ABSTRACT
Objective: The immune response initiated by SARS-CoV-2 infection in pregnancy is poorly elucidated. We aimed to access and compare the antiviral cellular responses and lymphocytes activation between healthy pregnancies and pregnant women infected with SARS-CoV-2. Methods: We detected the immunological changes of lymphocytes in peripheral blood of healthy non-pregnant women, non-pregnant women with COVID-19, healthy pregnant women, pregnant women with COVID-19 and convalescent group by flow cytometry. In vitro blockade was used to identify NKT-like cell activation through ICOS-ICOSL pathway. Results: We found that CD3+CD56+ NKT-like cells decreased significantly in COVID-19 positive pregnant women compared to healthy pregnant women. NKT-like cells of pregnant women expressed higher level of activating receptors CD69 and NKp46 after SARS-CoV-2 infection. Particularly, they also increased the expression of the co-stimulatory molecule ICOS. NKT-like cells of pregnant women with COVID-19 up-regulated the expression of IFN-γ, CD107a and Ki67. Meanwhile, we found that ICOSL expression was significantly increased on pDCs in pregnant women with COVID-19. Blocking ICOS in vitro significantly decreased the antiviral activity of NKT-like cells in COVID-19 positive pregnant women, suggesting that ICOS-ICOSL may play an important role in the virus clearance by NKT-like cells. Conclusions: During SARS-CoV-2 infection, NKT-like cells of pregnant women activated through ICOS-ICOSL pathway and played an important role in the antiviral response.
Subject(s)
COVID-19 , Inducible T-Cell Co-Stimulator Ligand , Inducible T-Cell Co-Stimulator Protein , Natural Killer T-Cells , Pregnancy Complications, Infectious , SARS-CoV-2 , Humans , Female , Pregnancy , Natural Killer T-Cells/immunology , Natural Killer T-Cells/metabolism , COVID-19/immunology , COVID-19/virology , Inducible T-Cell Co-Stimulator Protein/metabolism , Adult , Inducible T-Cell Co-Stimulator Ligand/metabolism , Pregnancy Complications, Infectious/immunology , Pregnancy Complications, Infectious/virology , SARS-CoV-2/immunology , Lymphocyte Activation/immunology , Antigens, CD/metabolism , Antigens, Differentiation, T-Lymphocyte/metabolism , Signal Transduction/immunology , Interferon-gamma/metabolism , Natural Cytotoxicity Triggering Receptor 1/metabolism , Lectins, C-Type/metabolismABSTRACT
Elevated levels of miR-155 in solid and liquid malignancies correlate with aggressiveness of the disease. In this manuscript, we show that miR-155 targets transcripts encoding IcosL, the ligand for Inducible T-cell costimulator (Icos), thus impairing the ability of T cells to recognize and eliminate malignant cells. We specifically found that overexpression of miR-155 in B cells of Eµ-miR-155 mice causes loss of IcosL expression as they progress toward malignancy. Similarly, in mice where miR-155 expression is controlled by a Cre-Tet-OFF system, miR-155 induction led to malignant infiltrates lacking IcosL expression. Conversely, turning miR-155 OFF led to tumor regression and emergence of infiltrates composed of IcosL-positive B cells and Icos-positive T cells forming immunological synapses. Therefore, we next engineered malignant cells to express IcosL, in order to determine whether IcosL expression would increase tumor infiltration by cytotoxic T cells and reduce tumor progression. Indeed, overexpressing an IcosL-encoding cDNA in MC38 murine colon cancer cells before injection into syngeneic C57BL6 mice reduced tumor size and increased intratumor CD8+ T cell infiltration, that formed synapses with IcosL-expressing MC38 cells. Our results underscore the fact that by targeting IcosL transcripts, miR-155 impairs the infiltration of tumors by cytotoxic T cells, as well as the importance of IcosL on enhancing the immune response against malignant cells. These findings should lead to the development of more effective anticancer treatments based on maintaining, increasing, or restoring IcosL expression by malignant cells, along with impairing miR-155 activity.
