CD1d-restricted Invariant Natural Killer Research Articles

Abstract 1386: CD1d-restricted iNKT cells promote liver metastasis from colon cancer

Abstract The liver is a prominent site for metastasis, yet little is known about the role of liver immunity in antimetastatic defense. Compared to other organs, the liver harbors a unique immune environment characterized by the abundance of innate cell lineages, including CD1d-restricted invariant natural killer T (iNKT) cells. To investigate the influence of iNKT cells on metastasis, we established an orthotopic model of colorectal cancer (CRC) with spontaneous dissemination to the liver. We induce the primary tumor by implanting CRC organoids into the colon mucosa of syngeneic C57BL/6 mice, resulting in hepatic macrometastases within 5 weeks. We observed that liver metastasis hardly occurred in mouse strains lacking iNKT cells (Cd1d−/− and Traj18−/− mice), while the growth of the primary tumor was comparable to that in wild-type mice. These results indicate a tumor-promoting function for hepatic iNKT cells. We uncovered that hepatic iNKT cells respond to disseminating cancer by producing IL-4 and IL-13, two cytokines known to activate hepatic stellate cells (HSCs) into fibrogenic myofibroblasts. Indeed, we saw rapid activation of HSCs and deposition of extracellular matrix upon arrival of cancer cells in the liver of wild-type but not iNKT cell-deficient mice. We propose that hepatic iNKT cells contribute to the formation of a supportive niche for disseminated tumor cells by activating HSCs early in the development of hepatic metastasis. Taken together, our findings highlight an essential tumor-promoting role for iNKT cells during the development of liver metastasis and contribute to our understanding of immune regulation of metastasis. Citation Format: Marc Nater, Giulia Lucchiari, Michael Brügger, Tomas Valenta, Virginia Cecconi, Geo Forni, Stephan Benke, Maries van den Broek. CD1d-restricted iNKT cells promote liver metastasis from colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1386.

Opposing Roles of DCs and iNKT Cells in the Induction of Foxp3 Expression by MLN CD25+CD4+ T Cells during IFNγ-Driven Colitis

We have previously shown that a deficiency of CD1d-restricted invariant natural killer T (iNKT) cells exacerbates dextran sulfate sodium (DSS)-induced colitis in Yeti mice that exhibit IFNγ-mediated hyper-inflammation. Although iNKT cell-deficiency resulted in reduced Foxp3 expression by mesenteric lymph node (MLN) CD4+ T cells in DSS-treated Yeti mice, the cellular mechanisms that regulate Foxp3 expression by CD25+CD4+ T cells during intestinal inflammation remain unclear. We found that Foxp3-CD25+CD4+ T cells expressing Th1 and Th17 phenotypic hallmarks preferentially expanded in the MLNs of DSS-treated Yeti/CD1d knockout (KO) mice. Moreover, adoptive transfer of Yeti iNKT cells into iNKT cell-deficient Jα18 KO mice effectively suppressed the expansion of MLN Foxp3-CD25+CD4+ T cells during DSS-induced colitis. Interestingly, MLN dendritic cells (DCs) purified from DSS-treated Yeti/CD1d KO mice promoted the differentiation of naive CD4+ T cells into Foxp3-CD25+CD4+ T cells rather than regulatory T (Treg) cells, indicating that MLN DCs might mediate Foxp3+CD25+CD4+ T cell expansion in iNKT cell-sufficient Yeti mice. Furthermore, we showed that Foxp3-CD25+CD4+ T cells were pathogenic in DSS-treated Yeti/CD1d KO mice. Our result suggests that pro-inflammatory DCs and CD1d-restricted iNKT cells play opposing roles in Foxp3 expression by MLN CD25+CD4+ T cells during IFNγ-mediated intestinal inflammation, with potential therapeutic implications.

Longitudinal analysis of invariant natural killer T cell activation reveals a cMAF-associated transcriptional state of NKT10 cells.

