Activation of natural killer T cells inhibits the development of induced regulatory T cells via IFNγ

Recent reports have provided evidence for cross-talk between regulatory T (Treg) cells and natural killer T (NKT) cells. However, it is unclear whether NKT cells play a role in the differentiation of Treg cells. By employing NKT cell-abundant Vα14 TCR transgenic (Tg) and NKT cell-deficient CD1d knock-out (KO) mice, we examined the effects of NKT cells on the in vitro differentiation of induced Treg (iTreg) cells with IL2 and TGFβ. We found that iTreg induction from CD1d KO mice was significantly increased compared to the control. Also, the addition of isolated NKT cells from Vα14 TCR Tg mice to naïve CD4⁺ T cells from CD1d KO mice during iTreg differentiation caused a remarkable reduction of iTreg cells. Through IFNγ neutralization, we showed that this reduction was mediated by IFNγ. Furthermore, the main source of IFNγ during iTreg differentiation was NK1.1⁻CD4⁺Foxp3⁻ T cells. This finding implied that early-activated NKT cells induced Th1-type cells and subsequently underwent apoptosis. Taken together, our results suggest that NKT cells inhibit the in vitro development of iTreg cells by increasing IFNγ.     

Changes of frequency and expression level of CD161 in CD8+ T cells and natural killer T cells in peripheral blood of patients with systemic lupus erythematosus

Immunologic abnormalities of natural killer (NK) cells and T cells play a role in the pathogenesis of systemic lupus erythematosus (SLE). CD161 is expressed on most of the NK cells and on some T cells. The quantities of CD161-expressing cells and expression levels of CD161 were analyzed in T cells and NK cells from patients with SLE compared with normal controls. The expression of CD161 on NK cells, NKT cells, CD4⁺ T cells, and CD8⁺ T cells in peripheral blood from patients with inactive SLE and active SLE, and from the normal controls group were determined using flow cytometry. The frequency and expression level of CD161 in the lymphocyte subsets and its relationship with the quantity of regulatory T cells, anti-double stranded DNA antibody, and the titer of antinuclear antibody were evaluated. Both the percentages of the CD161⁺ subpopulation and the mean fluorescence intensities (MFIs) of CD161 in CD8⁺ T cells and NKT cells decreased significantly in SLE patients compared with normal controls (P < .001). The CD161 expression in CD8⁺ T cells and NKT cells also decreased in the anti-dsDNA (+) group (P < .05). The counts of Treg cells were lower in SLE patients and were weakly correlated with the percentage of the CD161 subpopulation (r = 0.229, P = .016) and the MFIs of CD161 expression in CD8⁺ T cells (r = .232, P = .014). The frequencies and levels of CD161 expression on CD8⁺ T cells and NKT cells were reduced in SLE patients, suggesting that an abnormality of these cells was related to the pathogenesis of SLE.     

Heterogeneity in phenotype and function of CD8+ and CD4/CD8 double‐negative Natural Killer T cell subsets in sooty mangabeys

Background We have recently reported the presence of CD8⁺ and CD4/8 double‐negative (DN) natural killer T (NKT) lymphocytes in sooty mangabeys. To investigate differences in the two NKT cell subsets, we compared the phenotype and function of sooty mangabey CD8⁺ and DN NKT cells. Methods Flow‐sorted NKT lymphocytes from one SIV‐negative sooty mangabey were subjected to limiting dilution cloning. Invariant NKT clones were characterized by flow cytometry and cytokine ELISA. Results The majority of NKT clones displayed an effector memory phenotype and expressed CXCR3 and NKG2D. While CD8⁺ NKT subsets expressed significantly higher levels of granzyme B and perforin and produced more IFN‐γ, the DN NKT subsets secreted significantly more IL‐4, IL‐13, and IL‐10. Conclusions The Th1 and Th2 cytokine bias of CD8⁺ and DN NKT cells, respectively, indicates the presence of functionally heterogeneous populations of NKT cells in sooty mangabeys.     

