NKT cell subsets as key participants in liver physiology and pathology

PubMed Central

Bandyopadhyay, Keya; Marrero, Idania; Kumar, Vipin

2016-01-01

Natural killer T (NKT) cells are innate-like lymphocytes that generally recognize lipid antigens and are enriched in microvascular compartments of the liver. NKT cells can be activated by self- or microbial-lipid antigens and by signaling through toll-like receptors. Following activation, NKT cells rapidly secrete pro-inflammatory or anti-inflammatory cytokines and chemokines, and thereby determine the milieu for subsequent immunity or tolerance. It is becoming clear that two different subsets of NKT cells—type I and type II—have different modes of antigen recognition and have opposing roles in inflammatory liver diseases. Here we focus mainly on the roles of both NKT cell subsets in the maintenance of immune tolerance and inflammatory diseases in liver. Furthermore, how the differential activation of type I and type II NKT cells influences other innate cells and adaptive immune cells to result in important consequences for tissue integrity is discussed. It is crucial that better reagents, including CD1d tetramers, be used in clinical studies to define the roles of NKT cells in liver diseases in patients. PMID:26972772

NKT Cells as an Ideal Anti-Tumor Immunotherapeutic

PubMed Central

Fujii, Shin-ichiro; Shimizu, Kanako; Okamoto, Yoshitaka; Kunii, Naoki; Nakayama, Toshinori; Motohashi, Shinichiro; Taniguchi, Masaru

2013-01-01

Human natural killer T (NKT) cells are characterized by their expression of an invariant T cell antigen receptor α chain variable region encoded by a Vα24Jα18 rearrangement. These NKT cells recognize α-galactosylceramide (α-GalCer) in conjunction with the MHC class I-like CD1d molecule and bridge the innate and acquired immune systems to mediate efficient and augmented immune responses. A prime example of one such function is adjuvant activity: NKT cells augment anti-tumor responses because they can rapidly produce large amounts of IFN-γ, which acts on NK cells to eliminate MHC negative tumors and also on CD8 cytotoxic T cells to kill MHC positive tumors. Thus, upon administration of α-GalCer-pulsed DCs, both MHC negative and positive tumor cells can be effectively eliminated, resulting in complete tumor eradication without tumor recurrence. Clinical trials have been completed in a cohort of 17 patients with advanced non-small cell lung cancers and 10 cases of head and neck tumors. Sixty percent of advanced lung cancer patients with high IFN-γ production had significantly prolonged median survival times of 29.3 months with only the primary treatment. In the case of head and neck tumors, 10 patients who completed the trial all had stable disease or partial responses 5 weeks after the combination therapy of α-GalCer-DCs and activated NKT cells. We now focus on two potential powerful treatment options for the future. One is to establish artificial adjuvant vector cells containing tumor mRNA and α-GalCer/CD1d. This stimulates host NKT cells followed by DC maturation and NK cell activation but also induces tumor-specific long-term memory CD8 killer T cell responses, suppressing tumor metastasis even 1 year after the initial single injection. The other approach is to establish induced pluripotent stem (iPS) cells that can generate unlimited numbers of NKT cells with adjuvant activity. Such iPS-derived NKT cells produce IFN-γ in vitro and in vivo upon

Pro-Inflammatory Activated Kupffer Cells by Lipids Induce Hepatic NKT Cells Deficiency through Activation-Induced Cell Death

PubMed Central

Tang, Tongfang; Sui, Yongheng; Lian, Min; Li, Zhiping; Hua, Jing

2013-01-01

Background Dietary lipids play an important role in the progression of non-alcoholic fatty liver disease (NAFLD) through alternation of liver innate immune response. Aims The present study was to investigate the effect of lipid on Kupffer cells phenotype and function in vivo and in vitro. And further to investigate the impact of lipid on ability of Kupffer cell lipid antigen presentation to activate NKT cells. Methods Wild type male C57BL/6 mice were fed either normal or high-fat diet. Hepatic steatosis, Kupffer cell abundance, NKT cell number and cytokine gene expression were evaluated. Antigen presentation assay was performed with Kupffer cells treated with certain fatty acids in vitro and co-cultured with NKT cells. Results High-fat diet induced hepatosteatosis, significantly increased Kupffer cells and decreased hepatic NKT cells. Lipid treatment in vivo or in vitro induced increase of pro-inflammatory cytokines gene expression and toll-like receptor 4 (TLR4) expression in Kupffer cells. Kupffer cells expressed high levels of CD1d on cell surface and only presented exogenous lipid antigen to activate NKT cells. Ability of Kupffer cells to present antigen and activate NKT cells was enhanced after lipid treatment. In addition, pro-inflammatory activated Kupffer cells by lipid treatment induced hepatic NKT cells activation-induced apoptosis and necrosis. Conclusion High-fat diet increase Kupffer cells number and induce their pro-inflammatory status. Pro-inflammatory activated Kupfffer cells by lipid promote hepatic NKT cell over-activation and cell death, which lead to further hepatic NKT cell deficiency in the development of NAFLD. PMID:24312613

Invariant NKT Cell Response to Dengue Virus Infection in Human

PubMed Central

Matangkasombut, Ponpan; Chan-in, Wilawan; Opasawaschai, Anunya; Pongchaikul, Pisut; Tangthawornchaikul, Nattaya; Vasanawathana, Sirijitt; Limpitikul, Wannee; Malasit, Prida; Duangchinda, Thaneeya; Screaton, Gavin; Mongkolsapaya, Juthathip

2014-01-01

Background Dengue viral infection is a global health threat without vaccine or specific treatment. The clinical outcome varies from asymptomatic, mild dengue fever (DF) to severe dengue hemorrhagic fever (DHF). While adaptive immune responses were found to be detrimental in the dengue pathogenesis, the roles of earlier innate events remain largely uninvestigated. Invariant natural killer T (iNKT) cells represent innate-like T cells that could dictate subsequent adaptive response but their role in human dengue virus infection is not known. We hypothesized that iNKT cells play a role in human dengue infection. Methods Blood samples from a well-characterized cohort of children with DF, DHF, in comparison to non-dengue febrile illness (OFI) and healthy controls at various time points were studied. iNKT cells activation were analyzed by the expression of CD69 by flow cytometry. Their cytokine production was then analyzed after α-GalCer stimulation. Further, the CD1d expression on monocytes, and CD69 expression on conventional T cells were measured. Results iNKT cells were activated during acute dengue infection. The level of iNKT cell activation associates with the disease severity. Furthermore, these iNKT cells had altered functional response to subsequent ex vivo stimulation with α-GalCer. Moreover, during acute dengue infection, monocytic CD1d expression was also upregulated and conventional T cells also became activated. Conclusion iNKT cells might play an early and critical role in the pathogenesis of severe dengue viral infection in human. Targeting iNKT cells and CD1d serve as a potential therapeutic strategy for severe dengue infection in the future. PMID:24945350

Fas/FasL interaction mediates imbalanced cytokine/cytotoxicity responses of iNKT cells against Jurkat cells.

PubMed

Dou, Rui; Hong, Zhenya; Tan, Xiaosheng; Hu, Fenfen; Ding, Yajie; Wang, Wei; Liang, Zhihui; Zhong, Rongrong; Wu, Xiongwen; Weng, Xiufang

2018-07-01

The rapid antitumor cytokine production and direct cytotoxicity confer invariant NKT (iNKT) cells ideal candidates for cancer therapy. However, the therapeutic potential of iNKT cells in T-cell malignant diseases remains elusive, as antigen presentation by T cells (T-T presentation) has been suggested to induce hyporesponsiveness of iNKT cells. In this study, we found discrepancies in iNKT cell responses against two T cell-origin cell lines (Jurkat and Molt-4). Human iNKT cells exhibited more intensive cytotoxicity and less efficient cytokine production in response to Fas-bearing Jurkat cells than those to the Fas-negative tumor cells (Molt-4 and myeloid-derived K562). The imbalanced cytokine/cytotoxicity responses of iNKT cells against Jurkat cells were CD1d-dependent and relied mostly on Fas/FasL interaction. The impairment in cytokine production could be overcome by Fas/FasL blocking antibodies and exogenous IL-2. Elevated CD1d levels as well as CD1d and Fas co-localization were found in T-cell lymphomas. However, defects in frequency and function of circulating iNKT cells were observed in the patients, which could be partly rescued by exogenous IL-2. Collectively, the Fas/FasL-dependent aberrant iNKT cell responses and the reversibility of the defects suggest the distinct iNKT cell manipulation in CD1d- and Fas-bearing T cell malignancies. Copyright © 2018. Published by Elsevier Ltd.

iNKT cells ameliorate human autoimmunity: Lessons from alopecia areata.