Subject(s)
Inducible T-Cell Co-Stimulator Ligand , MicroRNAs , MicroRNAs/genetics , MicroRNAs/metabolism , Animals , Mice , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Ligand/genetics , B-Lymphocytes/immunology , B-Lymphocytes/metabolism , Cell Line, Tumor , Mice, Inbred C57BL , Humans , T-Lymphocytes, Cytotoxic/immunology , Gene Expression Regulation, Neoplastic , Inducible T-Cell Co-Stimulator Protein/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Neoplasms/immunology , Neoplasms/genetics , Neoplasms/pathologyABSTRACT
Background and aims: Inducible T-cell Co-Stimulator (ICOS) present on T-lymphocytes and its ligand ICOSL expressed by myeloid cells play multiple roles in regulating T-cell functions. However, recent evidence indicates that reverse signalling involving ICOSL is also important in directing the differentiation of monocyte-derived cells. In this study, we investigated the involvement of ICOS/ICOSL dyad in modulating macrophage functions during the evolution of metabolic dysfunction-associated steatohepatitis (MASH). Results: In animal models of MASH, ICOS was selectively up-regulated on CD8+ T-cells in parallel with an expansion of ICOSL-expressing macrophages. An increase in circulating soluble ICOSL was also evident in patients with MASH as compared to healthy individuals. ICOSL knockout (ICOSL-/-) mice receiving choline/methionine deficient (MCD) diet for 6 weeks had milder steatohepatitis than wild type mice. MASH improvement was confirmed in mice fed with cholesterol-enriched Western diet for 24 weeks in which ICOSL deficiency greatly reduced liver fibrosis along with the formation of crown-like macrophage aggregates producing the pro-fibrogenic mediators osteopontin (OPN) and galectin-3 (Gal-3). These effects associated with a selective shewing of F4-80+/CD11bhigh monocyte-derived macrophages (MoMFs) expressing the Triggering Receptor Expressed on Myeloid cells 2 (TREM2) to CD11blow/F4-80+ cells positive for the Kupffer cell marker C-type lectin-like type 2 receptor (CLEC-2), thus indicating an increased MoMF maturation toward monocyte-derived Kupffer cells. Conclusions: These results suggest that CD8+ T-cells interaction with monocyte-derived macrophages through ICOS/ICOSL critically supports a specific subset of TREM2+-expressing cells contributing to the evolution of steatohepatitis. The data also point ICOS/ICOSL dyad as a possible target for therapeutic interventions in MASH.
Subject(s)
CD8-Positive T-Lymphocytes , Fatty Liver , Animals , Humans , Mice , CD8-Positive T-Lymphocytes/metabolism , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Interleukin-2 , Ligands , Signal TransductionABSTRACT
Skin-resident CD8+ T cells include distinct interferon-γ-producing [tissue-resident memory T type 1 (TRM1)] and interleukin-17 (IL-17)-producing (TRM17) subsets that differentially contribute to immune responses. However, whether these populations use common mechanisms to establish tissue residence is unknown. In this work, we show that TRM1 and TRM17 cells navigate divergent trajectories to acquire tissue residency in the skin. TRM1 cells depend on a T-bet-Hobit-IL-15 axis, whereas TRM17 cells develop independently of these factors. Instead, c-Maf commands a tissue-resident program in TRM17 cells parallel to that induced by Hobit in TRM1 cells, with an ICOS-c-Maf-IL-7 axis pivotal to TRM17 cell commitment. Accordingly, by targeting this pathway, skin TRM17 cells can be ablated without compromising their TRM1 counterparts. Thus, skin-resident T cells rely on distinct molecular circuitries, which can be exploited to strategically modulate local immunity.
Subject(s)
CD8-Positive T-Lymphocytes , Immunologic Memory , Memory T Cells , Skin , CD8-Positive T-Lymphocytes/immunology , Memory T Cells/immunology , Skin/immunology , Humans , Th17 Cells/immunology , Inducible T-Cell Co-Stimulator Ligand/metabolism , Proto-Oncogene Proteins c-maf/metabolism , Interleukin-7/metabolismABSTRACT
A20 is a ubiquitin-modifying protein that negatively regulates NF-κB signaling. Mutations in A20/TNFAIP3 are associated with a variety of autoimmune diseases, including multiple sclerosis (MS). We found that deletion of A20 in central nervous system (CNS) endothelial cells (ECs) enhances experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. A20ΔCNS-EC mice showed increased numbers of CNS-infiltrating immune cells during neuroinflammation and in the steady state. While the integrity of the blood-brain barrier (BBB) was not impaired, we observed a strong activation of CNS-ECs in these mice, with dramatically increased levels of the adhesion molecules ICAM-1 and VCAM-1. We discovered ICOSL to be expressed by A20-deficient CNS-ECs, which we found to function as adhesion molecules. Silencing of ICOSL in CNS microvascular ECs partly reversed the phenotype of A20ΔCNS-EC mice without reaching statistical significance and delayed the onset of EAE symptoms in WT mice. In addition, blocking of ICOSL on primary mouse brain microvascular ECs impaired the adhesion of T cells in vitro. Taken together, we propose that CNS EC-ICOSL contributes to the firm adhesion of T cells to the BBB, promoting their entry into the CNS and eventually driving neuroinflammation.