Systemic analyses of immunophenotypes of peripheral T cells in non-segmental vitiligo: implication of defective natural killer T cells

Although it is widely believed that non‐segmental vitiligo (NSV) results from the autoimmune destruction of melanocytes, a clear understanding of defects in immune tolerance, which mediate this uncontrolled self‐reactivity, is still lacking. In the present study, we systemically evaluated circulating regulatory T (Treg) cells, including CD4⁺CD25⁺FoxP3⁺ Treg cells and invariant natural killer T (iNKT) cells, as well as naïve and memory CD4⁺ and CD8⁺ T cells and their cytokine production, in a cohort of 43 progressive NSV patients with race‐, gender‐, and age‐matched healthy controls. We found that the general immunophenotypes of CD4⁺ and CD8⁺ T cells and the percentage of CD4⁺CD25⁺FoxP3⁺ Tregs were comparable between NSV and healthy controls. However, percentages of peripheral iNKT cells were significantly decreased in NSV patients compared to that in healthy controls. Our data confirm the previous notion that the percentage of peripheral CD4⁺CD25⁺FoxP3⁺ Tregs remains unaltered in NSV and suggests the involvement of defective iNKT cells in the pathogenesis of NSV.     

Expression of murine killer immunoglobulin-like receptor KIRL1 on CD1d-independent NK1.1<Superscript>+</Superscript> T cells

Three mouse killer immunoglobulin-like receptors (KIRs), namely, KIR3DL1, KIRL1, and KIRL2, have recently been identified in C56BL/6 (B6) mice. However, only two Kir genes are found in the B6 mouse genome sequence data base. To clarify this discrepancy, we cloned Kir cDNAs from multiple strains of mice. Sequencing of the cDNA clones showed that the Kir3dl1 gene is found in C3H/HeJ and CBA/J but not in B6 mice. Analysis of the single nucleotide polymorphism data base suggested that Kir3dl1 is the C3H/HeJ and CBA/J allele of Kirl1. We generated mAb to the recombinant KIRL1 protein to investigate its expression pattern. The anti-KIRL1 mAb bound to NK1.1⁺ T cells but only very weakly or at undetectable levels to other lymphocytes including natural killer (NK) cells and conventional T cells. Among NK1.1⁺ T cells, conventional NK T cells stained with CD1d tetramer did not significantly bind anti-KIRL1 mAb, whereas CD1d-tetramer-negative subset was KIRL1-positive. Furthermore, the expression of KIRL1 is readily detected on NK1.1⁺ T cells from β₂-microglobulin-deficient B6 mice. Thus, KIRL1 is predominantly expressed on CD1d-independent NK1.1⁺ T cells.     

Reduced tumorigenesis of EG7 after interleukin-10 gene transfer and enhanced efficacy in combination with intratumorally injection of adenovirus-mediated lymphotactin and the underlying mechanism

Although interleukin-10 (IL-10) is commonly regarded as an immunosuppressive cytokine, a wealth of evidence is accumulating that IL-10 also possesses some immunostimulating antitumor properties. Previous studies demonstrated that forced expression of the IL-10 gene in tumor cells could unexpectedly produce antitumor effects. In this study, we explored the tumorigenesis of EG7 cells transduced with IL-10 gene. In vivo, IL-10 gene transfer reduced tumorigenic capacity of EG7 cells and prolonged survival of the EG7 tumor-bearing mice. It was found that the cytotoxicities of cytotoxic T lymphocytes (CTL) and natural killer cells (NK cells) were enhanced. Assessment of the immune status of the animals showed prevalence of a systemic and tumor-specific Th2 response (high levels of IL-4 and IL-10). To improve the therapeutic efficacy, we combined with intratumoral injection of adenovirus-mediated lymphotactin (Ad-Lptn) into the overestablished EG7 tumor model. More significant inhibition of tumor growth were observed in EG7 tumor-bearing mice that received combined treatment with IL-10 and Lptn gene than those of mice treated with IL-10 or Lptn gene alone. The highest NK cells and CTL activity was induced in the combined therapy group, increasing the production of IL-2 and interferon-γ (IFN-γ) significantly but decreasing the expression of immune suppressive cells (CD4⁺Foxp3⁺ Treg cells and Gr1⁺CD11b⁺ MDSCs). The necrosis of tumor cells was markedly observed in the tumor tissues, accompanying with strongest expression of Mig (monokine induced by interferon-gamma) and IP-10 (interferon-inducible protein 10), weakest expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinases-2 (MMP-2). In vivo, depletion analysis demonstrated that CD8⁺ T cells and NK cells were the predominant effector cell subset responsible for the antitumor effect of IL-10 or Lptn gene. These findings may provide a potential strategy to improve the antitumor efficacy of IL-10 and Lptn.     