PubMed

Ghraieb, Amal; Keren, Aviad; Ginzburg, Alex; Ullmann, Yehuda; Schrum, Adam G; Paus, Ralf; Gilhar, Amos

2018-04-18

Alopecia areata (AA) is understood to be a CD8+/NKG2D+ T cell-dependent autoimmune disease. Here, we demonstrate that human AA pathogenesis of is also affected by iNKT10 cells, an unconventional T cell subtype whose number is significantly increased in AA compared to healthy human skin. AA lesions can be rapidly induced in healthy human scalp skin xenotransplants on Beige-SCID mice by intradermal injections of autologous healthy-donor PBMCs pre-activated with IL-2. We show that in this in vivo model, the development of AA lesions is prevented by recognized the iNKT cell activator, α-galactosylceramide (α-GalCer), which stimulates iNKT cells to expand and produce IL-10. Moreover, in pre-established humanized mouse AA lesions, hair regrowth is promoted by α-GalCer treatment through a process requiring both effector-memory iNKT cells, which can interact directly with CD8+/NKG2D+ T cells, and IL-10. This provides the first in vivo evidence in a humanized model of autoimmune disease that iNKT10 cells are key disease-protective lymphocytes. Since these regulatory NKT cells can both prevent the development of AA lesions and promote hair re-growth in established AA lesions, targeting iNKT10 cells may have preventive and therapeutic potential also in other autoimmune disorders related to AA. Copyright © 2018 Elsevier Ltd. All rights reserved.

Type II NKT Cells in Inflammation, Autoimmunity, Microbial Immunity, and Cancer

PubMed Central

Marrero, Idania; Ware, Randle; Kumar, Vipin

2015-01-01

Natural killer T cells (NKT) recognize self and microbial lipid antigens presented by non-polymorphic CD1d molecules. Two major NKT cell subsets, type I and II, express different types of antigen receptors (TCR) with distinct mode of CD1d/lipid recognition. Though type II NKT cells are less frequent in mice and difficult to study, they are predominant in human. One of the major subsets of type II NKT cells reactive to the self-glycolipid sulfatide is the best characterized and has been shown to induce a dominant immune regulatory mechanism that controls inflammation in autoimmunity and in anti-cancer immunity. Recently, type II NKT cells reactive to other self-glycolipids and phospholipids have been identified suggesting both promiscuous and specific TCR recognition in microbial immunity as well. Since the CD1d pathway is highly conserved, a detailed understanding of the biology and function of type II NKT cells as well as their interplay with type I NKT cells or other innate and adaptive T cells will have major implications for potential novel interventions in inflammatory and autoimmune diseases, microbial immunity, and cancer. PMID:26136748

NKT Cell Subsets Can Exert Opposing Effects in Autoimmunity, Tumor Surveillance and Inflammation

PubMed Central

Viale, Rachael; Ware, Randle; Maricic, Igor; Chaturvedi, Varun; Kumar, Vipin

2014-01-01

The innate-like natural killer T (NKT) cells are essential regulators of immunity. These cells comprise at least two distinct subsets and recognize different lipid antigens presented by the MHC class I like molecules CD1d. The CD1d-dependent recognition pathway of NKT cells is highly conserved from mouse to humans. While most type I NKT cells can recognize αGalCer and express a semi-invariant T cell receptor (TCR), a major population of type II NKT cells reactive to sulfatide utilizes an oligoclonal TCR. Furthermore TCR recognition features of NKT subsets are also distinctive with almost parallel as opposed to perpendicular footprints on the CD1d molecules for the type I and type II NKT cells respectively. Here we present a view based upon the recent studies in different clinical and experimental settings that while type I NKT cells are more often pathogenic, they may also be regulatory. On the other hand, sulfatide-reactive type II NKT cells mostly play an inhibitory role in the control of autoimmune and inflammatory diseases. Since the activity and cytokine secretion profiles of NKT cell subsets can be modulated differently by lipid ligands or their analogs, novel immunotherapeutic strategies are being developed for their differential activation for potential intervention in inflammatory diseases. PMID:25288922

Type 1 Diabetes and NKT Cells: A Report on the 3rd International Workshop on NKT Cells and CD1-Mediated Antigen Presentation, September 2004, Heron Island, QLD, Australia

PubMed Central

Fletcher, Julie M.; Jordan, Margaret A.; Baxter, Alan G.

2004-01-01

NKT cells play a major role in regulating the vigor and character of a broad range of immune responses. Defects in NKT cell numbers and function have been associated with type 1 diabetes, especially in the NOD mouse model. The 3rd International Workshop on NKT Cells and CD1-Mediated Antigen Presentation provided an opportunity for researchers in the field of NKT cell biology to discuss their latest results, many of which have direct relevance to understanding the etiology and pathogenesis of diabetes. PMID:17491677

Invariant NKT cells from HIV-1 or Mycobacterium tuberculosis-infected patients express an activated phenotype.

PubMed

Montoya, Carlos J; Cataño, Juan C; Ramirez, Zoraida; Rugeles, Maria T; Wilson, S Brian; Landay, Alan L

2008-04-01

The frequency, subsets and activation status of peripheral blood invariant NKT (iNKT) cells were evaluated in pulmonary tuberculosis (TB) patients and in chronically HIV-1-infected subjects. The absolute numbers of iNKT cells were significantly decreased in TB patients and in HIV-1+ individuals who were antiretroviral therapy naive or had detectable viremia despite receiving HAART. iNKT cell subset analysis demonstrated a decreased percentage of CD4(+) iNKT cells in HIV-1+ subjects, and a decreased percentage of double negative iNKT cells in TB patients. Peripheral blood iNKT cells from HIV-1+ and TB patients had significantly increased expression of CD69, CD38, HLA-DR, CD16, CD56, and CD62L. The expression of CD25 was significantly increased only on iNKT cells from TB patients. These findings indicate that peripheral blood iNKT cells in these two chronic infections show an up-regulated expression of activation markers, suggesting their role in the immune response to infection.

iNKT cell cytotoxic responses control T-lymphoma growth in vitro and in vivo

PubMed Central

Bassiri, Hamid; Das, Rupali; Guan, Peng; Barrett, David M.; Brennan, Patrick J.; Banerjee, Pinaki P.; Wiener, Susan J.; Orange, Jordan S.; Brenner, Michael B.; Grupp, Stephan A.; Nichols, Kim E.

2013-01-01

Invariant natural killer T (iNKT) cells comprise a lineage of CD1d-restricted glycolipid-reactive T lymphocytes with important roles in host immunity to cancer. iNKT cells indirectly participate in antitumor responses by inducing dendritic cell maturation and producing cytokines that promote tumor clearance by CD8+ T and NK cells. Although iNKT cells thereby act as potent cellular adjuvants, it is less clear whether they directly control the growth of tumors. To gain insights into the direct contribution of iNKT cells to tumor immune surveillance, we developed in vitro and in vivo systems to selectively examine the antitumor activity of iNKT cells in the absence of other cytolytic effectors. Using the EL4 T-lymphoma cell line as a model, we find that iNKT cells exert robust and specific lysis of tumor cells in vitro in a manner that is differentially-induced by iNKT cell agonists of varying TCR affinities, such as OCH, α-galactosyl ceramide and PBS44. In vitro blockade of CD1d-mediated lipid antigen presentation, disruption of T cell receptor (TCR) signaling, or loss of perforin expression significantly reduce iNKT cell killing. Consistent with these findings, iNKT cell reconstitution of T, B, and NK cell-deficient mice slows EL4 growth in vivo via TCR-CD1d and perforin-dependent mechanisms. Together, these observations establish that iNKT cells are sufficient to control the growth of T-lymphoma in vitro and in vivo. They also suggest that the induction of iNKT cell cytotoxic responses in situ might serve as a more effective strategy to prevent and/or treat CD1d+ cancers, such as T-lymphoma. PMID:24563871

iNKT cell cytotoxic responses control T-lymphoma growth in vitro and in vivo .

PubMed

Bassiri, Hamid; Das, Rupali; Guan, Peng; Barrett, David M; Brennan, Patrick J; Banerjee, Pinaki P; Wiener, Susan J; Orange, Jordan S; Brenner, Michael B; Grupp, Stephan A; Nichols, Kim E

2014-01-01

Invariant natural killer T (iNKT) cells comprise a lineage of CD1d-restricted glycolipid-reactive T lymphocytes with important roles in host immunity to cancer. iNKT cells indirectly participate in antitumor responses by inducing dendritic cell maturation and producing cytokines that promote tumor clearance by CD8+ T and NK cells. Although iNKT cells thereby act as potent cellular adjuvants, it is less clear whether they directly control the growth of tumors. To gain insights into the direct contribution of iNKT cells to tumor immune surveillance, we developed in vitro and in vivo systems to selectively examine the antitumor activity of iNKT cells in the absence of other cytolytic effectors. Using the EL4 T-lymphoma cell line as a model, we found that iNKT cells exert robust and specific lysis of tumor cells in vitro in a manner that is differentially induced by iNKT cell agonists of varying T-cell receptor (TCR) affinities, such as OCH, α-galactosyl ceramide, and PBS44. In vitro blockade of CD1d-mediated lipid antigen presentation, disruption of TCR signaling, or loss of perforin expression significantly reduce iNKT cell killing. Consistent with these findings, iNKT cell reconstitution of T, B, and NK cell–deficient mice slows EL4 growth in vivo via TCR-CD1d and perforin-dependent mechanisms. Together, these observations establish that iNKT cells are sufficient to control the growth of T lymphoma in vitro and in vivo. They also suggest that the induction of iNKT cell cytotoxic responses in situ might serve as a more effective strategy to prevent and/or treat CD1d+ cancers, such as T lymphoma. ©2013 AACR.