Innate T cells, including CD1d-restricted invariant natural killer T (iNKT) cells, are characterized by their rapid activation in response to non-peptide antigens, such as lipids. While the transcriptional profiles of naive, effector, and memory adaptive T cells have been well studied, less is known about the transcriptional regulation of different iNKT cell activation states. Here, using single-cell RNA-sequencing, we performed longitudinal profiling of activated murine iNKT cells, generating a transcriptomic atlas of iNKT cell activation states. We found that transcriptional signatures of activation are highly conserved among heterogeneous iNKT cell populations, including NKT1, NKT2, and NKT17 subsets, and human iNKT cells. Strikingly, we found that regulatory iNKT cells, such as adipose iNKT cells, undergo blunted activation and display constitutive enrichment of memory-like cMAF+ and KLRG1+ populations. Moreover, we identify a conserved cMAF-associated transcriptional network among NKT10 cells, providing novel insights into the biology of regulatory and antigen-experienced iNKT cells.

TCR-engineered iNKT cells induce robust antitumor response by dual targeting cancer and suppressive myeloid cells.

Adoptive immunotherapy with T cells engineered with tumor-specific T cell receptors (TCRs) holds promise for cancer treatment. However, suppressive cues generated in the tumor microenvironment (TME) can hinder the efficacy of these therapies, prompting the search for strategies to overcome these detrimental conditions and improve cellular therapeutic approaches. CD1d-restricted invariant natural killer T (iNKT) cells actively participate in tumor immunosurveillance by restricting suppressive myeloid populations in the TME. Here, we showed that harnessing iNKT cells with a second TCR specific for a tumor-associated peptide generated bispecific effectors for CD1d- and major histocompatibility complex (MHC)-restricted antigens in vitro. Upon in vivo transfer, TCR-engineered iNKT (TCR-iNKT) cells showed the highest efficacy in restraining the progression of multiple tumors that expressed the cognate antigen compared with nontransduced iNKT cells or CD8+ T cells engineered with the same TCR. TCR-iNKT cells achieved robust cancer control by simultaneously modulating intratumoral suppressive myeloid populations and killing malignant cells. This dual antitumor function was further enhanced when the iNKT cell agonist α-galactosyl ceramide (α-GalCer) was administered as a therapeutic booster through a platform that ensured controlled delivery at the tumor site, named multistage vector (MSV). These preclinical results support the combination of tumor-redirected TCR-iNKT cells and local α-GalCer boosting as a potential therapy for patients with cancer.

Human iNKT Cells Modulate Macrophage Survival and Phenotype.

CD1d-restricted invariant Natural Killer T (iNKT) cells are unconventional innate-like T cells whose functions highly depend on the interactions they establish with other immune cells. Although extensive studies have been reported on the communication between iNKT cells and macrophages in mice, less data is available regarding the relevance of this crosstalk in humans. Here, we dove into the human macrophage-iNKT cell axis by exploring how iNKT cells impact the survival and polarization of pro-inflammatory M1-like and anti-inflammatory M2-like monocyte-derived macrophages. By performing in vitro iNKT cell-macrophage co-cultures followed by flow cytometry analysis, we demonstrated that antigen-stimulated iNKT cells induce a generalized activated state on all macrophage subsets, leading to upregulation of CD40 and CD86 expression. CD40L blocking with a specific monoclonal antibody prior to co-cultures abrogated CD40 and CD86 upregulation, thus indicating that iNKT cells required CD40-CD40L co-stimulation to trigger macrophage activation. In addition, activated iNKT cells were cytotoxic towards macrophages in a CD1d-dependent manner, killing M1-like macrophages more efficiently than their naïve M0 or anti-inflammatory M2-like counterparts. Hence, this work highlighted the role of human iNKT cells as modulators of macrophage survival and phenotype, untangling key features of the human macrophage-iNKT cell axis and opening perspectives for future therapeutic modulation.