Peripheral blood but not synovial fluid natural killer T cells are biased towards a Th1-like phenotype in rheumatoid arthritis

Natural killer T (NKT) cells have been implicated in the regulatory immune mechanisms that control autoimmunity. However, their precise role in the pathogenesis of rheumatoid arthritis (RA) remains unclear. The frequency, cytokine profile and heterogeneity of NKT cells were studied in peripheral blood mononuclear cells (PBMCs) from 23 RA patients and 22 healthy control individuals, including paired PBMC–synovial fluid samples from seven and paired PBMC–synovial tissue samples from four RA patients. Flow cytometry revealed a decreased frequency of NKT cells in PBMCs from RA patients. NKT cells were present in paired synovial fluid and synovial tissue samples. Based on the reactivity of PBMC-derived NKT cells toward α-galactosylceramide, RA patients could be divided into responders (53.8%) and nonresponders (46.2%). However, NKT cells isolated from synovial fluid from both responders and nonresponders expanded upon stimulation with α-galactosylceramide. Analysis of the cytokine profile of CD4⁺ and CD4^- PBMC derived NKT cell lines from RA patients revealed a significantly reduced number of IL-4 producing cells. In contrast, synovial fluid derived NKT cell lines exhibited a Th0-like phenotype, which was comparable to that in healthy control individuals. This suggests that synovial fluid NKT cells are functional, even in patients with nonresponding NKT cells in their blood. We conclude that, because the number of Vα24⁺Vβ11⁺CD3⁺ NKT cells is decreased and the cytokine profile of blood-derived NKT cells is biased toward a Th1-like phenotype in RA patients, NKT cells might be functionally related to resistance or progression of RA. Providing a local boost to the regulatory potential of NKT cells might represent a useful candidate therapy for RA.     

Participation of NK1.1+ T Cells in the Rejection of lpr αβT Cells When Bone Marrow Cells of Ipr Mice Are Transplanted into B6 Mice

When C57BL/6 (B6) mice were irradiated (9 Gy) and received bone marrow (BM) cells of B6-lpr/lpr mouse origin (i.e., lpr→B6), all mice died within 6 days. In the irradiated B6 mice, radioresistant CD3^? IL-2Rβ⁺ NK cells and IL-2Rβ CD3^int cells (i.e., CD3^int cells of extrathymic origin) remained, especially in the liver. There were two subsets, NK1.1⁺ and NK1.1^?, among the IL-2Rβ⁺ CD3^int cells. However, the NK1.1⁺ subset (i.e., NK1.1⁺ T cells) was much more radioresistant, and the majority of CD3^int cells belonged to this subset in irradiated mice. The expansion of lymphocytes from injected BM cells did not occur in the irradiated B6 mice. However, such expansion did take place in irradiated B6-lpr/lpr mice injected with both BM cells of B6-lpr/lpr and B6 origin. As a result, the mice subjected to BM cells survived. Irradiated B6 mice were treated in vivo with anti-NK1.1 mAb or anti-asialoGM₁ antibody to eliminate NK cells alone or both NK cells and NK1.1⁺ T cells. When irradiated B6 mice were pretreated with anti-NK1.1 mAb, the mice could survive. These results suggest that intact NK1.1⁺ T cells of extrathymic origin may recognize abnormal BM cells with the lpr gene and inhibit the expansion of lymphocytes, including abnormal double-negative CD4^?8^? cells, in B6-lpr/lpr mice. To inhibit the expansion of lymphocytes, mechanisms other than Fas ligand/Fas molecules on extrathymic T cells may be responsible.     