[Nasal NK/T cell lymphoma with outstanding performance of ocular symptoms].

PubMed

Liu, Lei; Zhao, Yulin; Wang, Jia; Ma, Fei

2012-09-01

To investigate the clinical features and misdiagnosis of nasal NK/T cell lymphoma with outstanding performance in ocular symptoms. Clinical data of 11 patients who had nasal NK/T cell lymphoma with the outstanding performances in ocular symptoms during 2009 to 2011 were retrospectively analyzed. The rate of misdiagnosis in the first diagnosis and first pathological diagnosis were 72.7% and 27.3% respectively. Nasal NK/T cell lymphoma with obvious ocular symptoms developed quickly and had almost special imaging findings. Nasal NK/T cell lymphoma with outstanding performance of ocular symptoms can be easily misdiagnosed. Comprehensive consideration of the clinical features, imaging findings and pathological examination do help to make accurate diagnosis early.

Regulatory iNKT cells lack PLZF expression and control Treg cell and macrophage homeostasis in adipose tissue

PubMed Central

Lynch, Lydia; Michelet, Xavier; Zhang, Sai; Brennan, Patrick J.; Moseman, Ashley; Lester, Chantel; Besra, Gurdyal; Vomhof-Dekrey, Emilie E.; Tighe, Mike; Koay, Hui-Fern; Godfrey, Dale I.; Leadbetter, Elizabeth A.; Sant’Angelo, Derek B.; von Andrian, Ulrich; Brenner, Michael B.

2015-01-01

iNKT cells are CD1d-restricted lipid-sensing innate T cells that express the transcription factor PLZF. iNKT cells accumulate in adipose tissue, where they are anti-inflammatory, but the factors that contribute to their anti-inflammatory nature, and their targets in adipose tissue are unknown. Here we report that adipose tissue iNKT cells have a unique transcriptional program and produce interleukin 2 (IL-2) and IL-10. Unlike other iNKT cells, they lack PLZF, but express the transcription factor E4BP4, which controls their IL-10 production. Adipose iNKT cells are a tissue resident population that induces an anti-inflammatory phenotype in macrophages and, through production of IL-2, controls the number, proliferation and suppressor function of adipose regulatory T (Treg) cells. Thus, adipose tissue iNKT cells are unique regulators of immune homeostasis in this tissue. PMID:25436972

Sodium chloride inhibits IFN-γ, but not IL-4, production by invariant NKT cells.

PubMed

Jeong, Dongjin; Kim, Hye Young; Chung, Doo Hyun

2018-01-01

Invariant NKT (iNKT) cells are a distinct subset of T cells that exert Janus-like functions in vivo by producing IFN-γ and IL-4. Sodium chloride modulates the functions of various immune cells, including conventional CD4 + T cells and macrophages. However, it is not known whether sodium chloride affects iNKT cell function, so we addressed this issue. Sodium chloride inhibited IFN-γ, but not IL-4, production by iNKT cells upon TCR or TCR-independent (IL-12 and IL-18) stimulation in a dose-dependent manner. Consistently, sodium chloride reduced the expression level of tbx21, but not gata-3, in iNKT cells stimulated with TCR engagement or IL-12 + IL-18. Sodium chloride increased phosphorylated p38 expression in iNKT cells and inhibitors of p38, NFAT5, SGK1, and TCF-1 restored IFN-γ production by iNKT cells stimulated with sodium chloride and TCR engagement. Furthermore, adoptive transfer of iNKT cells pretreated with sodium chloride restored antibody-induced joint inflammation to a lesser extent than for untreated iNKT cells in Jα18 knockout mice. These findings suggest that sodium chloride inhibits IFN-γ production by iNKT cells in TCR-dependent and TCR-independent manners, which is dependent on p38, NFAT5, SGK1, and TCF-1. These findings highlight the functional role of sodium chloride in iNKT cell-mediated inflammatory diseases. ©2017 Society for Leukocyte Biology.

Differential requirements for the Ets transcription factor Elf-1 in the development of NKT cells and NK cells

PubMed Central

Choi, Hak-Jong; Geng, Yanbiao; Cho, Hoonsik; Li, Sha; Giri, Pramod Kumar; Felio, Kyrie

2011-01-01

E26 Transformation specific (Ets) family transcription factors control the expression of a large number of genes regulating hematopoietic cell development and function. Two such transcription factors, Ets-1 and myeloid Elf-1–like factor (MEF), have been shown to play critical roles in both natural killer (NK)– and NKT-cell development, but not in the development of conventional T cells. In this study, we address the role of E74-like factor 1 (Elf-1), another Ets family transcription factor that is closely related to MEF but divergent from Ets-1, in NK- and NKT-cell development using Elf-1–deficient (Elf-1−/−) mice. Whereas the proportion of NK cells in Elf-1−/− mice was normal, the proportion of NKT cells was significantly reduced in the thymus and periphery of Elf-1−/− mice compared with wild-type (WT) mice. Although Ets-1–deficient mice lack NKT cells altogether, Elf-1−/− mice exhibited only a partial block in NKT-cell development caused by a cell-intrinsic defect in the selection, survival, and maturation of NKT cells. In addition, residual NKT cells found in Elf-1−/− mice produced less cytokine upon antigen stimulation compared with WT NKT cells. Our data demonstrate that Elf-1 plays an important and nonredundant role in the development and function of NKT cells, but is not involved in NK-cell development. PMID:21148815

Application of tissue-specific NK and NKT cell activity for tumor immunotherapy

PubMed Central

Subleski, Jeff J.; Wiltrout, Robert H.; Weiss, Jonathan M.

2009-01-01

Natural killer (NK) and NKT cells are a first line of defense against pathogens and transformed cells. However, dysregulation of their function can lead to autoimmune disease. A better understanding of the mechanisms controlling NK and NKT effector function should lead to the development of improved strategies for the treatment of many diseases. The site in which NK and NKT cells reside should be taken into account, because accumulating evidence suggests that the tissue microenvironment strongly influences their function. In this regard, the liver represents a unique immunologic organ in which the balance between the need for tolerance and the ability to respond rapidly to pathogens and tissue injury is tightly regulated. NK cells in the liver have augmented cytolytic activity as compared to other organs, which is consistent with a role for liver-associated NK cells in being critical effector cells for inhibiting tumor metastasis in the liver. Several studies also suggest that hepatic NKT cells have different functions than those in other organs. Whereas splenic and thymic NKT cells have been shown to suppress diabetes development, facilitate the induction of systemic tolerance and are regulated by IL-4 and other Th2 cytokines, certain subsets of NKT cells in the liver are important sources of Th1 cytokines such as Interferon gamma, and are the primary mediators of anti-tumor responses. The unique properties and roles as critical effector cells make NK and NKT cells within the liver microenvironment attractive targets of immunotherapeutic approaches that have the goal of controlling tumor metastasis in the liver. PMID:19682859

The adaptor molecule SAP plays essential roles during invariant NKT cell cytotoxicity and lytic synapse formation.

PubMed

Das, Rupali; Bassiri, Hamid; Guan, Peng; Wiener, Susan; Banerjee, Pinaki P; Zhong, Ming-Chao; Veillette, André; Orange, Jordan S; Nichols, Kim E

2013-04-25

The adaptor molecule signaling lymphocytic activation molecule-associated protein (SAP) plays critical roles during invariant natural killer T (iNKT) cell ontogeny. As a result, SAP-deficient humans and mice lack iNKT cells. The strict developmental requirement for SAP has made it difficult to discern its possible involvement in mature iNKT cell functions. By using temporal Cre recombinase-mediated gene deletion to ablate SAP expression after completion of iNKT cell development, we demonstrate that SAP is essential for T-cell receptor (TCR)-induced iNKT cell cytotoxicity against T-cell and B-cell leukemia targets in vitro and iNKT-cell-mediated control of T-cell leukemia growth in vivo. These findings are not restricted to the murine system: silencing RNA-mediated suppression of SAP expression in human iNKT cells also significantly impairs TCR-induced cytolysis. Mechanistic studies reveal that iNKT cell killing requires the tyrosine kinase Fyn, a known SAP-binding protein. Furthermore, SAP expression is required within iNKT cells to facilitate their interaction with T-cell targets and induce reorientation of the microtubule-organizing center to the immunologic synapse (IS). Collectively, these studies highlight a novel and essential role for SAP during iNKT cell cytotoxicity and formation of a functional IS.

The adaptor molecule SAP plays essential roles during invariant NKT cell cytotoxicity and lytic synapse formation

PubMed Central

Das, Rupali; Bassiri, Hamid; Guan, Peng; Wiener, Susan; Banerjee, Pinaki P.; Zhong, Ming-Chao; Veillette, André; Orange, Jordan S.