Expansion of invariant natural killer T cells from systemic lupus erythematosus patients by alpha-Galactosylceramide and IL-15

CD1d-restricted invariant natural killer T cells (iNKT cells) may play an important role in the pathogenesis of systemic lupus erythematosus (SLE). Interleukin (IL)-15 is a pro-inflammatory cytokine which is over-expressed in SLE patients. In the present study, we investigated the iNKT cell expansion of mononuclear cells (MNCs) from SLE patients following 10 days’ culture with α-galactosylceramide (α-Galcer) and /or IL-15. We sought to determine the phenotypic and functional characteristics of the expanded iNKT cells compared to healthy controls and correlated with disease activity. We observed that 1. The percentages of Vα24+/Vβ11+ iNKT cells following 10-day incubation was lower in SLE groups compared to controls; 2. The percentages and absolute numbers of Vα24+/Vβ11+ iNKT cells were expanded by α-galactosylceramide (α-Galcer), and further enhanced with IL-15 in SLE patient, but the effect of IL-15 was much lower than controls; 3.IL-15 +α-Galcer expanded CD3+/CD56+ NKT-like cells from SLE patients, especially with active disease 4. The CD161+ Vα24+/Vβ11+ iNKT cells in SLE were more responsive to α-Galcer stimulation than the CD161- counterpart; 5. IL-15 decreased apoptosis of α-Galcer activated SLE iNKT cells; 6. IL-15 enhanced CD69, CD1d and CD11a expression on α-Galcer treated iNKT cells; 7. The IL-4 production of iNKT cells was decreased in SLE patients compared to controls; 8. IL-15 increased IFN-γ and IL-4 production of SLE iNKT cells; 8. IL-15 failed to augment the ability of iNKT cells to aid NK-mediated K562 cytolysis in SLE patients; 9. CD161 positivity, granzyme B and perforin expression of α-Galcer+IL-15 expanded iNKT cells correlated with C3 levels in SLE patients. Taken together, our results demonstrated numeric and functional deficiency of iNKT cells and their response to IL-15 in SLE patients. Our finding may provide insight for using adoptive iNKT cell therapy in autoimmune diseases.

High Dimensional Single-Cell Analysis Reveals iNKT Cell Developmental Trajectories and Effector Fate Decision

CD1d-restricted invariant Natural Killer T (iNKT) cells represent a unique class of T lymphocytes endowed with potent regulatory and effector immune functions. Although these functions are acquired during thymic ontogeny, the sequence of events that gives rise to discrete effector subsets remains unclear. Using an unbiased single-cell transcriptomic analysis combined with functional assays, we reveal an unappreciated diversity among thymic iNKT cells, especially among iNKT1 cells. Mathematical modeling and biological methods unravel a developmental map whereby iNKT2 cells constitute a transient branching point toward the generation of iNKT1 and iNKT17 cells, which reconciles the two previously proposed models. In addition, we identify the transcription co-factor Four-and-a-half LIM domains protein 2 (FHL2) as a critical cell-intrinsic regulator of iNKT1 specification. Thus, these data illustrate the changing transcriptional network that guides iNKT cell effector fate.

ER stress in antigen-presenting cells promotes NKT cell activation through endogenous neutral lipids.

CD1d‐restricted invariant natural killer T (iNKT) cells constitute a common glycolipid‐reactive innate‐like T‐cell subset with a broad impact on innate and adaptive immunity. While several microbial glycolipids are known to activate iNKT cells, the cellular mechanisms leading to endogenous CD1d‐dependent glycolipid responses remain largely unclear. Here, we show that endoplasmic reticulum (ER) stress in APCs is a potent inducer of CD1d‐dependent iNKT cell autoreactivity. This pathway relies on the presence of two transducers of the unfolded protein response: inositol‐requiring enzyme‐1a (IRE1α) and protein kinase R‐like ER kinase (PERK). Surprisingly, the neutral but not the polar lipids generated within APCs undergoing ER stress are capable of activating iNKT cells. These data reveal that ER stress is an important mechanism to elicit endogenous CD1d‐restricted iNKT cell responses through induction of distinct classes of neutral lipids.

High Dimensional Single-Cell Analysis Reveals iNKT Cell Developmental Trajectories and Effector Fate Decision

CD1d-restricted invariant Natural Killer T (iNKT) cells represent a unique class of T lymphocytes endowed with potent regulatory and effector immune functions. Although these functions are acquired during thymic ontogeny, the sequence of events that give rise to discrete effector subsets remains unclear. Using an unbiased single-cell transcriptomic analysis combined with functional assays, we revealed an unappreciated diversity among thymic iNKT cells, especially among iNKT1 cells. Mathematical modelling and biological methods unravelled a developmental map whereby iNKT2 cells constitute a transient branching point towards the generation of iNKT1 and iNKT17 cells, which reconciles the two previously proposed models. In addition, we identified the transcription co-factor Four-and-a-half LIM domains protein 2 (FHL2) as a critical cell-intrinsic regulator of iNKT1 specification. Thus, these data illustrate the changing transcriptional network that guides iNKT cell effector fate.