Subject(s)
Encephalomyelitis, Autoimmune, Experimental , Neuroinflammatory Diseases , Tumor Necrosis Factor alpha-Induced Protein 3 , Animals , Mice , Blood-Brain Barrier/metabolism , Central Nervous System/metabolism , Endothelial Cells/metabolism , Mice, Inbred C57BL , Multiple Sclerosis/metabolism , Neuroinflammatory Diseases/metabolism , T-Lymphocytes/metabolism , Inducible T-Cell Co-Stimulator Ligand/metabolism , Tumor Necrosis Factor alpha-Induced Protein 3/metabolismABSTRACT
Inducible T cell co-stimulator (ICOS), an immune checkpoint protein expressed on activated T cells and its unique ligand, ICOSL, which is expressed on antigen-presenting cells and non-hematopoietic cells, have been extensively investigated in the immune response. Recent findings showed that a soluble recombinant form of ICOS (ICOS-Fc) can act as an innovative immunomodulatory drug as both antagonist of ICOS and agonist of ICOSL, modulating cytokine release and cell migration to inflamed tissues. Although the ICOS-ICOSL pathway has been poorly investigated in the septic context, a few studies have reported that septic patients have reduced ICOS expression in whole blood and increased serum levels of osteopontin (OPN), that is another ligand of ICOSL. Thus, we investigated the pathological role of the ICOS-ICOSL axis in the context of sepsis and the potential protective effects of its immunomodulation by administering ICOS-Fc in a murine model of sepsis. Polymicrobial sepsis was induced by cecal ligation and puncture (CLP) in five-month-old male wild-type (WT) C57BL/6, ICOS-/-, ICOSL-/- and OPN-/- mice. One hour after the surgical procedure, either CLP or Sham (control) mice were randomly assigned to receive once ICOS-Fc, F119SICOS-Fc, a mutated form uncapable to bind ICOSL, or vehicle intravenously. Organs and plasma were collected 24 h after surgery for analyses. When compared to Sham mice, WT mice that underwent CLP developed within 24 h a higher clinical severity score, a reduced body temperature, an increase in plasma cytokines (TNF-α, IL-1ß, IL-6, IFN-γ and IL-10), liver injury (AST and ALT) and kidney (creatinine and urea) dysfunction. Administration of ICOS-Fc to WT CLP mice reduced all of these abnormalities caused by sepsis. Similar beneficial effects were not seen in CLP-mice treated with F119SICOS-Fc. Treatment of CLP-mice with ICOS-Fc also attenuated the sepsis-induced local activation of FAK, P38 MAPK and NLRP3 inflammasome. ICOS-Fc seemed to act at both sides of the ICOS-ICOSL interaction, as the protective effect was lost in septic knockout mice for the ICOS or ICOSL genes, whereas it was maintained in OPN knockout mice. Collectively, our data show the beneficial effects of pharmacological modulation of the ICOS-ICOSL pathway in counteracting the sepsis-induced inflammation and organ dysfunction.
Subject(s)
Osteopontin , Sepsis , Animals , Male , Mice , Creatinine , Cytokines/metabolism , Immune Checkpoint Proteins , Immunity , Immunomodulation , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Inflammasomes , Inflammation , Interleukin-10 , Interleukin-6 , Ligands , Mice, Inbred C57BL , Mice, Knockout , NLR Family, Pyrin Domain-Containing 3 Protein , p38 Mitogen-Activated Protein Kinases , Sepsis/drug therapy , Tumor Necrosis Factor-alpha , UreaABSTRACT
BACKGROUND: ICOS and its ligand ICOSL are immune receptors whose interaction triggers bidirectional signals that modulate the immune response and tissue repair. AIM: The aim of this study was to assess the in vivo effects of ICOSL triggering by ICOS-Fc, a recombinant soluble form of ICOS, on skin wound healing. METHODS: The effect of human ICOS-Fc on wound healing was assessed, in vitro, and, in vivo, by skin wound healing assay using ICOS-/- and ICOSL-/- knockout (KO) mice and NOD-SCID-IL2R null (NSG) mice. RESULTS: We show that, in wild type mice, treatment with ICOS-Fc improves wound healing, promotes angiogenesis, preceded by upregulation of IL-6 and VEGF expression; increases the number of fibroblasts and T cells, whereas it reduces that of neutrophils; and increases the number of M2 vs. M1 macrophages. Fittingly, ICOS-Fc enhanced M2 macrophage migration, while it hampered that of M1 macrophages. ICOS-/- and ICOSL-/- KO, and NSG mice showed delayed wound healing, and treatment with ICOS-Fc improved wound closure in ICOS-/- and NSG mice. CONCLUSION: These data show that the ICOS/ICOSL network cooperates in tissue repair, and that triggering of ICOSL by ICOS-Fc improves cutaneous wound healing by increasing angiogenesis and recruitment of reparative macrophages.