2013-01-01

The adaptor molecule signaling lymphocytic activation molecule–associated protein (SAP) plays critical roles during invariant natural killer T (iNKT) cell ontogeny. As a result, SAP-deficient humans and mice lack iNKT cells. The strict developmental requirement for SAP has made it difficult to discern its possible involvement in mature iNKT cell functions. By using temporal Cre recombinase–mediated gene deletion to ablate SAP expression after completion of iNKT cell development, we demonstrate that SAP is essential for T-cell receptor (TCR)–induced iNKT cell cytotoxicity against T-cell and B-cell leukemia targets in vitro and iNKT-cell–mediated control of T-cell leukemia growth in vivo. These findings are not restricted to the murine system: silencing RNA–mediated suppression of SAP expression in human iNKT cells also significantly impairs TCR-induced cytolysis. Mechanistic studies reveal that iNKT cell killing requires the tyrosine kinase Fyn, a known SAP-binding protein. Furthermore, SAP expression is required within iNKT cells to facilitate their interaction with T-cell targets and induce reorientation of the microtubule-organizing center to the immunologic synapse (IS). Collectively, these studies highlight a novel and essential role for SAP during iNKT cell cytotoxicity and formation of a functional IS. PMID:23430111

Characterization of human invariant natural killer T subsets in health and disease using a novel invariant natural killer T cell-clonotypic monoclonal antibody, 6B11.

PubMed

Montoya, Carlos J; Pollard, David; Martinson, Jeffrey; Kumari, Kumud; Wasserfall, Clive; Mulder, Candice B; Rugeles, Maria T; Atkinson, Mark A; Landay, Alan L; Wilson, S Brian

2007-09-01

Identification of human CD1d-restricted T-cell receptor (TCR)-invariant natural killer T (iNKT) cells has been dependent on utilizing combinations of monoclonal antibodies or CD1d tetramers, which do not allow for the most specific analysis of this T-cell subpopulation. A novel monoclonal antibody (clone 6B11), specific for the invariant CDR3 loop of human canonical Valpha24Jalpha18 TCR alpha chain, was developed and used to specifically characterize iNKT cells. In healthy individuals studied for up to 1 year, a wide but stable frequency of circulating iNKT cells (range: 0.01-0.92%) was observed, with no differences in frequency by gender. Four stable iNKT cell subsets were characterized in peripheral blood based on the expression of CD4 and CD8, with CD8(+) iNKT cells being a phenotypic and functionally different subset from CD4(+) and double negative iNKT cells; in particular, LAG-3 was preferentially expressed on CD8(+) iNKT cells. In addition, a strong negative linear correlation between the frequency of total iNKT cells and percentage of the CD4(+) subset was observed. In terms of their potential association with disease, patients at risk for type 1 diabetes had significantly expanded frequencies of double negative iNKT cells when compared to matched controls and first-degree relatives. Moreover, peripheral blood CD4(+) iNKT cells were the highest producers of interleukin-4, while the production of interferon-gamma and tumour necrosis factor-alpha was similar amongst all iNKT cell subsets. These differences in iNKT cell subsets suggest that in humans the relative ratio of iNKT cell subsets may influence susceptibility vs. resistance to immune-mediated diseases.

The transcription factor Th-POK negatively regulates Th17 differentiation in Vα14i NKT cells

PubMed Central

Engel, Isaac; Zhao, Meng; Kappes, Dietmar; Taniuchi, Ichiro

2012-01-01

The majority of mouse Vα14 invariant natural killer T (Vα14i NKT) cells produce several cytokines, including IFNγ and IL-4, very rapidly after activation. A subset of these cells, known as NKT17 cells, however, differentiates in the thymus to preferentially produce IL-17. Here, we show that the transcription factor—known as T helper, Poxviruses, and Zinc-finger and Krüppel family, (Th-POK)—represses the formation of NKT17 cells. Vα14i NKT cells from Th-POK–mutant helper deficient (hd/hd) mice have increased transcripts of genes normally expressed by Th17 and NKT17 cells, and even heterozygosity for this mutation leads to dramatically increased numbers of Vα14i NKT cells that are poised to express IL-17, especially in the thymus and lymph nodes. In addition, using gene reporter mice, we demonstrate that NKT17 cells from wild-type mice express lower amounts of Th-POK than the majority population of Vα14i NKT cells. We also show that retroviral transduction of Th-POK represses the expression of the Th17 master regulator RORγT in Vα14i NKT-cell lines. Our data suggest that NKT17-cell differentiation is intrinsically regulated by Th-POK activity, with only low levels of Th-POK permissive for the differentiation of NKT17 cells. PMID:23034280

Activation of plasmacytoid dendritic cells with TLR9 agonists initiates invariant NKT cell-mediated cross-talk with myeloid dendritic cells.

PubMed

Montoya, Carlos J; Jie, Hyun-Bae; Al-Harthi, Lena; Mulder, Candice; Patiño, Pablo J; Rugeles, María T; Krieg, Arthur M; Landay, Alan L; Wilson, S Brian

2006-07-15

CD1d-restricted invariant NK T (iNKT) cells and dendritic cells (DCs) have been shown to play crucial roles in various types of immune responses, including TLR9-dependent antiviral responses initiated by plasmacytoid DCs (pDCs). However, the mechanism by which this occurs is enigmatic because TLRs are absent in iNKT cells and human pDCs do not express CD1d. To explore this process, pDCs were activated with CpG oligodeoxyribonucleotides, which stimulated the secretion of several cytokines such as type I and TNF-alpha. These cytokines and other soluble factors potently induced the expression of activation markers on iNKT cells, selectively enhanced double-negative iNKT cell survival, but did not induce their expansion or production of cytokines. Notably, pDC-derived factors licensed iNKT cells to respond to myeloid DCs: an important downstream cellular target of iNKT cell effector function and a critical contributor to the initiation of adaptive immune responses. This interaction supports the notion that iNKT cells can mediate cross-talk between DC subsets known to express mutually exclusive TLR and cytokine profiles.

TET proteins regulate the lineage specification and TCR-mediated expansion of iNKT cells.

PubMed

Tsagaratou, Ageliki; González-Avalos, Edahí; Rautio, Sini; Scott-Browne, James P; Togher, Susan; Pastor, William A; Rothenberg, Ellen V; Chavez, Lukas; Lähdesmäki, Harri; Rao, Anjana

2017-01-01

TET proteins oxidize 5-methylcytosine in DNA to 5-hydroxymethylcytosine and other oxidation products. We found that simultaneous deletion of Tet2 and Tet3 in mouse CD4 + CD8 + double-positive thymocytes resulted in dysregulated development and proliferation of invariant natural killer T cells (iNKT cells). Tet2-Tet3 double-knockout (DKO) iNKT cells displayed pronounced skewing toward the NKT17 lineage, with increased DNA methylation and impaired expression of genes encoding the key lineage-specifying factors T-bet and ThPOK. Transfer of purified Tet2-Tet3 DKO iNKT cells into immunocompetent recipient mice resulted in an uncontrolled expansion that was dependent on the nonclassical major histocompatibility complex (MHC) protein CD1d, which presents lipid antigens to iNKT cells. Our data indicate that TET proteins regulate iNKT cell fate by ensuring their proper development and maturation and by suppressing aberrant proliferation mediated by the T cell antigen receptor (TCR).

The Interaction between Regulatory T Cells and NKT Cells in the Liver: A CD1d Bridge Links Innate and Adaptive Immunity

PubMed Central

Webb, Tonya J.; Potter, James P.; Li, Zhiping

2011-01-01

Background/Aims Regulatory T cells (Tregs) and natural killer T (NKT) cells are two distinct lymphocyte subsets that independently regulate hepatic adaptive and innate immunity, respectively. In the current study, we examine the interaction between Tregs and NKT cells to understand the mechanisms of cross immune regulation by these cells. Methods The frequency and function of Tregs were evaluated in wild type and NKT cell deficient (CD1dko) mice. In vitro lymphocyte proliferation and apoptosis assays were performed with NKT cells co-cultured with Tregs. The ability of Tregs to inhibit NKT cells in vivo was examined by adoptive transfer of Tregs in a model of NKT cell mediated hepatitis. Results CD1dko mice have a significant reduction in hepatic Tregs. Although, the Tregs from CD1dko mice remain functional and can suppress conventional T cells, their ability to suppress activation induced NKT cell proliferation and to promote NKT cell apoptosis is greatly diminished. These effects are CD1d dependent and require cell to cell contact. Adoptive transfer of Tregs inhibits NKT cell-mediated liver injury. Conclusions NKT cells promote Tregs, and Tregs inhibit NKT cells in a CD1d dependent manner requiring cell to cell contact. These cross-talk immune regulations provide a linkage between innate and adaptive immunity. PMID:22073248

Cyclooxygenase-2 Inhibition Enhances Proliferation of NKT Cells Derived from Patients with Laryngeal Cancer.