Tweaking α -Galactoceramides: Probing the Dynamical Mechanisms of Improved Recognition for Invariant Natural Killer T-cell Receptor in Cancer Immunotherapeutics.

The last few decades have witnessed groundbreaking research geared towards immune surveillance mechanisms and have yielded significant improvements in the field of cancer immunotherapy. This approach narrows down on the development of therapeutic agents that either activate or enhance the recognitive function of the immune system to facilitate the destruction of malignant cells. The α -galactosylceramide derivative, KRN7000, is an immunotherapeutic agent that has gained attention due to its pharmacological ability to activate CD1d-restricted invariant natural killer T(iNKT) cells with notable potency against cancer cells in mouse models; a therapeutic success was not well replicated in human models. Dual structural modification of KRN7000 entailing the incorporation of hydrocinnamoyl ester on C6" and C4-OH truncation of the sphingoid base led to the development of AH10-7 which, interestingly, exhibited high potency in human cells. Therefore, to gain molecular insights into the structural dynamics and selective mechanisms of AH10-7 for human variants, we employed integrative molecular dynamics simulations and thermodynamic calculations to investigate the inhibitory activities of KRN7000 andAH10-7 on hTCR-CD1d towards activating iNKT. Interestingly, our findings revealed that AH10-7 exhibited higher affinity binding and structural effects on hTCR-CD1d, as mediated by the incorporated hydrocinnamoyl ester moiety which accounted for stronger intermolecular interactions with 'non-common' binding site residues. Findings extracted from this study further reveal important molecular and structural perspectives that could aid in the design of novel α-GalCer derivatives for cancer immunotherapeutics.

Adipocytes harbor a glucosylceramide biosynthesis pathway involved in iNKT cell activation

BackgroundNatural killer T (NKT) cells in adipose tissue (AT) contribute to whole body energy homeostasis. ResultsInhibition of the glucosylceramide synthesis in adipocytes impairs iNKT cell activity. ConclusionGlucosylceramide biosynthesis pathway is important for endogenous lipid antigen activation of iNKT cells in adipocytes. SignificanceUnraveling adipocyte-iNKT cell communication may help to fight obesity-induced AT dysfunction.Overproduction and/or accumulation of ceramide and ceramide metabolites, including glucosylceramides, can lead to insulin resistance. However, glucosylceramides also fulfill important physiological functions. They are presented by antigen presenting cells (APC) as endogenous lipid antigens via CD1d to activate a unique lymphocyte subspecies, the CD1d-restricted invariant (i) natural killer T (NKT) cells. Recently, adipocytes have emerged as lipid APC that can activate adipose tissue-resident iNKT cells and thereby contribute to whole body energy homeostasis. Here we investigate the role of the glucosylceramide biosynthesis pathway in the activation of iNKT cells by adipocytes.UDP-glucose ceramide glucosyltransferase (Ugcg), the first rate limiting step in the glucosylceramide biosynthesis pathway, was inhibited via chemical compounds and shRNA knockdown in vivo and in vitro. β-1,4-Galactosyltransferase (B4Galt) 5 and 6, enzymes that convert glucosylceramides into potentially inactive lactosylceramides, were subjected to shRNA knock down. Subsequently, (pre)adipocyte cell lines were tested in co-culture experiments with iNKT cells (IFNγ and IL4 secretion).Inhibition of Ugcg activity shows that it regulates presentation of a considerable fraction of lipid self-antigens in adipocytes. Furthermore, reduced expression levels of either B4Galt5 or -6, indicate that B4Galt5 is dominant in the production of cellular lactosylceramides, but that inhibition of either enzyme results in increased iNKT cell activation. Additionally, in vivo inhibition of Ugcg by the aminosugar AMP-DNM results in decreased iNKT cell effector function in adipose tissue.Inhibition of endogenous glucosylceramide production results in decreased iNKT cells activity and cytokine production, underscoring the role of this biosynthetic pathway in lipid self-antigen presentation by adipocytes.