Subject(s)
Immunoglobulin Fc Fragments , Inducible T-Cell Co-Stimulator Ligand , Inducible T-Cell Co-Stimulator Protein , Wound Healing , Animals , Immunoglobulin Fc Fragments/immunology , Immunoglobulin Fc Fragments/pharmacology , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Ligand/immunology , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Inducible T-Cell Co-Stimulator Protein/immunology , Mice , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Recombinant Proteins/pharmacology , Wound Healing/drug effectsABSTRACT
Immunofibroblasts have been described within tertiary lymphoid structures (TLS) that regulate lymphocyte aggregation at sites of chronic inflammation. Here we report, for the first time, an immunoregulatory property of this population, dependent on inducible T-cell co-stimulator ligand and its ligand (ICOS/ICOS-L). During inflammation, immunofibroblasts, alongside other antigen presenting cells, like dendritic cells (DCs), upregulate ICOSL, binding incoming ICOS + T cells and inducing LTα3 production that, in turn, drives the chemokine production required for TLS assembly via TNFRI/II engagement. Pharmacological or genetic blocking of ICOS/ICOS-L interaction results in defective LTα expression, abrogating both lymphoid chemokine production and TLS formation. These data provide evidence of a previously unknown function for ICOSL-ICOS interaction, unveil a novel immunomodulatory function for immunofibroblasts, and reveal a key regulatory function of LTα3, both as biomarker of TLS establishment and as first driver of TLS formation and maintenance in mice and humans.
Subject(s)
Tertiary Lymphoid Structures , Animals , Chemokines , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Inducible T-Cell Co-Stimulator Protein/metabolism , Inflammation , MiceABSTRACT
Negative selection of developing T cells plays a significant role in T-cell tolerance to self-antigen. This process relies on thymic antigen-presenting cells which express both self-antigens and cosignaling molecules. Inducible T-cell costimulator (ICOS) belongs to the CD28 family of cosignaling molecules and binds to ICOS ligand (ICOSL). The ICOS signaling pathway plays important roles in shaping the immune response to infections, but its role in central tolerance is less well understood. Here we show that ICOSL is expressed by subsets of thymic dendritic cells and medullary thymic epithelial cells as well as thymic B cells. ICOS expression is upregulated as T cells mature in the thymus and correlates with T-cell receptor signal strength during thymic selection. We also provide evidence of a role for ICOS signaling in mediating negative selection. Our findings suggest that ICOS may fine-tune T-cell receptor signals during thymic selection contributing to the generation of a tolerant T-cell population.
Subject(s)
Antigen-Presenting Cells , T-Lymphocytes , Antigen-Presenting Cells/metabolism , B-Lymphocytes/metabolism , CD28 Antigens/metabolism , Inducible T-Cell Co-Stimulator Ligand/metabolismABSTRACT
Elevated gene expression of the costimulatory receptor Icos is a hallmark of CD8+ tissue-resident memory (Trm) T cells. Here, we examined the contribution of ICOS in Trm cell differentiation. Upon transfer into WT mice, Icos-/- CD8+ T cells exhibited defective Trm generation but produced recirculating memory populations normally. ICOS deficiency or ICOS-L blockade compromised establishment of CD8+ Trm cells but not their maintenance. ICOS ligation during CD8+ T cell priming did not determine Trm induction; rather, effector CD8+ T cells showed reduced Trm differentiation after seeding into Icosl-/- mice. IcosYF/YF CD8+ T cells were compromised in Trm generation, indicating a critical role for PI3K signaling. Modest transcriptional changes in the few Icos-/- Trm cells suggest that ICOS-PI3K signaling primarily enhances the efficiency of CD8+ T cell tissue residency. Thus, local ICOS signaling promotes production of Trm cells, providing insight into the contribution of costimulatory signals in the generation of tissue-resident populations.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Inducible T-Cell Co-Stimulator Protein/metabolism , Memory T Cells/immunology , Adoptive Transfer , Animals , Antibodies, Blocking/metabolism , Cell Differentiation , Cells, Cultured , Inducible T-Cell Co-Stimulator Ligand/immunology , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Lymphocyte Activation , Mice , Mice, Inbred C57BL , Mice, Knockout , Phosphatidylinositol 3-Kinases/metabolism , Signal TransductionABSTRACT
The inducible T-cell co-stimulator (ICOS) is a T-cell receptor that, once bound to ICOS ligand (ICOSL) expressed on several cell types including the B-cell lineage, plays a decisive role in adaptive immunity by regulating the interplay between B and T cells. In addition to its immunomodulatory functions, we have shown that ICOS/ICOSL signalling can inhibit the activity of osteoclasts, unveiling a novel mechanism of lymphocyte-bone cells interactions. ICOS and ICOSL can also be found as soluble forms, namely sICOS and sICOSL. Here we show that: (i) levels of sICOS and sICOSL are increased in multiple myeloma (MM) compared to monoclonal gammopathy of undetermined significance and smouldering MM; (ii) levels of sICOS and sICOSL variably correlate with several markers of tumour burden; and (iii) sICOS levels tend to be higher in Durie-Salmon stage II/III versus stage I MM and correlate with overall survival as an independent variable. Moreover, surface ICOS and ICOSL are expressed in both myeloma cells and normal plasma cells, where they probably regulate different functional stages. Finally, ICOSL triggering inhibits the migration of myeloma cell lines in vitro and the growth of ICOSL+ MOPC-21 myeloma cells in vivo. These results suggest that ICOS and ICOSL represent novel markers and therapeutic targets for MM.