PubMed

Klatka, Janusz; Grywalska, Ewelina; Hymos, Anna; Guz, Małgorzata; Polberg, Krzysztof; Roliński, Jacek; Stepulak, Andrzej

2017-08-01

The aim of this study was to analyze whether inhibition of cyclooxygenase-2 by celecoxib and the subsequent enhancement in the proliferation of natural killer T (NKT) cells could play a role in dendritic cell (DC)-based laryngeal cancer (LC) immunotherapy. Peripheral blood mononuclear cells were obtained from 48 male patients diagnosed with LC and 30 control patients without cancer disease. Neoplastic cell lysate preparations were made from cancer tissues obtained after surgery and used for in vitro DCs generation. NKT cells proliferation assay was performed based on 3 H-thymidine incorporation assay. An increased proliferation of NKT cells was obtained from control patients compared to NKT cells obtained from LC patients regardless of the type of stimulation or treatment. In the patient group diagnosed with LC, COX-2 inhibition resulted in a significantly enhanced proliferation of NKT cells when stimulated with autologous DCs than NKT cells stimulated with DCs without COX-2 inhibition. These correlations were not present in the control group. Higher proliferation rate of NKT cells was also observed in non-metastatic and highly differentiated LC, which was independent of the type of stimulation or treatment. COX-2 inhibition could be regarded as immunotherapy-enhancing tool in patients with LC. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells

PubMed Central

Torreno-Pina, Juan A.; Manzo, Carlo; Salio, Mariolina; Aichinger, Michael C.; Oddone, Anna; Lakadamyali, Melike; Shepherd, Dawn; Besra, Gurdyal S.; Cerundolo, Vincenzo

2016-01-01

Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such “tonic” activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters. PMID:26798067

Critical role for the chemokine receptor CXCR6 in homeostasis and activation of CD1d-restricted NKT cells.

PubMed

Germanov, Elitza; Veinotte, Linnea; Cullen, Robyn; Chamberlain, Erin; Butcher, Eugene C; Johnston, Brent

2008-07-01

NK T (NKT) cells play important roles in the regulation of diverse immune responses. However, little is known about the mechanisms that regulate homeostasis and activation of these cells. Thymic NKT cells up-regulated the chemokine receptor CXCR6 following positive selection and migrated toward CXCL16 in vitro. However, CXCR6 was not essential for thymic development or maturation. In contrast, liver and lung NKT cells were depleted in CXCR6+/- and CXCR6-/- mice. The reduction in liver and lung NKT cells coincided with an increase in bone marrow NKT cells, suggesting a redistribution of NKT cells in CXCR6-/- animals. In wild-type mice, CXCL16 neutralization reduced accumulation of mature NK1.1+, but not immature NK1.1- NKT cell recent thymic emigrants in the liver. Given that thymic NKT cells are preferentially exported as NK1.1- cells, this suggests an additional role for CXCR6/CXCL16 in maturation or survival of immature liver NKT cells. CXCL16 blockade did not deplete resident NK1.1+ NKT cells, indicating that CXCR6/CXCL16 are not required to retain mature NKT cells in the liver. Cytokine production by liver and spleen NKT cells was impaired in CXCR6-/- mice following in vivo stimulation with alpha-galactosylceramide, implicating a novel role for CXCR6 in NKT cell activation. Reduced IFN-gamma production was not due to an intrinsic defect as production was normal following PMA and ionomycin stimulation. Preformed transcripts for IL-4, but not IFN-gamma, were reduced in CXCR6-/- liver NKT cells. These data identify critical roles for CXCR6/CXCL16 in NKT cell activation and the regulation of NKT cell homeostasis.

The Functions of Type I and Type II Natural Killer T (NKT) Cells in Inflammatory Bowel Diseases

PubMed Central

Liao, Chia-Min; Zimmer, Michael I.; Wang, Chyung-Ru

2013-01-01

CD1d-restricted natural killer T (NKT) cells are a distinct subset of T cells that rapidly produce an array of cytokines upon activation and play a critical role in regulating various immune responses. NKT cells are classified into two groups based on differences in T cell receptor (TCR) usage. Type I NKT cells have an invariant TCRα-chain and are readily detectable by α-galactosylceramide (α-GalCer)-loaded CD1d tetramers. Type II NKT cells have a more diverse TCR repertoire and cannot be directly identified. Both types of NKT cells as well as multiple CD1d-expressing cell types are present in the intestine and their interactions are likely to be modulated by pathogenic and commensal microbes, which in turn contribute to the intestinal immune responses in health and disease. Indeed, in several animal models of inflammatory bowel disease (IBD), Type I NKT cells have been shown to make both protective and pathogenic contributions to disease. In contrast, in human patients suffering from ulcerative colitis (UC), and a mouse model in which both CD1d expression and the frequency of Type II NKT cells are increased, Type II NKT cells appear to promote intestinal inflammation. In this review, we summarize present knowledge on the antigen recognition, activation and function of NKT cells with a particular focus on their role in IBD, and discuss factors that may influence the functional outcome of NKT cell responses in intestinal inflammation. PMID:23518808

The clinical implication and molecular mechanism of preferential IL-4 production by modified glycolipid-stimulated NKT cells

PubMed Central

Oki, Shinji; Chiba, Asako; Yamamura, Takashi; Miyake, Sachiko

2004-01-01

OCH, a sphingosine-truncated analog of α-galactosylceramide (αGC), is a potential therapeutic reagent for a variety of Th1-mediated autoimmune diseases through its selective induction of Th2 cytokines from natural killer T (NKT) cells. We demonstrate here that the NKT cell production of IFN-γ is more susceptible to the sphingosine length of glycolipid ligand than that of IL-4 and that the length of the sphingosine chain determines the duration of NKT cell stimulation by CD1d-associated glycolipids. Furthermore, IFN-γ production by NKT cells requires longer T cell receptor stimulation than is required for IL-4 production by NKT cells stimulated either with immobilized mAb to CD3 or with immobilized “αGC-loaded” CD1d molecules. Interestingly, transcription of IFN-γ but not that of IL-4 was sensitive to cycloheximide treatment, indicating the intrinsic involvement of de novo protein synthesis for IFN-γ production by NKT cells. Finally, we determined c-Rel was preferentially transcribed in αGC-stimulated but not in OCH-stimulated NKT cells and was essential for IFN-γ production by activated NKT cells. Given the dominant immune regulation by the remarkable cytokine production of ligand-stimulated NKT cells in vivo, in comparison with that of (antigen-specific) T cells or NK cells, the current study confirms OCH as a likely therapeutic reagent for use against Th1-mediated autoimmune diseases and provides a novel clue for the design of drugs targeting NKT cells. PMID:15173890

Functional Invariant NKT Cells in Pig Lungs Regulate the Airway Hyperreactivity: A Potential Animal Model

PubMed Central

Manickam, Cordelia; Khatri, Mahesh; Rauf, Abdul; Li, Xiangming; Tsuji, Moriya; Rajashekara, Gireesh; Dwivedi, Varun

2015-01-01

Important roles played by invariant natural killer T (iNKT) cells in asthma pathogenesis have been demonstrated. We identified functional iNKT cells and CD1d molecules in pig lungs. Pig iNKT cells cultured in the presence of α-GalCer proliferated and secreted Th1 and Th2 cytokines. Like in other animal models, direct activation of pig lung iNKT cells using α-GalCer resulted in acute airway hyperreactivity (AHR). Clinically, acute AHR-induced pigs had increased respiratory rate, enhanced mucus secretion in the airways, fever, etc. In addition, we observed petechial hemorrhages, infiltration of CD4+ cells, and increased Th2 cytokines in AHR-induced pig lungs. Ex vivo proliferated iNKT cells of asthma induced pigs in the presence of C-glycoside analogs of α-GalCer had predominant Th2 phenotype and secreted more of Th2 cytokine, IL-4. Thus, baby pigs may serve as a useful animal model to study iNKT cell-mediated AHR caused by various environmental and microbial CD1d-specific glycolipid antigens. PMID:21042929

Type II Natural Killer T (NKT) Cells And Their Emerging Role In Health And Disease

PubMed Central

Dhodapkar, Madhav V.; Kumar, Vipin

2016-01-01

Natural killer T (NKT) cells recognize lipid antigens presented by a class I MHC-like molecule CD1d, a member of the CD1 family. While most of the initial studies on NKT cells focused on a subset with semi-invariant T cell receptor (TCR) termed iNKT cells, majority of CD1d-restricted lipid-reactive human T cells express diverse TCRs and are termed as type II NKT cells. These cells constitute a distinct population of circulating and tissue-resident effector T cells with immune-regulatory properties. They react to a growing list of self- as well as non-self lipid ligands, and share some properties with both iNKT as well as conventional T cells. Emerging body of evidence points to their role in the regulation of immunity to pathogens/tumors and in autoimmune/metabolic disorders. Improved understanding of the biology of these cells and the ability to manipulate their function may be of therapeutic benefit in diverse disease conditions. PMID:28115591

Components of Streptococcus pneumoniae suppress allergic airways disease and NKT cells by inducing regulatory T cells.

PubMed

Thorburn, Alison N; Foster, Paul S; Gibson, Peter G; Hansbro, Philip M

2012-05-01

Asthma is an allergic airways disease (AAD) caused by dysregulated immune responses and characterized by eosinophilic inflammation, mucus hypersecretion, and airway hyperresponsiveness (AHR). NKT cells have been shown to contribute to AHR in some mouse models. Conversely, regulatory T cells (Tregs) control aberrant immune responses and maintain homeostasis. Recent evidence suggests that Streptococcus pneumoniae induces Tregs that have potential to be harnessed therapeutically for asthma. In this study, mouse models of AAD were used to identify the S. pneumoniae components that have suppressive properties, and the mechanisms underlying suppression were investigated. We tested the suppressive capacity of type-3-polysaccharide (T3P), isolated cell walls, pneumolysoid (Ply) and CpG. When coadministered, T3P + Ply suppressed the development of: eosinophilic inflammation, Th2 cytokine release, mucus hypersecretion, and AHR. Importantly, T3P + Ply also attenuated features of AAD when administered during established disease. We show that NKT cells contributed to the development of AAD and also were suppressed by T3P + Ply treatment. Furthermore, adoptive transfer of NKT cells induced AHR, which also could be reversed by T3P + Ply. T3P + Ply-induced Tregs were essential for the suppression of NKT cells and AAD, which was demonstrated by Treg depletion. Collectively, our results show that the S. pneumoniae components T3P + Ply suppress AAD through the induction of Tregs that blocked the activity of NKT cells. These data suggest that S. pneumoniae components may have potential as a therapeutic strategy for the suppression of allergic asthma through the induction of Tregs and suppression of NKT cells.