TLR9-mediated dendritic cell activation uncovers mammalian ganglioside species with specific ceramide backbones that activate invariant natural killer T cells.

CD1d-restricted invariant natural killer T (iNKT) cells represent a heterogeneous population of lipid-reactive T cells that are involved in many immune responses, mediated through T-cell receptor (TCR)–dependent and/or independent activation. Although numerous microbial lipid antigens (Ags) have been identified, several lines of evidence have suggested the existence of relevant Ags of endogenous origin. However, the identification of their precise nature as well as the molecular mechanisms involved in their generation are still highly controversial and ill defined. Here, we identified two mammalian gangliosides—namely monosialoganglioside GM3 and disialoganglioside GD3—as endogenous activators for mouse iNKT cells. These glycosphingolipids are found in Toll-like receptor-stimulated dendritic cells (DC) as several species varying in their N-acyl fatty chain composition. Interestingly, their ability to activate iNKT cells is highly dependent on the ceramide backbone structure. Thus, both synthetic GM3 and GD3 comprising a d18:1-C24:1 ceramide backbone were able to activate iNKT cells in a CD1d-dependent manner. GM3 and GD3 are not directly recognized by the iNKT TCR and required the Ag presenting cell intracellular machinery to reveal their antigenicity. We propose a new concept in which iNKT cells can rapidly respond to pre-existing self-molecules after stress-induced structural changes in CD1d-expressing cells. Moreover, these gangliosides conferred partial protection in the context of bacterial infection. Thus, this report identified new biologically relevant lipid self-Ags for iNKT cells.

Abstract A151: Immunotherapy against cancer using iNKT ligands in combination with an attenuated Listeria monocytogenes in humanized mouse models

Abstract Listeria monocytogenes (Lm) is a gram-positive bacterium associated with gastrointestinal infections. Initially, infects epithelial cells in the small intestine, then migrates to other organs where it is taken up mainly by phagocytic cells. Once phagocytized, Lm may then be processed in the phagosome to generate peptides presented on MHC Class II for activation of Lm-specific CD4+ T-cell responses. Lm can also escape the phagosome, leading to secretion of protein antigens into the cytosol. These are processed and presented by MHC Class I molecules to CD8+ T-cells, generating strong protective immunity that can clear the infection and prevent subsequent reinfection. Lm has been exploited for its use as a vaccine vector, particularly for the delivery of tumor-associated antigens. Our previous work showed that the efficacy of Lm as an anti-umor therapeutic vaccine could be significantly improved by incorporation of synthetic glycolipid activators of CD1d-restricted invariant natural killer T-cells (iNKT-cells) into the bacteria. Using an attenuated Lm strain expressing the tumor-associated antigen Mage-B, this approach gave improvements in prevention of metastasis in the 4T1 breast carcinoma model in BALB/c mice, and also gave modest reductions in the size of primary tumors. Currently, our efforts are directed at further improving this approach to more effectively target primary tumors, and also at validating the effects in a more humanized experimental models. To this end, we are using two models: 1. human CD1d knock-in (hCD1dKI) mouse line in which the mouse CD1d protein has been replaced by human CD1d, and 2. VaKI mouse, which is a new model made by our lab that is a hCD1dKI mouse that has as a transgene the α-chain of the human iNKT TCR (Vα24Jα18) and is deficient of endogenous murine iNKT-cells due to a murine Jα18 KO. These mice models have diminished percentages of iNKT-cells closely resembling what is seen in most humans, and thus mount iNKT-cell responses that are comparable to those seen in humans. We found that incorporation of iNKT-cell activating glycolipids into Lm leads to improved iNKT-cell-dependent immune responses in our models, and induces synergistic antitumor immune mechanisms against implanted primary syngeneic tumors (B16 F10 and MC38). Besides, administration of iNKT-cell activating glycolipids in association with Lm did not induce anergy or exhaustion of the responding cells, thus enabling repeated treatment to boost antitumor immune responses. Citation Format: Noemi Alejandra Saavedra-Avila, Shalu Sharma, Christopher T. Johndrow, Tony Ng, Claudia Gravekamp, Steven A. Porcelli. Immunotherapy against cancer using iNKT ligands in combination with an attenuated Listeria monocytogenes in humanized mouse models [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A151.