Essential Role of NK Cells in IgG Therapy for Experimental Autoimmune Encephalomyelitis

Intravenous immunoglobulin has long been used in treating autoimmune diseases, although mechanisms remain uncertain. Activating Fcγ receptors are receptors of IgG and reported to be essential in intravenous immunoglobulin (IVIG) therapy. Therefore, we hypothesized natural killer (NK) cells, which express abundant activating Fcγ receptors, are the potential cellular target. In experimental autoimmune encephalomyelitis (EAE), we demonstrated that IgG suppressed disease development in intact, but not in NK cell depleted mice. Adoptive transfer of IgG-treated NK cell could protect mice against EAE, and suppressed interferon γ and interleukin 17 production. The percentage of CD4⁺Foxp3⁺ regulatory T cells was significantly increased. The increase of regulatory T cells was also observed in IgG-treated EAE mice but not in NK cell depleted mice. In vitro experiments confirmed that IgG-treated NK cells enhanced regulatory T cell induction from naïve CD4⁺ T cells. Interestingly, cells from draining lymph nodes produced more interleukin 2 after the adoptive transfer of IgG-treated NK cells. We neutralized interleukin 2 and the induction of CD4⁺Foxp3⁺ T cells by IgG-treated NK cells was significantly reduced. To our knowledge, we identified for the first time the critical role of NK cells in the mechanism of IgG-induced induction of Treg cells in treatment of autoimmunity.     

In vivo priming of natural killer T cells by dendritic cells pulsed with hepatoma-derived acid-eluted substances

Many murine tumor cells express not only individual haplotype-matched class I MHC molecules, but also species-specific CD1d molecules. The former class I MHC molecules generally present internally synthesized tumor-derived peptide antigens to highly specific CD8⁺ cytotoxic T lymphocytes (CTLs) in acquired immunity. In contrast, the latter CD1d molecules may present tumor-associated glycolipid antigens to broadly crossreactive natural killer T (NKT) cells, which might correlate with controlling tumor metastasis. Here, we showed that murine hepatoma cell line Hepa1-6-derived acid-eluted substances might contain both D^b class I MHC-restricted antigens and CD1d-restriced substances, which could sensitize not only syngeneic bone marrow-derived DCs (BM-DCs), but also allogeneic BM-DCs expressing haplotype-mismatched class I MHC and species-specific CD1d molecules. To our surprise, intravenous (i.v.) immunization of C57BL/6 mice with the former syngeneic BM-DCs carrying acid-eluted materials primed both CD4^–CD8^– and CD8⁺ NKT cells in the spleen, whereas immunization with the latter allogeneic BM-DCs loaded the tumor-derived substances primed CD4^–CD8^–, but not CD8⁺ NKT cells. The findings shown in the present study will open a new area for cancer immunotherapy using allogeneic DCs and tumor-derived acid-eluted substances.Abbreviations CTLs cytotoxic T lymphocytes - NKT natural killer T - BM-DCs bone marrow-derived dendritic cells - CTM complete T-cell medium - FCS fetal calf serum - MMC mitomycin C - TCRs T cell receptors     

Modulation of lymphocyte function with inhibitory CD2: loss of NK and NKT cells

Analysis of the NK cell developmental pathway suggests that CD2 expression may be important in regulating NK maturation. To test this hypothesis, we developed mice containing only an inhibitory CD2 molecule by linking the extracellular domain of CD2 to an intracellular immunoreceptor tyrosine-based inhibitory motif (ITIM) motif. Mice containing the CD2 Tg(ITIM) transgene, introduced into a CD2 KO background, have no morphologically detectable lymph nodes, although development of the thymus appears normal. In addition, these mice had major loss of both NK and NKT subsets in peripheral organs, while T and B cell frequencies were intact. Expression of CD2 was low on T cells and lacking on B cells and functional defects were observed in these populations. NKT cells expressing CD4 were absent, while the CD8⁺ and double negative NKT cells were retained. Small subsets of NK cells were detected but expression of CD2 on these cells was very low or absent, and their maturation was impaired. Based on the phenotype described here, we believe that these mice represent a unique model to study lymphoid organ and lymphocyte development.     