Dual immune effect of iNKT cells considering human cutaneous and visceral leishmaniasis: an example of cell plasticity according to different disease scenarios.

PubMed

de Gois Macêdo, Bruna; Peixoto, Rephany Fonseca; Maciel, Bruna Leal Lima; de Assis Silva Gomes, Juliana; de Lourdes Assunção Araújo de Azevedo, Fátima; Veras, Robson Cavalcanti; de Medeiros, Isac Almeida; de Lima Grisi, Teresa Cristina Soares; de Araújo, Demétrius Antônio Machado; Amaral, Ian Porto Gurgel do; de Souza Lima, Tatjana Keesen

2018-04-27

Although the semi-invariant natural killer T cells (iNKT) are a small subpopulation of cells in the peripheral blood, they are presumed to play a role in early stages of infection against various pathogens, including protozoa. This work investigates the activation status and cytokine profile of iNKT cells during human Leishmania infantum and Leishmania braziliensis infection. We studied iNKT cells in symptomatic active visceral patients (AVL) (n=8), symptomatic active cutaneous patients (ACL) (n=13), negative endemic controls (NEC) (n=6) and non-endemic controls (NonEC) (n=6), with and without Total Leishmania antigen stimulus (TLA). The number of iNKT cells in the peripheral blood of ACL and AVL patients unaltered in relation to control groups. Moreover, the iNKT cells from ACL showed a hyperactivation profile compared to AVL patients. Additionally, TLA induced IFN-gamma production in iNKT cells from ACL patients, while in iNKT of AVL patients, TLA induced a decrease of this cytokine. Higher IL-17 and IL-10 production by iNKT cells from ACL patients were also observed compared to all other groups. There were no changes in IL-10 iNKT producing cells in AVL after TLA stimulation. However, TLA induced increase of IL-10 in iNKT cells in ACL patients. These findings suggest that although iNKT cells showed distinct profiles in ACL and AVL patients, they play a dual role in immune modulation in both Leishmania infections. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Dendritic cells and anergic type I NKT cells play a crucial role in sulfatide-mediated immune regulation in experimental autoimmune encephalomyelitis

PubMed Central

Maricic, Igor; Halder, Ramesh; Bischof, Felix; Kumar, Vipin

2014-01-01

CD1d-restricted NKT cells can be divided into two groups: type I NKT cells utilize a semi-invariant TCR whereas type II express a relatively diverse set of TCRs. A major subset of type II NKT cells recognizes myelin-derived sulfatides and is selectively enriched in the central nervous system tissue during experimental autoimmune encephalomyelitis (EAE). We have shown that activation of sulfatide-reactive type II NKT cells by sulfatide prevents induction of EAE. Here we have addressed the mechanism of regulation as well as whether a single immunodominant form of synthetic sulfatide can treat ongoing chronic and relapsing EAE in SJL/J mice. We have shown that the activation of sulfatide-reactive type II NKT cells leads to a significant reduction in the frequency and effector function of PLP139-151/I-As–tetramer+ cells in lymphoid and CNS tissues. In addition, type I NKT cells and dendritic cells in the periphery as well as CNS-resident microglia are inactivated following sulfatide administration, and mice deficient in type I NKT cells are not protected from disease. Moreover tolerized DCs from sulfatide-treated animals can adoptively transfer protection into naive mice. Treatment of SJL/J mice with a synthetic cis-tetracosenoyl sulfatide, but not αGalCer, reverses ongoing chronic and relapsing EAE. Our data highlight a novel immune regulatory pathway involving NKT subset interactions leading to inactivation of type I NKT cells, DCs, and microglial cells in suppression of autoimmunity. Since CD1 molecules are non-polymorphic, the sulfatide-mediated immune regulatory pathway can be targeted for development of non-HLA-dependent therapeutic approaches to T cell-mediated autoimmune diseases. PMID:24973441

Cross-activating invariant NKT cells and kupffer cells suppress cholestatic liver injury in a mouse model of biliary obstruction.

PubMed

Duwaerts, Caroline C; Sun, Eric P; Cheng, Chao-Wen; van Rooijen, Nico; Gregory, Stephen H

2013-01-01

Both Kupffer cells and invariant natural killer T (iNKT) cells suppress neutrophil-dependent liver injury in a mouse model of biliary obstruction. We hypothesize that these roles are interdependent and require iNKT cell-Kupffer cell cross-activation. Female, wild-type and iNKT cell-deficient C57Bl/6 mice were injected with magnetic beads 3 days prior to bile duct ligation (BDL) in order to facilitate subsequent Kupffer cell isolation. On day three post-BDL, the animals were euthanized and the livers dissected. Necrosis was scored; Kupffer cells were isolated and cell surface marker expression (flow cytometry), mRNA expression (qtPCR), nitric oxide (NO (.) ) production (Griess reaction), and protein secretion (cytometric bead-array or ELISAs) were determined. To address the potential role of NO (.) in suppressing neutrophil accumulation, a group of WT mice received 1400W, a specific inducible nitric oxide synthase (iNOS) inhibitor, prior to BDL. To clarify the mechanisms underlying Kupffer cell-iNKT cell cross-activation, WT animals were administered anti-IFN-γ or anti-lymphocyte function-associated antigen (LFA)-1 antibody prior to BDL. Compared to their WT counterparts, Kupffer cells obtained from BDL iNKT cell-deficient mice expressed lower iNOS mRNA levels, produced less NO (.) , and secreted more neutrophil chemoattractants. Both iNOS inhibition and IFN-γ neutralization increased neutrophil accumulation in the livers of BDL WT mice. Anti-LFA-1 pre-treatment reduced iNKT cell accumulation in these same animals. These data indicate that the LFA-1-dependent cross-activation of iNKT cells and Kupffer cells inhibits neutrophil accumulation and cholestatic liver injury.

Cross-Activating Invariant NKT Cells and Kupffer Cells Suppress Cholestatic Liver Injury in a Mouse Model of Biliary Obstruction

PubMed Central

Duwaerts, Caroline C.; Sun, Eric P.; Cheng, Chao-Wen; van Rooijen, Nico; Gregory, Stephen H.

2013-01-01

Both Kupffer cells and invariant natural killer T (iNKT) cells suppress neutrophil-dependent liver injury in a mouse model of biliary obstruction. We hypothesize that these roles are interdependent and require iNKT cell-Kupffer cell cross-activation. Female, wild-type and iNKT cell-deficient C57Bl/6 mice were injected with magnetic beads 3 days prior to bile duct ligation (BDL) in order to facilitate subsequent Kupffer cell isolation. On day three post-BDL, the animals were euthanized and the livers dissected. Necrosis was scored; Kupffer cells were isolated and cell surface marker expression (flow cytometry), mRNA expression (qtPCR), nitric oxide (NO.) production (Griess reaction), and protein secretion (cytometric bead-array or ELISAs) were determined. To address the potential role of NO. in suppressing neutrophil accumulation, a group of WT mice received 1400W, a specific inducible nitric oxide synthase (iNOS) inhibitor, prior to BDL. To clarify the mechanisms underlying Kupffer cell-iNKT cell cross-activation, WT animals were administered anti-IFN-γ or anti-lymphocyte function-associated antigen (LFA)-1 antibody prior to BDL. Compared to their WT counterparts, Kupffer cells obtained from BDL iNKT cell-deficient mice expressed lower iNOS mRNA levels, produced less NO., and secreted more neutrophil chemoattractants. Both iNOS inhibition and IFN-γ neutralization increased neutrophil accumulation in the livers of BDL WT mice. Anti-LFA-1 pre-treatment reduced iNKT cell accumulation in these same animals. These data indicate that the LFA-1-dependent cross-activation of iNKT cells and Kupffer cells inhibits neutrophil accumulation and cholestatic liver injury. PMID:24260285

The Lysine Acetyltransferase GCN5 Is Required for iNKT Cell Development through EGR2 Acetylation.