The Role of Autophagy in iNKT Cell Development.

CD1d-restricted invariant natural killer T (iNKT) cells are innate-like T cells that express an invariant T cell receptor (TCR) α-chain and recognize self and foreign glycolipid antigens. They can rapidly respond to agonist activation and stimulate an extensive array of immune responses. Thymic development and function of iNKT cells are regulated by many different cellular processes, including autophagy, a self-degradation mechanism. In this mini review, we discuss the current understanding of how autophagy regulates iNKT cell development and effector lineage differentiation. Importantly, we propose that iNKT cell development is tightly controlled by metabolic reprogramming.

Enhanced Anti-lymphoma Activity of CAR19-iNKT Cells Underpinned by Dual CD19 and CD1d Targeting

SummaryChimeric antigen receptor anti-CD19 (CAR19)-T cell immunotherapy-induced clinical remissions in CD19+ B cell lymphomas are often short lived. We tested whether CAR19-engineering of the CD1d-restricted invariant natural killer T (iNKT) cells would result in enhanced anti-lymphoma activity. CAR19-iNKT cells co-operatively activated by CD1d- and CAR19-CD19-dependent interactions are more effective than CAR19-T cells against CD1d-expressing lymphomas in vitro and in vivo. The swifter in vivo anti-lymphoma activity of CAR19-iNKT cells and their enhanced ability to eradicate brain lymphomas underpinned an improved tumor-free and overall survival. CD1D transcriptional de-repression by all-trans retinoic acid results in further enhanced cytotoxicity of CAR19-iNKT cells against CD19+ chronic lymphocytic leukemia cells. Thus, iNKT cells are a highly efficient platform for CAR-based immunotherapy of lymphomas and possibly other CD1d-expressing cancers.

Lipid-Reactive T Cells in Immunological Disorders of the Lung.

Regulation of T cell-mediated immunity in the lungs is critical for prevention of immune-related lung disorders and for host protection from pathogens. While the prevalent view of pulmonary T cell responses is based on peptide recognition by antigen receptors, called T cell receptors (TCR), on the T cell surface in the context of classical major histocompatibility complex (MHC) molecules, novel pathways involving the presentation of lipid antigens by cluster of differentiation 1 (CD1) molecules to lipid-reactive T cells are emerging as key players in pulmonary immune system. Whereas, genetic conservation of group II CD1 (CD1d) in mouse and human genomes facilitated numerous in vivo studies of CD1d-restricted invariant natural killer T (iNKT) cells in lung diseases, the recent development of human CD1-transgenic mice has made it possible to examine the physiological roles of group I CD1 (CD1a-c) molecules in lung immunity. Here, we discuss current understanding of the biology of CD1-reactive T cells with a specific focus on their roles in several pulmonary disorders.

CD1d-Invariant Natural Killer T Cell-Based Cancer Immunotherapy: α-Galactosylceramide and Beyond.

CD1d-restricted invariant natural killer T (iNKT) cells are considered an attractive target for cancer immunotherapy. Upon their activation by glycolipid antigen and/or cytokines, iNKT cells can induce direct lysis of tumor cells but can also induce an antitumor immune response via their rapid production of proinflammatory cytokines that trigger the cytotoxic machinery of other components of the innate and adaptive immune system. Here, we provide an overview of various therapeutic approaches that have been evaluated or that are currently being developed and/or explored. These include administration of α-GalCer or alternative (glyco) lipid antigens, glycolipid-loaded antigen-presenting cells and liposomes, strategies that enhance CD1d expression levels or are based on ligation of CD1d, adoptive transfer of iNKT cells or chimeric antigen receptor iNKT cells, and tumor targeting of iNKT cells.

Activation and Regulation of B Cell Responses by Invariant Natural Killer T Cells.