Subject(s)
Multiple Myeloma , Humans , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/metabolism , Ligands , Multiple Myeloma/metabolism , T-Lymphocytes , Tumor MicroenvironmentABSTRACT
The liver capacity to recover from acute liver injury is a critical factor in the development of acute liver failure (ALF) caused by viral infections, ischemia/reperfusion or drug toxicity. Liver healing requires the switching of pro-inflammatory monocyte-derived macrophages(MoMFs) to a reparative phenotype. However, the mechanisms involved are still incompletely characterized. In this study we investigated the contribution of T-lymphocyte/macrophage interaction through the co-stimulatory molecule Inducible T-cell co-stimulator (ICOS; CD278) and its ligand (ICOSL; CD275) in modulating liver repair. The role of ICOS/ICOSL dyad was investigated during the recovery from acute liver damage induced by a single dose of carbon tetrachloride (CCl4). Flow cytometry of non-parenchymal liver cells obtained from CCl4-treated wild-type mice revealed that the recovery from acute liver injury associated with a specific up-regulation of ICOS in CD8+ T-lymphocytes and with an increase in ICOSL expression involving CD11bhigh/F4-80+ hepatic MoMFs. Although ICOS deficiency did not influence the severity of liver damage and the evolution of inflammation, CCl4-treated ICOS knockout (ICOS-/- ) mice showed delayed clearance of liver necrosis and increased mortality. These animals were also characterized by a significant reduction of hepatic reparative MoMFs due to an increased rate of cell apoptosis. An impaired liver healing and loss of reparative MoMFs was similarly evident in ICOSL-deficient mice or following CD8+ T-cells ablation in wild-type mice. The loss of reparative MoMFs was prevented by supplementing CCl4-treated ICOS-/- mice with recombinant ICOS (ICOS-Fc) which also stimulated full recovery from liver injury. These data demonstrated that CD8+ T-lymphocytes play a key role in supporting the survival of reparative MoMFs during liver healing trough ICOS/ICOSL-mediated signaling. These observations open the possibility of targeting ICOS/ICOSL dyad as a novel tool for promoting efficient healing following acute liver injury.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Inducible T-Cell Co-Stimulator Protein/metabolism , Liver Failure, Acute/immunology , Liver Regeneration/immunology , Macrophages/immunology , Animals , CD8-Positive T-Lymphocytes/metabolism , Carbon Tetrachloride/administration & dosage , Carbon Tetrachloride/toxicity , Cell Communication/immunology , Disease Models, Animal , Humans , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/genetics , Liver/cytology , Liver/drug effects , Liver/immunology , Liver/pathology , Liver Failure, Acute/chemically induced , Liver Failure, Acute/pathology , Macrophages/metabolism , Male , Mice , Mice, Knockout , Signal Transduction/immunologyABSTRACT
Mouse embryonic stem cells (mESCs) can be maintained in vitro in defined N2B27 medium supplemented with two chemical inhibitors for GSK3 and MEK (2i) and the cytokine leukemia inhibitory factor (LIF), which act synergistically to promote self-renewal and pluripotency. Here, we find that genetic deletion of the four genes encoding the TCF/LEF transcription factors confers mESCs with the ability to self-renew in N2B27 medium alone. TCF/LEF quadruple knockout (qKO) mESCs display dysregulation of several genes, including Aire, Dnmt3l, and IcosL, located adjacent to each other within a topologically associated domain (TAD). Aire, Dnmt3l, and IcosL appear to be regulated by TCF/LEF in a ß-catenin independent manner. Moreover, downregulation of Aire and Dnmt3l in wild-type mESCs mimics the loss of TCF/LEF and increases mESC survival in the absence of 2iL. Hence, this study identifies TCF/LEF effectors that mediate exit from the pluripotent state.