CD83+CCR7+ NK cells induced by interleukin 18 by dendritic cells promote experimental autoimmune uveitis

Natural killer (NK) cells have been reported to play a pathological role in autoimmune uveitis. However, the mechanisms regarding NK cells in uveitis and factors that affect NK‐cell activation in this condition remain unclear. Here, we report that the number of CD3^‐NK1.1⁺CD83⁺CCR7⁺ cells is increased in the inflamed eyes within a mouse model of experimental autoimmune uveitis (EAU), and these cells express elevated levels of NKG2D, CD69 and IFN‐γ. Adoptively transferring CD83⁺CCR7⁺NK cells aggravates EAU symptoms and increases the number of CD4⁺IFN‐γ⁺T cells and dendritic cells (DCs) within the eye. These CD83⁺CCR7⁺NK cells then promote the maturation of DCs and IFN‐γ expression within T cells as demonstrated in vitro. Furthermore, IL‐18, as primarily secreted by DCs in the eyes, is detected to induce CD83⁺CCR7⁺NK cells. In EAU mice, anti‐IL‐18R antibody treatment also decreases retinal tissue damage, as well as the number of infiltrating CD83⁺CCR7⁺NK cells, T cells and DCs in the inflamed eyes and spleens of EAU mice. These results suggest that CD83⁺CCR7⁺NK cells, as induced by IL‐18 that primarily secreted by DCs, play a critical pathological role in EAU. Anti‐IL‐18R antibody might serve as a potential therapeutic agent for uveitis through its capacity to inhibit CD83⁺CCR7⁺NK cells infiltration.     

Mycobacterium paratuberculosis CobT Activates Dendritic Cells via Engagement of Toll-like Receptor 4 Resulting in Th1 Cell Expansion

Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne disease in animals and MAP involvement in human Crohn disease has been recently emphasized. Evidence from M. tuberculosis studies suggests mycobacterial proteins activate dendritic cells (DCs) via Toll-like receptor (TLR) 4, eventually determining the fate of immune responses. Here, we investigated whether MAP CobT contributes to the development of T cell immunity through the activation of DCs. MAP CobT recognizes TLR4, and induces DC maturation and activation via the MyD88 and TRIF signaling cascades, which are followed by MAP kinases and NF-κB. We further found that MAP CobT-treated DCs activated naive T cells, effectively polarized CD4⁺ and CD8⁺ T cells to secrete IFN-γ and IL-2, but not IL-4 and IL-10, and induced T cell proliferation. These data indicate that MAP CobT contributes to T helper (Th) 1 polarization of the immune response. MAP CobT-treated DCs specifically induced the expansion of CD4⁺/CD8⁺CD44^highCD62L^low memory T cells in the mesenteric lymph node of MAP-infected mice in a TLR4-dependent manner. Our results indicate that MAP CobT is a novel DC maturation-inducing antigen that drives Th1 polarized-naive/memory T cell expansion in a TLR4-dependent cascade, suggesting that MAP CobT potentially links innate and adaptive immunity against MAP.     

NKG2D ligand RAE1ε induces generation and enhances the inhibitor function of myeloid‐derived suppressor cells in mice

Expression of surface NKG2D ligands on tumour cells, which activates nature killer (NK) cells and CD8⁺ T cells, is crucial in antitumour immunity. Some types of tumours have evolved mechanisms to suppress NKG2D‐mediated immune cell activation, such as tumour‐derived soluble NKG2D ligands or sustained NKG2D ligands produced by tumours down‐regulate the expression of NKG2D on NK cells and CD8⁺ T cells. Here, we report that surface NKG2D ligand RAE1ε on tumour cells induces CD11b⁺Gr‐1⁺ myeloid‐derived suppressor cell (MDSC) via NKG2D in vitro and in vivo. MDSCs induced by RAE1ε display a robust induction of IL‐10 and arginase, and these MDSCs show greater suppressive activity by inhibiting antigen‐non‐specific CD8⁺ T‐cell proliferation. Consistently, upon adoptive transfer, MDSCs induced by RAE1ε significantly promote CT26 tumour growth in IL‐10‐ and arginase‐dependent manners. RAE1ε moves cytokine balance towards Th2 but not Th1 in vivo. Furthermore, RAE1ε enhances inhibitory function of CT26‐derived MDSCs and promotes IL‐4 rather than IFN‐γ production from CT26‐derived MDSCs through NKG2D in vitro. Our study has demonstrated a novel mechanism for NKG2D ligand⁺ tumour cells escaping from immunosurveillance by facilitating the proliferation and the inhibitory function of MDSCs.     