PubMed

Wang, Yajun; Yun, Chawon; Gao, Beixue; Xu, Yuanming; Zhang, Yana; Wang, Yiming; Kong, Qingfei; Zhao, Fang; Wang, Chyung-Ru; Dent, Sharon Y R; Wang, Jian; Xu, Xiangping; Li, Hua-Bin; Fang, Deyu

2017-07-18

The development of CD1d-restricted invariant natural killer T (iNKT) cells, a population that is critical for both innate and adaptive immunity, is regulated by multiple transcription factors, but the molecular mechanisms underlying how the transcriptional activation of these factors are regulated during iNKT development remain largely unknown. We found that the histone acetyltransferase general control non-derepressible 5 (GCN5) is essential for iNKT cell development during the maturation stage. GCN5 deficiency blocked iNKT cell development in a cell-intrinsic manner. At the molecular level, GCN5 is a specific lysine acetyltransferase of early growth responsive gene 2 (EGR2), a transcription factor required for iNKT cell development. GCN5-mediated acetylation positively regulated EGR2 transcriptional activity, and both genetic and pharmacological GCN5 suppression specifically inhibited the transcription of EGR2 target genes in iNKT cells, including Runx1, promyelocytic leukemia zinc finger protein (PLZF), interleukin (IL)-2Rb, and T-bet. Therefore, our study revealed GCN5-mediated EGR2 acetylation as a molecular mechanism that regulates iNKT development. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

On/off TLR signaling decides proinflammatory or tolerogenic dendritic cell maturation upon CD1d-mediated interaction with invariant NKT cells.

PubMed

Caielli, Simone; Conforti-Andreoni, Cristina; Di Pietro, Caterina; Usuelli, Vera; Badami, Ester; Malosio, Maria Luisa; Falcone, Marika

2010-12-15

Invariant NKT (iNKT) cells play an effector/adjuvant function during antimicrobial and antitumoral immunity and a regulatory role to induce immune tolerance and prevent autoimmunity. iNKT cells that differentially modulate adaptive immunity do not bear a unique phenotype and/or specific cytokine secretion profile, thus opening questions on how a single T cell subset can exert opposite immunological tasks. In this study, we show that iNKT cells perform their dual roles through a single mechanism of action relying on the cognate interaction with myeloid dendritic cells (DCs) and leading to opposite effects depending on the presence of other maturation stimuli simultaneously acting on DCs. The contact of murine purified iNKT cells with immature autologous DCs directly triggers the tolerogenic maturation of DCs, rendering them able to induce regulatory T cell differentiation and prevent autoimmune diabetes in vivo. Conversely, the interaction of the same purified iNKT cells with DCs, in the presence of simultaneous TLR4 stimulation, significantly enhances proinflammatory DC maturation and IL-12 secretion. The different iNKT cell effects are mediated through distinct mechanisms and activation of different molecular pathways within the DC: CD1d signaling and activation of the ERK1/2 pathway for the tolerogenic action, and CD40-CD40L interaction and NF-κB activation for the adjuvant effect. Our data suggest that the DC decision to undergo proinflammatory or tolerogenic maturation results from the integration of different signals received at the time of iNKT cell contact and could have important therapeutic implications for exploiting iNKT cell adjuvant/regulatory properties in autoimmune diseases, infections, and cancer.

The adaptor protein SAP regulates type II NKT-cell development, cytokine production, and cytotoxicity against lymphoma.

PubMed

Weng, Xiufang; Liao, Chia-Min; Bagchi, Sreya; Cardell, Susanna L; Stein, Paul L; Wang, Chyung-Ru

2014-12-01

CD1d-restricted NKT cells represent a unique lineage of immunoregulatory T cells that are divided into two groups, type I and type II, based on their TCR usage. Because there are no specific tools to identify type II NKT cells, little is known about their developmental requirements and functional regulation. In our previous study, we showed that signaling lymphocytic activation molecule associated protein (SAP) is essential for the development of type II NKT cells. Here, using a type II NKT-cell TCR transgenic mouse model, we demonstrated that CD1d-expressing hematopoietic cells, but not thymic epithelial cells, meditate efficient selection of type II NKT cells. Furthermore, we showed that SAP regulates type II NKT-cell development by controlling early growth response 2 protein and promyelocytic leukemia zinc finger expression. SAP-deficient 24αβ transgenic T cells (24αβ T cells) exhibited an immature phenotype with reduced Th2 cytokine-producing capacity and diminished cytotoxicity to CD1d-expressing lymphoma cells. The impaired IL-4 production by SAP-deficient 24αβ T cells was associated with reduced IFN regulatory factor 4 and GATA-3 induction following TCR stimulation. Collectively, these data suggest that SAP is critical for regulating type II NKT cell responses. Aberrant responses of these T cells may contribute to the immune dysregulation observed in X-linked lymphoproliferative disease caused by mutations in SAP. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Does an NKT-cell-based immunotherapeutic approach have a future in multiple myeloma?

PubMed Central

Favreau, Mérédis; Vanderkerken, Karin

2016-01-01

Natural killer T (NKT) cells constitute a unique subset of innate-like T lymphocytes which differ from conventional T cells by recognizing lipid antigens presented by the non-polymorphic major histocompatibility complex (MHC) I-like molecule CD1d. Despite being a relatively infrequent population of lymphocytes, NKT cells can respond rapidly upon activation with glycosphingolipids by production of cytokines which aim to polarize different axes of the immune system. Due to their dual effector capacities, NKT cells can play a vital role in cancer immunity, infection, inflammation and autoimmune diseases. It is believed that modulation of their activity towards immune activation can be a useful tool in anti-tumor immunotherapeutic strategies. Here we summarize the characteristics of NKT cells and discuss their involvement in immunosurveillance. Furthermore, an update is given about their role and the progress that has been made in the field of multiple myeloma (MM). Finally, some challenges are discussed that are currently hampering further progress. PMID:26895468

Distribution of intrahepatic T, NK and CD3(+)CD56(+)NKT cells alters after liver transplantation: Shift from innate to adaptive immunity?

PubMed

Werner, Jens M; Lang, Corinna; Scherer, Marcus N; Farkas, Stefan A; Geissler, Edward K; Schlitt, Hans J; Hornung, Matthias

2011-07-01

The liver is an immunological organ containing a large number of T, NK and NKT cells, but little is known about intrahepatic immunity after LTx. Here, we investigated whether the distribution of T, NK and CD3(+)CD56(+)NKT cells is altered in transplanted livers under different circumstances. Core biopsies of transplanted livers were stained with antibodies against CD3 and CD56. Several cell populations including T (CD3(+)CD56(-)), NK (CD3(-)CD56(+)) and NKT cells (CD3(+)CD56(+)) were studied by fluorescence microscopy. Cell numbers were analyzed in relation to the time interval after LTx, immunosuppressive therapy and stage of acute graft rejection (measured with the rejection activity index: RAI) compared to tumor free liver tissue from patients after liver resection due to metastatic disease as control. Recruitment of CD3(+)CD56(+)NKT cells revealed a significant decrease during high RAI scores in comparison to low and middle RAI scores (RAI 7-9: 0.03±0.01/HPF vs. RAI 4-6: 0.1±0.005/HPF). CD3(+)CD56(+)NKT cells were also lower during immunosuppressive therapy with tacrolimus (0.03±0.01/HPF) than with cyclosporine (0.1±0.003/HPF), cyclosporine/MMF (0.1±0.003/HPF) or sirolimus (0.1±0.01/HPF) treatment. Intrahepatic T cell numbers increased significantly 50days after LTx compared to control liver tissue (4.5±0.2/HPF vs. 1.9±0.1/HPF). In contrast, NK cells (0.3±0.004/HPF) were significantly fewer in all biopsies after LTx compared to the control (0.7±0.04/HPF). These data indicate significant alterations in the hepatic recruitment of T, NK and CD3(+)CD56(+)NKT cells after LTx. The increase in T cells and the decrease in NK and CD3(+)CD56(+)NKT cells suggest a shift from innate to adaptive hepatic immunity in the liver graft. Copyright © 2011 Elsevier B.V. All rights reserved.

The Adaptor Protein SAP Regulates Type II NKT Cell Development, Cytokine Production and Cytotoxicity Against Lymphoma1

PubMed Central

Weng, Xiufang; Liao, Chia-Min; Bagchi, Sreya; Cardell, Susanna L.; Stein, Paul L.; Wang, Chyung-Ru

2014-01-01

CD1d-restricted NKT cells represent a unique lineage of immunoregulatory T cells that are divided into two groups, type I and type II, based on their TCR usage. Because there are no specific tools to identify type II NKT cells, little is known about their developmental requirements and functional regulation. In our previous study, we showed that signaling lymphocytic activation molecule-associated protein (SAP) is essential for the development of type II NKT cells. Here, using a type II NKT cell TCR transgenic mouse model (24αβTg), we demonstrated that CD1d-expressing hematopoietic cells but not thymic epithelial cells meditate efficient selection of type II NKT cells. Further, we showed that SAP regulates type II NKT cell development by controlling Egr2 and PLZF expression. SAP-deficient 24αβ transgenic T cells (24αβ T cells) exhibited an immature phenotype with reduced Th2 cytokine-producing capacity and diminished cytotoxicity to CD1d-expressing lymphoma cells. The impaired IL-4 production by SAP-deficient 24αβ T cells was associated with reduced IRF4 and GATA-3 induction following TCR stimulation. Collectively, these data suggest that SAP is critical for regulating type II NKT cell responses. Aberrant responses of these T cells may contribute to the immune dysregulation observed in X-linked lymphoproliferative disease caused by mutations in SAP. PMID:25236978