Subject(s)
Cell Self Renewal , Hepatocyte Nuclear Factor 1-alpha/genetics , Lymphoid Enhancer-Binding Factor 1/genetics , Transcription Factor 7-Like 1 Protein/genetics , Transcription Factor 7-Like 2 Protein/genetics , Animals , Benzamides/pharmacology , Cell Self Renewal/drug effects , Culture Media/chemistry , Culture Media/pharmacology , DNA (Cytosine-5-)-Methyltransferases/antagonists & inhibitors , DNA (Cytosine-5-)-Methyltransferases/genetics , DNA (Cytosine-5-)-Methyltransferases/metabolism , Diphenylamine/analogs & derivatives , Diphenylamine/pharmacology , Down-Regulation/drug effects , Gene Editing , Hepatocyte Nuclear Factor 1-alpha/deficiency , Hepatocyte Nuclear Factor 1-alpha/metabolism , Inducible T-Cell Co-Stimulator Ligand/antagonists & inhibitors , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Ligand/metabolism , Lymphoid Enhancer-Binding Factor 1/deficiency , Lymphoid Enhancer-Binding Factor 1/metabolism , Mice , Mouse Embryonic Stem Cells/cytology , Mouse Embryonic Stem Cells/metabolism , Octamer Transcription Factor-3/genetics , Octamer Transcription Factor-3/metabolism , Pyridines/pharmacology , Pyrimidines/pharmacology , RNA Interference , RNA, Small Interfering/metabolism , Transcription Factor 7-Like 1 Protein/deficiency , Transcription Factor 7-Like 1 Protein/metabolism , Transcription Factor 7-Like 2 Protein/deficiency , Transcription Factor 7-Like 2 Protein/metabolism , Transcription Factors/antagonists & inhibitors , Transcription Factors/genetics , Transcription Factors/metabolism , beta Catenin/deficiency , beta Catenin/genetics , AIRE ProteinABSTRACT
RATIONALE: Neutrophils constitute massive cellular constituents in inflammatory human gastric cancer (GC) tissues, but their roles in pathogenesis of inflammatory T helper (Th) subsets are still unknown. METHODS: Flow cytometry analysis and immunohistochemistry were used to analyze the responses and phenotypes of neutrophils in different samples from 51 patients with GC. Kaplan-Meier plots and Multivariate analysis for the survival of patients were used by log-rank tests and Cox proportional hazards models. Neutrophils and CD4+ T cells were purified and cultured for ex vivo, in vitro and in vivo regulation and function assays. RESULTS: GC patients exhibited increased tumoral neutrophil infiltration with GC progression and poor patient prognosis. Intratumoral neutrophils accumulated in GC tumors via CXCL6/CXCL8-CXCR1-mediated chemotaxis, and expressed activated molecule CD54 and co-signaling molecule B7-H2. Neutrophils induced by tumors strongly expressed CD54 and B7-H2 in both dose- and time-dependent manners, and a close correlation was obtained between the expressions of CD54 and B7-H2 on intratumoral neutrophils. Tumor-derived tumor necrosis factor-α (TNF-α) promoted neutrophil activation and neutrophil B7-H2 expression through ERK-NF-κB pathway, and a significant correlation was found between the levels of TNF-α and CD54+ or B7-H2+ neutrophils in tumor tissues. Tumor-infiltrating and tumor-conditioned neutrophils effectively induced IL-17A-producing Th subset polarization through a B7-H2-dependent manner ex vivo and these polarized IL-17A-producing Th cells exerted protumorigenic roles by promoting GC tumor cell proliferation via inflammatory molecule IL-17A in vitro, which promoted the progression of human GC in vivo; these effects could be reversed when IL-17A is blocked. Moreover, increased B7-H2+ neutrophils and IL-17A in tumors were closely related to advanced GC progression and predicted poor patient survival. CONCLUSION: We illuminate novel underlying mechanisms that TNF-α-activated neutrophils link B7-H2 to protumorigenic IL-17A-producing Th subset polarization in human GC. Blocking this pathological TNF-α-B7-H2-IL-17A pathway may be useful therapeutic strategies for treating GC.
Subject(s)
Inducible T-Cell Co-Stimulator Ligand/metabolism , Interleukin-17/metabolism , Neutrophil Activation , Neutrophils/immunology , Stomach Neoplasms/pathology , T-Lymphocytes, Helper-Inducer/immunology , Tumor Necrosis Factor-alpha/metabolism , Animals , Apoptosis , Cell Proliferation , Female , Gene Expression Regulation, Neoplastic , Humans , Inducible T-Cell Co-Stimulator Ligand/genetics , Mice , Mice, Inbred NOD , Mice, SCID , Prognosis , Stomach Neoplasms/immunology , Stomach Neoplasms/metabolism , Survival Rate , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha/genetics , Xenograft Model Antitumor AssaysABSTRACT
Autosomal recessive mutations in Inducible T Cell Costimulator Ligand (ICOSLG) result in a combined immunodeficiency syndrome of humans, saliently marked by recurrent respiratory tract infections and significant disease with DNA-based viruses at epithelial barriers, including human papillomavirus (HPV). These features are also seen in persons with loss of function of the complementary gene, ICOS. The infection phenotypes associated with these natural experiments disclose a critical role of the corresponding proteins, ICOSL and ICOS, in human immunity at mucocutaneous barriers. Here, we review the syndromes of ICOSL and ICOS deficiency and explore the mechanisms by which the ICOSL:ICOS axis mediates epithelial host defenses.
