stimulation of T cells in vitro may thus explain part of the potent adjuvant effects of CpG-ODN in vivo.
PloS one, 2015
CD56 + cells have been recognized as being involved in bridging the innate and acquired immune systems. Herein, we assessed the effect of two major classes of immunostimulatory oligonucleotides (ODNs), PyNTTTTGT and CpG, on CD56 + cells. Incubation of human peripheral blood mononuclear cells (hPBMC) with some of these ODNs led to secretion of significant amounts of interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α) and granulocyte/monocyte colony-stimulating factor (GM-CSF), but only if interleukin 2 (IL2) was present. IMT504, the prototype of the PyNTTTTGT ODN class, was the most active. GM-CSF secretion was very efficient when non-CpG ODNs with high T content and PyNTTTTGT motifs lacking CpGs were used. On the other hand, CpG ODNs and IFNα inhibited this GM-CSF secretion. Selective cell type removal from hPBMC indicated that CD56 + cells were responsible for GM-CSF secretion and that plasmacytoid dendritic cells (PDCs) regulate this process. In addition, PyNTTTTGT ODNs inhibited the IFNα secretion induced by CpG ODNs in PDCs by interference with the TLR9 signaling pathway. Since IFNα is essential for CD56 + stimulation by CpG ODNs, there is a reciprocal interference of CpG and PyNTTTTGT ODNs when acting on this cell population. This suggests that these synthetic ODNs mimic different natural alarm signals for activation of the immune system.
Veterinary Immunology and Immunopathology, 2008
The immune stimulatory effects of synthetic CpG DNA, on porcine peripheral blood mononuclear cells (PBMC) have been reported, but little is known about CpG-induced responses in other lymphoid tissues of pigs. We investigated innate immune responses induced by CpG DNA in cells from blood, lymph nodes (LN) and spleens of pigs. Porcine PBMC and lymph node cells (LNC) were stimulated in vitro with three classes (A-, B-and C-class) of CpG oligodeoxynucleotides (ODNs), and a non-CpG control ODN. All three classes of CpG ODNs induced significant production of IFNa, TNFa, IL-1, IL-6 and IL-12 in PBMC. In contrast, in LNC, only IL-12 was stimulated by all three classes of CpG ODNs, while IFNa, and IL-6 were induced by A-and Cclass ODNs. No TNFa was induced in LNC by any of the ODNs. Significant lymphocyte proliferation was induced in PBMC by all three classes of CpG ODNs and non-CpG control. However, in LNC, B-and C-class ODNs induced significant proliferation, while no proliferation was seen with A-class and non-CpG control ODN. All three classes of ODNs induced NK-like cytotoxicity in PBMC and spleen cells, but were less effective in inducing NK cytotoxicity in LNC. We then investigated the reasons for the relatively poor CpG-induced responses in LNC. Our investigations revealed that LNC had a lower frequency of IFNa-secreting cells and expressed low levels of TLR9 mRNA compared to PBMC. We conclude that the lower number of IFNa-secreting cells and receptor expression may contribute to the attenuated responses in LNC following stimulation with CpG ODN. #
Veterinary Immunology and Immunopathology, 2007
CpG ODN signal through Toll-like receptor 9 (TLR9) and trigger a cascade of events that lead to activation of innate and adaptive immune responses. Our current understanding of the immunobiology of host responses to CpG is based largely on studies on peripheral blood mononuclear cells (PBMC) and splenocytes. Little is known regarding CpG-induced responses in other lymphoid tissues. In the present study, we investigated responses induced by CpG in both PBMC and lymph nodes. Cells were isolated from the superficial cervical lymph node (LNC) and blood and then stimulated with CpG ODN (either A-, or B-or C-class ODN). Cytokine production was assayed by ELISA, and lymphocyte proliferation was determined by 3 H-thymidine incorporation. NK-like cytotoxicity was analyzed by lysis of 51 Cr-labelled target cells. All three classes of CpG induced IFNa and IFNg in LNC. In contrast, only A and C-class ODN induced IFNa and IFNg in PBMC. Moreover, the IFN levels in LNC were 20-40-fold higher than in PBMC. Furthermore, all classes of ODN induced higher IL-12 levels in LNC (five-to six-fold) than in PBMC. Both B and C-class ODN induced good proliferative responses in PBMC and LNC, but the A-class ODN did not induce proliferation of PBMC and only induced moderate proliferation of LNC. A-class ODN induced significant NK-like activity in LNC. Thus, all three classes of CpG ODN induced similar responses in LNC, and these responses were consistently higher than in PBMC. These observations indicate that CpG ODN-induced responses differ between blood and lymph nodes, and suggest that the functional classification of CpG ODN based on PBMC responses may not be directly applicable to cells from other immune tissues. #
Immunomodulatory Effects of CpG Oligodeoxynucleotides on Established Th2 Responses