Efficacy of TLR7 agonistic imidazoquinoline as immunochemotherapeutic agent against P. Berghei ANKA infected rodent host

Malaria is a serious disease and is one of the most alarming public health issues. Plasmodium is being resistant to various antimalarials including Chloroquine (CQ) which was the first-line therapy for malaria treatment. WHO recommended several combination therapies but declining efficacy was reported to many of these therapies. Despite a great amount of research, efficient malaria vaccine still seems to be a distant dream. Immunochemotherapy could be an alternate strategy to deal with malaria. Based on the differential activity of various cytokines in the pathogenesis of and protection against malaria, the efficacy of highly active TLR7 agonistic imidazoquinoline (BBIQ) in combination with a suboptimal dose of CQ against P. berghei ANKA (PbA) in vivo was investigated. In mice treated with CQ alone, parasite appeared on Day 17 and all mice of this group died by Day 21. Whereas, mice treated with BBIQ along with CQ exhibited no appearance of parasite till Day 23. Frequencies of T cells (CD3⁺, CD4⁺and CD8⁺) and T regulatory cells (CD4⁺, CD25 ⁺and FoxP3⁺) were found to be lower in brain of BBIQ?+?CQ treated mice as compared to BBIQ alone and CQ alone treated mice on Day 10. Inhibition of infiltration of inflammatory T cells and activation of T helper and T cytotoxic cells against the parasite was observed in the mice treated with this combination therapy. Serum levels of IFN-γ and IL-12 were found to be higher on same day in mice treated with BBIQ?+?CQ which revealed the generation of strong Th1 immune response in mice against the infection. Overall, TLR7 agonist acted as an efficient partner when combined with potent antimalarial drug.     

Targeting NKT cells and PD-L1 pathway results in augmented anti-tumor responses in a melanoma model

Invariant or Type 1 NKT cells (iNKT cells) are a unique population of lymphocytes that share characteristics of T cells and natural killer (NK) cells. Various studies have shown that positive costimulatory pathways such as the CD28 and CD40 pathways can influence the expansion and cytokine production by iNKT cells. However, little is understood about the regulation of iNKT cells by negative costimulatory pathways. Here, we show that in vivo activation with α-GalCer results in increased cytokine production and expansion of iNKT cells in the absence of programmed cell death ligand-1 (PD-L1, B7-H1, and CD274). To study whether PD-L1 deficiency on NKT cells would enhance antigen-specific T-cell responses, we utilized CD8⁺ OT-1 OVA transgenic T cells. α-GalCer enhanced the expansion and cytokine production of OT-1 CD8⁺ cells after adoptive transfer into wild-type recipients. However, this expansion was significantly enhanced when OT-1 CD8⁺ T cells were adoptively transferred into PD-L1^−/− recipients. To extend these results to a tumor model, we used the B16 melanoma system. PD-L1^−/− mice given dendritic cells loaded with antigen and α-GalCer had a significant reduction in tumor growth and this was associated with increased trafficking of antigen-presenting cells and CD8⁺ T cells to the tumors. These data demonstrate that abrogating PDL1:PD-1 interactions during the activation of iNKT cells amplifies an anti-tumor response when coupled with DC vaccination.     

CD1d-restricted NKT cells modulate placental and uterine leukocyte populations during chlamydial infection in mice

Invariant CD1d-restricted natural killer T cells play an important immunoregulatory role and can influence a broad spectrum of immunological responses including against bacterial infections. They are present at the fetal–maternal interface and although it has been reported that experimental systemic iNKT cell activation can induce mouse abortion, their role during pregnancy remain poorly understood. In the present work, using a physiological Chlamydia muridarum infection model, we have shown that, in vaginally infected pregnant mice, C. muridarum is cleared similarly in C57BL/6 wild type (WT) and CD1d^−/− mice. We have also shown that infected- as well as uninfected-CD1d^−/− mice have the same litter size as WT counterparts. Thus, CD1d-restricted cells are required neither for the resolution of chlamydial infection of the lower-genital tract, nor for the maintenance of reproductive capacity. However, unexpected differences in T cell populations were observed in uninfected pregnant females, as CD1d^−/− placentas contained significantly higher percentages of CD4⁺ and CD8⁺ T cells than WT counterparts. However, infection triggered a significant decrease in the percentages of CD4⁺ T cells in CD1d^−/− mice. In infected WT pregnant mice, the numbers of uterine CD4⁺ and CD8⁺ T cells, monocytes and granulocytes were greatly increased, changes not observed in infected CD1d^−/− mice. An increase in the percentage of CD8⁺ T cells seems independent of CD1d-restricted cells as it occurred in both WT and CD1d^−/− mice. Thus, in the steady state, the lack of CD1d-restricted NKT cells affects leukocyte populations only in the placenta. In Chlamydia-infected pregnant mice, the immune response against Chlamydia is dampened in the uterus. Our results suggest that CD1d-restricted NKT cells play a role in the recruitment or homeostasis of leukocyte populations at the maternal–fetal interface in the presence or absence of Chlamydia infection.     