Subject(s)
Disease Resistance/genetics , Epithelium/immunology , Epithelium/metabolism , Host-Pathogen Interactions/genetics , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Ligand/metabolism , Animals , Disease Resistance/immunology , Disease Susceptibility/immunology , Genetic Predisposition to Disease , Genotype , Germinal Center/immunology , Germinal Center/metabolism , Host-Pathogen Interactions/immunology , Humans , Infections/etiology , Infections/metabolism , Mutation , Organ SpecificityABSTRACT
Genome-wide association studies have identified ICOSLG, which encodes the inducible costimulator ligand (ICOSLG or ICOSL) as a susceptibility locus for inflammatory bowel disease. ICOSL has been implicated in the enhancement of pattern recognition receptor signaling in dendritic cells, induction of IL-10 production by CD4 T cells, and the generation of high-affinity antibodies to specific antigens-all of which can potentially explain its involvement in gastrointestinal inflammation. Here, we show that murine ICOSL deficiency results in significant enrichment of IL-10-producing CD4 T cells particularly in the proximal large intestine. Transient depletion of IL-10-producing cells from adult ICOSL-deficient mice induced severe colonic inflammation that was prevented when mice were first treated with metronidazole. ICOSL-deficient mice displayed reduced IgA and IgG antibodies in the colon mucus and impaired serum antibody recognition of microbial antigens, including flagellins derived from mucus-associated bacteria of the Lachnospiraceae family. Confirming the synergy between ICOSL and IL-10, ICOSL deficiency coupled with CD4-specific deletion of the Il10 gene resulted in juvenile onset colitis that was impeded when pups were fostered by ICOSL-sufficient dams. In this setting, we found that both maternally acquired and host-derived antibodies contribute to the life anti-commensal antibody repertoire that mediates this protection in early life. Collectively, our findings reveal a partnership between ICOSL-dependent anti-commensal antibodies and IL-10 in adaptive immune regulation of the microbiota in the large intestine. Furthermore, we identify ICOSL deficiency as an effective platform for exploring the functions of anti-commensal antibodies in host-microbiota mutualism.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , Gastrointestinal Microbiome/immunology , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inflammatory Bowel Diseases/immunology , Interleukin-10/metabolism , Animals , Antibodies, Bacterial/immunology , Antigens, Bacterial/immunology , CD4-Positive T-Lymphocytes/metabolism , Colon/immunology , Colon/microbiology , Colon/pathology , Disease Models, Animal , Female , Host Microbial Interactions/immunology , Humans , Inducible T-Cell Co-Stimulator Ligand/genetics , Inducible T-Cell Co-Stimulator Protein/genetics , Inducible T-Cell Co-Stimulator Protein/metabolism , Inflammatory Bowel Diseases/microbiology , Inflammatory Bowel Diseases/pathology , Interleukin-10/genetics , Intestinal Mucosa/immunology , Intestinal Mucosa/microbiology , Intestinal Mucosa/pathology , Male , Mice , Mice, Knockout , Signal Transduction/immunology , Symbiosis/immunologyABSTRACT
Group 2 innate lymphoid cells (ILC2s) are recognized as key controllers and effectors of type 2 inflammation. Mesenchymal stem cells (MSCs) have been shown to alleviate type 2 inflammation by modulating T lymphocyte subsets and decreasing TH 2 cytokine levels. However, the effects of MSCs on ILC2s have not been investigated. In this study, we investigated the potential immunomodulatory effects of MSCs on ILC2s in peripheral blood mononuclear cells (PBMCs) from allergic rhinitis patients and healthy subjects. We further investigated the mechanisms involved in the MSC modulation using isolated lineage negative (Lin- ) cells. PBMCs and Lin- cells were cocultured with induced pluripotent stem cell-derived MSCs (iPSC-MSCs) under the stimulation of epithelial cytokines IL-25 and IL-33. And the ILC2 levels and functions were examined and the possible mechanisms were investigated based on regulatory T (Treg) cells and ICOS-ICOSL pathway. iPSC-MSCs successfully decreased the high levels of IL-13, IL-9, and IL-5 in PBMCs in response to IL-25, IL-33, and the high percentages of IL-13+ ILC2s and IL-9+ ILC2s in response to epithelial cytokines were significantly reversed after the treatment of iPSC-MSCs. However, iPSC-MSCs were found directly to enhance ILC2 levels and functions via ICOS-ICOSL interaction in Lin- cells and pure ILC2s. iPSC-MSCs exerted their inhibitory effects on ILC2s via activating Treg cells through ICOS-ICOSL interaction. The MSC-induced Treg cells then suppressed ILC2s by secreting IL-10 in the coculture system. This study revealed that human MSCs suppressed ILC2s via Treg cells through ICOS-ICOSL interaction, which provides further insight to regulate ILC2s in inflammatory disorders.