Clinical and Vaccine Immunology, 2002
CpG oligodeoxynucleotides (CpG ODNs) are known to induce type 1 T-helper-cell (Th1) responses. We have previously demonstrated that CpG ODNs administered during sensitization prevent Th2-mediated eosinophilic airway inflammation in vivo. We also reported that key Th1 cytokines, gamma interferon (IFN-␥) and interleukin 12 (IL-12), are not necessary for this protection. Recent in vivo data suggest that CpG ODNs might also reverse established pulmonary eosinophilia. In order to clarify how CpG ODNs can inhibit established Th2 responses, we evaluated the cytokine production from splenocytes from antigen-and alum-immunized mice. Restimulation with antigen induced IL-5, which was clearly inhibited by coculture with CpG ODNs in a concentration-dependent manner. CpG ODNs also induced IFN-␥, but in a concentration-independent manner. The inhibition of IL-5 production was not mediated through natural killer cells or via CD8 ؉ T lymphocytes. Although IFN-␥ plays an important role in inhibition of antigen-induced IL-5 production by CpG ODNs, IFN-␥ was not the sole factor in IL-5 inhibition. CpG ODNs also induced IL-10, and this induction correlated well with IL-5 inhibition. Elimination of IL-10 reduced the anti-IL-5 effect of CpG ODNs, although incompletely. This may be because IFN-␥, induced by CpG ODNs, is also inhibited by IL-10, serving as a homeostatic mechanism for the Th1-Th2 balance. Overproduction of IFN-␥ was downregulated by CpG ODN-induced IL-10 via modulation of IL-12 production. These data suggest that CpG ODNs may inhibit established Th2 immune responses through IFN-␥ and IL-10 production, the latter serving to regulate excessive Th1 bias. These properties of CpG ODNs might be a useful feature in the development of immunotherapy adjuvants against allergic diseases such as asthma.
European Journal of Immunology, 2006
Oligodeoxynucleotides (ODN) containing unmethylated CpG dinucleotides (CpG ODN) mimic the immunostimulatory activity of microbial DNA via Toll-like receptor (TLR)9. Previous studies indicated that human NK cells express functional TLR3 and TLR9, since their cytokine release and cytolytic function could be incremented by poly(I:C) or ODN A/B, respectively. We have now evaluated the capability of a novel class of CpG ODN, termed ODN C, to modulate the function of human NK cells in the presence of exogenous cytokines. We show that NK cells isolated from peripheral blood and cultured with ODN C, in the presence of either IL-12 or IL-8, express higher levels of CD69 as compared to those stimulated with either ODN A or ODN B. Moreover, NK cells cultured with ODN C displayed higher cytolytic activity against tumor cell lines. These effects were not confined to freshly isolated peripheral blood NK cells since polyclonal NK cell populations that had been cultured in the presence of exogenous IL-2 for several weeks also displayed higher cytolytic activity and cytokine release after culture in the presence of ODN C. Remarkably, NK cells displaying poor responses to ODN A/B were efficiently stimulated by ODN C.
Whole blood cultures to assess the immunostimulatory activities of CpG oligodeoxynucleotides
Journal of Immunological Methods, 2001
Specially designed oligodeoxynucleotide (ODN) sequences known as 'CpG' ODNs elicit innate and acquired immune responses. In general, screening of new CpG ODNs has been conducted by conventional lymphoproliferative assays or expression of activation markers in peripheral blood mononuclear cell (PBMC) cultures. Here, we compared conventional in vitro human PBMC assays with whole blood assays for screening the immunostimulatory properties of CpG ODNs. Commercially available DNA preparations and mycobacterial-based adjuvants were used as comparators. Activation was assessed by flow cytometry and cytokine production. CpG ODNs, identified by four-letter codes, consisted of 2006 (strong human cell stimulant), 1826 (strong murine cell stimulant), 1840 (weak immunostimulant), and 2041, a non-CpG ODN. In both test systems, and in accordance with previous reports, 2006 was an effective up-regulator of CD40 on human dendritic cells (DC1, DC2), monocytes, and B cells, and of CD69 on NK cells. In contrast to murine cells exposed to CpG ODNs, IL-12 (p40) and IFN-gamma production in human immune cells was negligible, but greatly enhanced by adding GM-CSF. Like 2006, two comparator mycobacterial adjuvant formulations activated DC1, DC2, monocytes and natural killer (NK) cells, but only 2006 had a strong effect on B cells. The usefulness of the whole blood assay was further demonstrated by studies in small volumes of umbilical cord mononuclear cells, that like adult blood cells, showed up-regulation of CD40 expression on B cells, DC, and monocytes, and CD69 on NK cells. The whole blood assay, in conjunction with flow cytometry, is useful for assessing the immunological properties of CpG ODN sequences.