High Chlamydia Burden Promotes Tumor Necrosis Factor-Dependent Reactive Arthritis in SKG Mice.

OBJECTIVE: Chlamydia trachomatis is a sexually transmitted obligate intracellular pathogen that causes inflammatory reactive arthritis, spondylitis, psoriasiform dermatitis, and conjunctivitis in some individuals after genital infection. The immunologic basis for this inflammatory response in susceptible hosts is poorly understood. As ZAP-70(W163C) -mutant BALB/c (SKG) mice are susceptible to spondylo-arthritis after systemic exposure to microbial ß-glucan, we undertook the present study to compare responses to infection with Chlamydia muridarum in SKG mice and BALB/c mice. METHODS: After genital or respiratory infection with C muridarum, conjunctivitis and arthritis were assessed clinically, and eye, skin, and joint specimens were analyzed histologically. Chlamydial major outer membrane protein antigen-specific responses were assessed in splenocytes. Treg cells were depleted from FoxP3-DTR BALB/c or SKG mice, and chlamydial DNA was quantified by polymerase chain reaction. RESULTS: Five weeks after vaginal infection with live C muridarum, arthritis, spondylitis, and psoriasiform dermatitis developed in female SKG mice, but not in BALB/c mice. Inflammatory bowel disease did not occur in mice of either strain. The severity of inflammatory disease was correlated with C muridarum inoculum size and vaginal burden postinoculation. Treatment with combination antibiotics starting 1 day postinoculation prevented disease. Chlamydial antigen was present in macrophages and spread from the infection site to lymphoid organs and peripheral tissue. In response to chlamydial antigen, production of interferon-γ and interleukin-17 was impaired in T cells from SKG mice but tumor necrosis factor (TNF) responses were exaggerated, compared to findings in T cells from BALB/c mice. Unlike previous observations in arthritis triggered by ß-glucan, no autoantibodies developed. Accelerated disease triggered by depletion of Treg cells was TNF dependent. CONCLUSION: In the susceptible SKG strain, Chlamydia-induced reactive arthritis develops as a result of deficient intracellular pathogen control, with antigen-specific TNF production upon dissemination of antigen, and TNF-dependent inflammatory disease.

TRAIL+ NK cells control CD4+ T cell responses during chronic viral infection to limit autoimmunity.

Natural killer (NK) cells have been reported to control adaptive immune responses that occur in lymphoid organs at the early stages of immune challenge. The physiological purpose of such regulatory activity remains unclear, because it generally does not confer a survival advantage. We found that NK cells specifically eliminated activated CD4(+) T cells in the salivary gland during chronic murine cytomegalovirus (MCMV) infection. This was dependent on TNF-related apoptosis inducing ligand (TRAIL) expression by NK cells. Although NK cell-mediated deletion of CD4(+) T cells prolonged the chronicity of infection, it also constrained viral-induced autoimmunity. In the absence of this activity, chronic infection was associated with a Sjogren's-like syndrome characterized by focal lymphocytic infiltration into the glands, production of autoantibodies, and reduced saliva and tear secretion. Thus, NK cells are an important homeostatic control that balances the efficacy of adaptive immune responses with the risk of developing autoimmunity.

MyD88-dependent and -independent murine cytomegalovirus sensing for IFN-alpha release and initiation of immune responses in vivo.

Antiviral immunity requires early and late mechanisms in which IFN-alpha and IL-12 play major roles. However, the initial events leading to their production remain largely unclear. Given the crucial role of TLR in innate recognition, we investigated their role in antiviral immunity in vivo. Upon murine CMV (MCMV) infection, both MyD88-/- and TLR9-/- mice were more susceptible and presented increased viral loads compared with C57BL/6, TLR2-/-, TLR3-/-, or TLR4-/- mice. However, in terms of resistance to infection, IFN-alpha production and in many other parameters of early inflammatory responses, the MyD88-/- mice showed a more defective response than TLR9-/- mice. In the absence of the TLR9/MyD88 signaling pathway, cytokine production was dramatically impaired with a complete abolition of bioactive IL-12p70 serum release contrasting with a high flexibility for IFN-alpha release, which is initially (36 h) plasmacytoid dendritic cell- and MyD88-dependent, and subsequently (44 h) PDC-, MyD88-independent and, most likely, TLR-independent. NK cells from MCMV-infected MyD88-/- and TLR9-/- mice displayed a severely impaired IFN-gamma production, yet retained enhanced cytotoxic activity. In addition, dendritic cell activation and critical inflammatory cell trafficking toward the liver were still effective. In the long term, except for isotype switching to MCMV-specific IgG1, the establishment of Ab responses was not significantly altered. Thus, our results demonstrate a critical requirement of TLR9 in the process of MCMV sensing to assure rapid antiviral responses, coordinated with other TLR-dependent and -independent events that are sufficient to establish adaptive immunity.

Interaction between conventional dendritic cells and natural killer cells is integral to the activation of effective antiviral immunity.