Subject(s)
Mesenchymal Stem Cells , T-Lymphocytes, Regulatory , Cytokines/metabolism , Humans , Immunity, Innate , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/metabolism , Leukocytes, Mononuclear , Lymphocytes , Mesenchymal Stem Cells/metabolism , T-Lymphocytes, Regulatory/metabolismABSTRACT
The interaction between the T-lymphocyte costimulatory molecule ICOS and its ligand (ICOS-L) is needed for efficient immune responses, but expression levels are tightly controlled, as altered expression of ICOS or ICOS-L may lead to immunodeficiency, or favor autoimmune diseases and tumor growth. Using cells of mouse B cell lymphoma (M12.C3) and melanoma (B16), or hamster CHO cells transfected with various forms of mouse ICOS-L, and ICOS+ T cell lines, we show that, within minutes, ICOS induces significant downmodulation of surface ICOS-L that is largely mediated by endocytosis and trans-endocytosis. So, after interaction with ICOS+ cells, ICOS-L was found inside permeabilized cells, or in cell lysates, with significant transfer of ICOS from ICOS+ T cells to ICOS-L-expressing cells, and simultaneous loss of surface ICOS by the T cells. Data from cells expressing ICOS-L mutants show that conserved, functionally important residues in the cytoplasmic domain of mouse ICOS-L (Arg300 , Ser307 and Tyr308 ), or removal of ICOS-L cytoplasmic tail have minor effect on its internalization. Internalization was dependent on temperature, and was partially dependent on actin polymerization, the GTPase dynamin, protein kinase C, or the integrity of lipid rafts. In fact, a fraction of ICOS-L was detected in lipid rafts. On the other hand, proteinase inhibitors had negligible effects on early modulation of ICOS-L from the cell surface. Our data add a new mechanism of control of ICOS-L expression to the regulation of ICOS-dependent responses.
Subject(s)
Endocytosis/physiology , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/metabolism , Animals , CHO Cells , Cricetinae , Cricetulus , Down-Regulation , Lymphocyte Activation/immunology , Melanoma, Experimental , Mice , Mice, Inbred C57BL , Mice, KnockoutABSTRACT
Viral infections are controlled, and very often cleared, by activated T lymphocytes. The inducible co-stimulator (ICOS) mediates its functions by binding to its ligand ICOSL, enhancing T-cell activation and optimal germinal center (GC) formation. Here, we show that ICOSL is heavily downmodulated during infection of antigen-presenting cells by different herpesviruses. We found that, in murine cytomegalovirus (MCMV), the immunoevasin m138/fcr-1 physically interacts with ICOSL, impeding its maturation and promoting its lysosomal degradation. This viral protein counteracts T-cell responses, in an ICOS-dependent manner, and limits virus control during the acute MCMV infection. Additionally, we report that blockade of ICOSL in MCMV-infected mice critically regulates the production of MCMV-specific antibodies due to a reduction of T follicular helper and GC B cells. Altogether, these findings reveal a novel mechanism evolved by MCMV to counteract adaptive immune surveillance, and demonstrates a role of the ICOS:ICOSL axis in the host defense against herpesviruses.
Subject(s)
Herpesviridae Infections/virology , Immune Evasion , Inducible T-Cell Co-Stimulator Ligand/metabolism , Inducible T-Cell Co-Stimulator Protein/metabolism , Muromegalovirus/physiology , T-Lymphocytes/immunology , Animals , MiceABSTRACT
CTLA4-Ig/abatacept dampens activation of naive T cells by blocking costimulation via CD28. It is an approved drug for rheumatoid arthritis but failed to deliver efficacy in a number of other autoimmune diseases. One explanation is that activated T cells rely less on CD28 signaling and use alternate coreceptors for effector function. ICOS is critical for activation of T-dependent humoral immune responses, which drives pathophysiology of IgG-mediated autoimmune diseases. In this study, we asked whether CD28 and ICOS play nonredundant roles for maintenance of T-dependent responses in mouse models. Using a hapten-protein immunization model, we show that during an ongoing germinal center response, combination treatment with CTLA4-Ig and ICOS ligand (ICOSL) blocking Ab completely dissolves ongoing germinal center responses, whereas single agents show only partial activity. Next, we took two approaches to engineer a therapeutic molecule that blocks both pathways. First, we engineered CTLA4-Ig to enhance binding to ICOSL while retaining affinity to CD80/CD86. Using a library approach, binding affinity of CTLA4-Ig to human ICOSL was increased significantly from undetectable to 15-42 nM; however, the affinity was still insufficient to completely block binding of ICOSL to ICOS. Second, we designed a bispecific costimulation inhibitor with high-affinity CTLA4 extracellular domains fused to anti-ICOSL Ab termed bifunctional costimulation inhibitor. With this bispecific approach, we achieved complete inhibition of CD80 and CD86 binding to CD28 as well as ICOS binding to ICOSL. Such bispecific molecules may provide greater therapeutic benefit in IgG-mediated inflammatory diseases compared with CTLA4-Ig alone.