Between adaptive and innate immunity: TLR4-mediated perforin production by CD28<Superscript>null</Superscript>T-helper cells in ankylosing spondylitis

CD3⁺CD4⁺CD28^null and CD3⁺CD8⁺CD28^null T cells are enriched in patients with immune-mediated diseases compared with healthy controls. This study shows that CD4⁺CD28^null T cells express Toll-like receptors recognizing bacterial lipopolysaccharides in ankylosing spondylitis, psoriatic arthritis and rheumatoid arthritis. In ankylosing spondylitis, TLR4 (23.1 ± 21.9%) and, to a smaller extent, TLR2 (4.1 ± 5.8%) were expressed on CD4⁺CD28^null T cells, whereas expression was negligible on CD4⁺CD28⁺ and CD8⁺ T cells. CD4⁺CD28^null T cells produced perforin upon stimulation with lipopolysaccharide, and this effect was enhanced by autologous serum or recombinant soluble CD14. Perforin production could be prevented with blocking antibodies directed against CD14 or TLR4. Incubation of peripheral blood mononuclear cells with tumour necrosis factor alpha led to an upregulation of TLR4 and TLR2 on CD4⁺CD28^null T cells in vitro, and treatment of patients with antibodies specifically directed against tumour necrosis factor alpha resulted in decreased expression of TLR4 and TLR2 on CD4⁺CD28^null T cells in vivo. We describe here a new pathway for direct activation of cytotoxic CD4⁺ T cells by components of infectious pathogens. This finding supports the hypothesis that CD4⁺CD28^null T cells represent an immunological link between the innate immune system and the adaptive immune system.     

Homing Receptor Expression Is Deviated on CD56+ Blood Lymphocytes during Pregnancy in Type 1 Diabetic Women

Type 1 Diabetes Mellitus (T1DM) is characterized by an augmented pro-inflammatory immune state. This contributes to the increased risk for gestational complications observed in T1DM mothers. In normal pregnancies, critical immunological changes occur, including the massive recruitment of lymphocytes, particularly CD56^bright NK cells, into early decidua basalis and a 2^nd trimester shift towards Type 2 immunity. Decidual CD56^bright NK cells arise at least partly from circulating progenitors expressing adhesion molecules SELL and ITGA4 and the chemokine receptors CXCR3 and CXCR4. In vitro studies show that T1DM reduces interactions between blood CD56⁺ NK cells and decidual endothelial cells by reducing SELL and ITGA4-based interactions. To address the mechanisms by which specific lymphocyte subsets may be recruited from the circulation during pregnancy and whether these mechanisms are altered in T1DM, flow cytometry was used to examine eight peripheral blood lymphocyte subsets (Type 1 (IL18R1⁺) and Type 2 (IL1RL1⁺) CD56^bright NK, CD56^dim NK, NKT and T cells) from control and T1DM women. Blood was collected serially over pregnancy and postpartum, and lymphocytes were compared for expression of homing receptors SELL, ITGA4, CXCR3, and CXCR4. The decline of Type 1/Type 2 immune cells in normal pregnancy was driven by an increase in Type 2 cells that did not occur in T1DM. CD56^bright NK cells from control women had the highest expression of all four receptors with greatest expression in 2^nd trimester. At this time, these receptors were expressed at very low levels by CD56^bright NK cells from TIDM patients. Type 1/Type 2 NKT cell ratios were not influenced by either pregnancy or TIDM. Our results suggest that T1DM alters immunological balances during pregnancy with its greatest impact on CD56^bright NK cells. This implicates CD56^bright NK cells in diabetic pregnancy complications.