CD1d-restricted invariant natural killer T (iNKT) cells play central roles in the activation and regulation of innate and adaptive immunity. Cytokine-mediated and CD1d-dependent interactions between iNKT cells and myeloid and lymphoid cells enable iNKT cells to contribute to the activation of multiple cell types, with important impacts on host immunity to infection and tumors and on the prevention of autoimmunity. Here, we review the mechanisms by which iNKT cells contribute to B cell maturation, antibody and cytokine production, and antigen presentation. Cognate interactions with B cells contribute to the rapid production of antibodies directed against conserved non-protein antigens resulting in rapid but short-lived innate humoral immunity. iNKT cells can also provide non-cognate help for the generation of antibodies directed against protein antigens, by promoting the activation of follicular helper T cells, resulting in long-lasting adaptive humoral immunity and B cell memory. iNKT cells can also regulate humoral immunity by promoting the development of autoreactive B cells into regulatory B cells. Depletions and functional impairments of iNKT cells are found in patients with infectious, autoimmune and malignant diseases associated with altered B cell function and in murine models of these conditions. The adjuvant and regulatory activities that iNKT cells have for B cells makes them attractive therapeutic targets for these diseases.

Functions of CD1d-Restricted Invariant Natural Killer T Cells in Antimicrobial Immunity and Potential Applications for Infection Control.

CD1d-restricted invariant natural killer T (iNKT) cells are innate-type lymphocytes that express a T-cell receptor (TCR) containing an invariant α chain encoded by the Vα14 gene in mice and Vα24 gene in humans. These iNKT cells recognize endogenous, microbial, and synthetic glycolipid antigens presented by the major histocompatibility complex (MHC) class I-like molecule CD1d. Upon TCR stimulation by glycolipid antigens, iNKT cells rapidly produce large amounts of cytokines, including interferon-γ (IFNγ) and interleukin-4 (IL-4). Activated iNKT cells contribute to host protection against a broad spectrum of microbial pathogens, and glycolipid-mediated stimulation of iNKT cells ameliorates many microbial infections by augmenting innate and acquired immunity. In some cases, however, antigen-activated iNKT cells exacerbate microbial infections by promoting pathogenic inflammation. Therefore, it is important to identify appropriate microbial targets for the application of iNKT cell activation as a treatment or vaccine adjuvant. Many studies have found that iNKT cell activation induces potent adjuvant activities promoting protective vaccine effects. In this review, we summarize the functions of CD1d-restricted iNKT cells in immune responses against microbial pathogens and describe the potential applications of glycolipid-mediated iNKT cell activation for preventing and controlling microbial infections.

Functional Invariant Natural Killer T Cells Secreting Cytokines Are Associated With Non-Progressive Human Immunodeficiency Virus-1 Infection but Not With Suppressive Anti-Retroviral Treatment.

CD1d restricted invariant natural killer T (iNKT) cells are important in the activation and regulation of immune responses. Limited information is available regarding the functional role of iNKT cells in the human immunodeficiency virus (HIV) disease progression. α-GalCer stimulated iNKT cells were characterized for their functionality in terms of cytokine production (IFN-γ, TNF-α, IL-2, IL-4, and IL-21) and CD107a expression in HIV-1 infected [23 long-term non progressors (LTNPs), 28 progressors, 18 patients before and after suppressive anti-retroviral treatment (ART)] along with 25 HIV-1 negative subjects using multicolor flow cytometry. The functional profile of α-GalCer stimulated iNKT cells was similar in LTNPs and healthy controls. The number of LTNPs showing functional response in terms of secretion of cytokines (IFN-γ/IL2/TNF-α) and CD107a expression was significantly higher than seen in the progressors. The cytokine secretion by the stimulated iNKT cells was predominantly Th1 type. The frequencies of iNKT cells showing secretion of IFN-γ or IL2 or TNF-α or expression of CD107a were higher in LTNPs (p < 0.05 for all) and also significantly associated with lower plasma viral load (p value ranged from 0.04 to 0.003) and higher CD4 count (p value ranged from 0.02 to <0.0001). The functional profile of the iNKT cells before and after ART did not differ significantly indicating absence of restoration of iNKT cells functionality after suppressive ART. The IL-4 and IL-21 secreting iNKT cells were rare in all study populations. The presence of functional iNKT cells secreting number of cytokines in non-progressive HIV infection could be one of the multiple factors required to achieve HIV control and hence have relevance in understanding the immunity in HIV infection. The failure of restoration of the iNKT functionality after ART should be potential area of future research.