Dendritic cells (DCs) regulate various aspects of innate immunity, including natural killer (NK) cell function. Here we define the mechanisms involved in DC-NK cell interactions during viral infection. NK cells were efficiently activated by murine cytomegalovirus (MCMV)-infected CD11b(+) DCs. NK cell cytotoxicity required interferon-alpha and interactions between the NKG2D activating receptor and NKG2D ligand, whereas the production of interferon-gamma by NK cells relied mainly on DC-derived interleukin 18. Although Toll-like receptor 9 contributes to antiviral immunity, we found that signaling pathways independent of Toll-like receptor 9 were important in generating immune responses to MCMV, including the production of interferon-alpha and the induction of NK cell cytotoxicity. Notably, adoptive transfer of MCMV-activated CD11b(+) DCs resulted in improved control of MCMV infection, indicating that these cells participate in controlling viral replication in vivo.

Type I interferon dependence of plasmacytoid dendritic cell activation and migration.

Differential expression of Toll-like receptor (TLR) by conventional dendritic cells (cDCs) and plasmacytoid DC (pDCs) has been suggested to influence the type of immune response induced by microbial pathogens. In this study we show that, in vivo, cDCs and pDCs are equally activated by TLR4, -7, and -9 ligands. Type I interferon (IFN) was important for pDC activation in vivo in response to all three TLR ligands, whereas cDCs required type I IFN signaling only for TLR9- and partially for TLR7-mediated activation. Although TLR ligands induced in situ migration of spleen cDC into the T cell area, spleen pDCs formed clusters in the marginal zone and in the outer T cell area 6 h after injection of TLR9 and TLR7 ligands, respectively. In vivo treatment with TLR9 ligands decreased pDC ability to migrate ex vivo in response to IFN-induced CXCR3 ligands and increased their response to CCR7 ligands. Unlike cDCs, the migration pattern of pDCs required type I IFN for induction of CXCR3 ligands and responsiveness to CCR7 ligands. These data demonstrate that mouse pDCs differ from cDCs in the in vivo response to TLR ligands, in terms of pattern and type I IFN requirement for activation and migration.

Mouse strain differences in plasmacytoid dendritic cell frequency and function revealed by a novel monoclonal antibody.

We report in this study the generation of a novel rat mAb that recognizes mouse plasmacytoid dendritic cells (pDC). This Ab, named 120G8, stains a small subset of CD11c(low) spleen cell with high specificity. This population produces high amounts of IFN-alpha upon in vitro viral stimulation. Both ex vivo- and in vitro-derived 120G8(+) cells display a phenotype identical with that of the previously described mouse pDC (B220(high)Ly6C(high)Gr1(low)CD11b(-)CD11c(low)). Mice treated with 120G8 mAb are depleted of B220(high)Ly6C(high)CD11c(low) cells and have a much-reduced ability to produce IFN-alpha in response to in vivo CpG stimulation. The mAb 120G8 stains all and only B220(high)Ly6C(high)CD11c(low) pDC in all lymphoid organs. Immunohistochemical studies performed with this mAb indicate that pDC are located in the T cell area of spleen, lymph nodes, and Peyer's patches. Although the Ag recognized by 120G8 is not yet known, we show that its expression is up-regulated by type I IFN on B cells and DC. Using this mAb in immunofluorescence studies demonstrates strain- and organ-specific differences in the frequency of pDC and other DC subsets. 129Sv mice have a much higher frequency of pDC, together with a lower frequency of conventional CD8alpha(+)CD11c(high) DC, compared with C57BL/6 mice, both in spleen and blood. The higher ability of 129Sv mice to produce IFN-alpha in vivo is related to a higher number of pDC, but also to a higher ability of pDC from 129Sv mice to produce IFN-alpha in vitro in response to viral stimulation.

Relevance of KIR gene polymorphisms in bone marrow transplantation outcome.

Natural Killer (NK) cells may be involved both in allogeneic bone marrow transplantation (BMT) rejection and graft-versus-host disease (GVHD). The physiologic functions of NK cells appear to be regulated by diverse non-inhibitory and inhibitory receptors including the killer cell immunoglobulin-like receptors (KIR). Although human leukocyte antigen (HLA) epitope mismatches are well-known causes of NK alloreactivity, the role of KIR genes in transplantation remains to be further investigated. In this study, we have evaluated whether KIR genotype differences between donors and recipients of HLA identical (related and unrelated) compared with HLA non-identical unrelated BMT, had an impact on transplantation outcome. Our results show that 5 of 15 KIR genes were always identical in donors and recipients and most variations were observed in the number and specificity of noninhibitory KIR genes. Based on the presence or absence of particular KIR genes, 70 different genotypes were obtained from all individuals. According to the donor or recipient KIR genotype, different combination patterns were described. Interestingly, when the recipient KIR genotype was "included" in the donor KIR genotype, 100% (11/11 pairs) of unrelated BMT developed GVHD compared with 60% (18/30) in all other combinations (p = 0.012). In contrast, no GVHD was observed in related BMT when the recipient KIR genotype was "included" in the donor KIR genotype (p = 0.0001). In conclusion, our results reveal a great diversity for KIR genotypes in donors and recipients of BMT and that the risk of GVHD was maximum in unrelated BMT when the recipient KIR genotype was "included" in the donor KIR genotype.