504 Participation of Invariant NKT Cells (Vα24Jα18) during Asthma Exacerbation in Children

PubMed Central

Carpio-Pedroza, Juan Carlos; del Rio-Navarro, Blanca Estela; del Río-Chivardí, Jaime Mariano; Morales-Flores, Amelia; Jiménez-Zamudio, Luis Antonio; Moreno-Lafont, Martha; Pedraza-Sánchez, Sigifredo; Vaughan-Figueroa, Juan Gilberto; Escobar-Gutiérrez, Alejandro

2012-01-01

Background Invariant NKT cells (or type 1 NKT cells) co-express CD3 marker and NK receptors (CD56, CD161) and use a single type of TCRα chain (Vα24Jα18 for humans), comprising CD4-CD8-, CD4+ and CD8+ subsets. Participation of these cells and their cytokines in asthmatic children, in stable conditions and under exacerbation, was studied. Methods Three groups on children (6–12 years old) were selected: 1) asthmatics under exacerbation attack (AE) within the first 24 hours after the attack and before starting any treatment; 2) asthmatics with stable asthma (SA), without symptoms for at least a month before bleeding; and 3) healthy controls (HC) without history of asthma, atopy and with normal lung function were selected in the Allergy and Clinical Immunology Service, Hospital Infantil de Mexico. Invariant NKT cells and subset levels as well as intracellular cytokines were evaluated in whole blood by 4-color flow cytometry (antibodies against CD3, CD4, CD8, CD161, Va24, IL-4 and IFN-g). Results Proportion of iNKT cells among total CD3+ cells in HC group was 0.9%, while in SA patients they were increased up to 2.6%; interestingly, during exacerbation such cells were dimished (1.8%). Concerning iNKT CD4+ cells were 0.6% in HC, 1.8% in SA, and 0.7% in AE, while iNKT CD8+ cells were 0.1% in HC, 0.7% in SA, and 0.4% in AE. Both iNKT cell subsets expressed intracellular IFN-g and IL-4 cytokines in AE, SA and HC but predominantly IFN-g in iNKT CD8+ cells from AE patients. Conclusions iNKT cells participation in asthma pathogenesis was confirmed. Increase of IFN-g production in patients with exacerbations, may provide a regulatory environment to stabilize the condition.

Leishmania infantum Exoproducts Inhibit Human Invariant NKT Cell Expansion and Activation

PubMed Central

Belo, Renata; Santarém, Nuno; Pereira, Cátia; Pérez-Cabezas, Begoña; Macedo, Fátima; Leite-de-Moraes, Maria; Cordeiro-da-Silva, Anabela

2017-01-01

Leishmania infantum is one of the major parasite species associated with visceral leishmaniasis, a severe form of the disease that can become lethal if untreated. This obligate intracellular parasite has developed diverse strategies to escape the host immune response, such as exoproducts (Exo) carrying a wide range of molecules, including parasite virulence factors, which are potentially implicated in early stages of infection. Herein, we report that L. infantum Exo and its two fractions composed of extracellular vesicles (EVs) and vesicle-depleted-exoproducts (VDEs) inhibit human peripheral blood invariant natural killer T (iNKT) cell expansion in response to their specific ligand, the glycolipid α-GalactosylCeramide (α-GalCer), as well as their capacity to promptly produce IL-4 and IFNγ. Using plate-bound CD1d and α-GalCer, we found that Exo, EV, and VDE fractions reduced iNKT cell activation in a dose-dependent manner, suggesting that they prevented α-GalCer presentation by CD1d molecules. This direct effect on CD1d was confirmed by the observation that CD1d:α-GalCer complex formation was impaired in the presence of Exo, EV, and VDE fractions. Furthermore, lipid extracts from the three compounds mimicked the inhibition of iNKT cell activation. These lipid components of L. infantum exoproducts, including EV and VDE fractions, might compete for CD1-binding sites, thus blocking iNKT cell activation. Overall, our results provide evidence for a novel strategy through which L. infantum can evade immune responses of mammalian host cells by preventing iNKT lymphocytes from recognizing glycolipids in a TCR-dependent manner. PMID:28674535

Leishmania infantum Exoproducts Inhibit Human Invariant NKT Cell Expansion and Activation.

PubMed

Belo, Renata; Santarém, Nuno; Pereira, Cátia; Pérez-Cabezas, Begoña; Macedo, Fátima; Leite-de-Moraes, Maria; Cordeiro-da-Silva, Anabela

2017-01-01

Leishmania infantum is one of the major parasite species associated with visceral leishmaniasis, a severe form of the disease that can become lethal if untreated. This obligate intracellular parasite has developed diverse strategies to escape the host immune response, such as exoproducts (Exo) carrying a wide range of molecules, including parasite virulence factors, which are potentially implicated in early stages of infection. Herein, we report that L. infantum Exo and its two fractions composed of extracellular vesicles (EVs) and vesicle-depleted-exoproducts (VDEs) inhibit human peripheral blood invariant natural killer T (iNKT) cell expansion in response to their specific ligand, the glycolipid α-GalactosylCeramide (α-GalCer), as well as their capacity to promptly produce IL-4 and IFNγ. Using plate-bound CD1d and α-GalCer, we found that Exo, EV, and VDE fractions reduced iNKT cell activation in a dose-dependent manner, suggesting that they prevented α-GalCer presentation by CD1d molecules. This direct effect on CD1d was confirmed by the observation that CD1d:α-GalCer complex formation was impaired in the presence of Exo, EV, and VDE fractions. Furthermore, lipid extracts from the three compounds mimicked the inhibition of iNKT cell activation. These lipid components of L. infantum exoproducts, including EV and VDE fractions, might compete for CD1-binding sites, thus blocking iNKT cell activation. Overall, our results provide evidence for a novel strategy through which L. infantum can evade immune responses of mammalian host cells by preventing iNKT lymphocytes from recognizing glycolipids in a TCR-dependent manner.

Regulatory iNKT cells lack expression of the transcription factor PLZF and control the homeostasis of T(reg) cells and macrophages in adipose tissue.

PubMed

Lynch, Lydia; Michelet, Xavier; Zhang, Sai; Brennan, Patrick J; Moseman, Ashley; Lester, Chantel; Besra, Gurdyal; Vomhof-Dekrey, Emilie E; Tighe, Mike; Koay, Hui-Fern; Godfrey, Dale I; Leadbetter, Elizabeth A; Sant'Angelo, Derek B; von Andrian, Ulrich; Brenner, Michael B

2015-01-01

Invariant natural killer T cells (iNKT cells) are lipid-sensing innate T cells that are restricted by the antigen-presenting molecule CD1d and express the transcription factor PLZF. iNKT cells accumulate in adipose tissue, where they are anti-inflammatory, but the factors that contribute to their anti-inflammatory nature, as well as their targets in adipose tissue, are unknown. Here we found that iNKT cells in adipose tissue had a unique transcriptional program and produced interleukin 2 (IL-2) and IL-10. Unlike other iNKT cells, they lacked PLZF but expressed the transcription factor E4BP4, which controlled their IL-10 production. The adipose iNKT cells were a tissue-resident population that induced an anti-inflammatory phenotype in macrophages and, through the production of IL-2, controlled the number, proliferation and suppressor function of regulatory T cells (Treg cells) in adipose tissue. Thus, iNKT cells in adipose tissue are unique regulators of immunological homeostasis in this tissue.

Dissection of SAP-dependent and SAP-independent SLAM family signaling in NKT cell development and humoral immunity.

PubMed

Chen, Shasha; Cai, Chenxu; Li, Zehua; Liu, Guangao; Wang, Yuande; Blonska, Marzenna; Li, Dan; Du, Juan; Lin, Xin; Yang, Meixiang; Dong, Zhongjun

2017-02-01

Signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) mutations in X-linked lymphoproliferative disease (XLP) lead to defective NKT cell development and impaired humoral immunity. Because of the redundancy of SLAM family receptors (SFRs) and the complexity of SAP actions, how SFRs and SAP mediate these processes remains elusive. Here, we examined NKT cell development and humoral immunity in mice completely deficient in SFR. We found that SFR deficiency severely impaired NKT cell development. In contrast to SAP deficiency, SFR deficiency caused no apparent defect in follicular helper T (T FH ) cell differentiation. Intriguingly, the deletion of SFRs completely rescued the severe defect in T FH cell generation caused by SAP deficiency, whereas SFR deletion had a minimal effect on the defective NKT cell development in SAP-deficient mice. These findings suggest that SAP-dependent activating SFR signaling is essential for NKT cell selection; however, SFR signaling is inhibitory in SAP-deficient T FH cells. Thus, our current study revises our understanding of the mechanisms underlying T cell defects in patients with XLP. © 2017 Chen et al.

Dissection of SAP-dependent and SAP-independent SLAM family signaling in NKT cell development and humoral immunity

PubMed Central

Cai, Chenxu; Liu, Guangao; Wang, Yuande; Du, Juan; Lin, Xin; Yang, Meixiang

2017-01-01

Signaling lymphocytic activation molecule (SLAM)–associated protein (SAP) mutations in X-linked lymphoproliferative disease (XLP) lead to defective NKT cell development and impaired humoral immunity. Because of the redundancy of SLAM family receptors (SFRs) and the complexity of SAP actions, how SFRs and SAP mediate these processes remains elusive. Here, we examined NKT cell development and humoral immunity in mice completely deficient in SFR. We found that SFR deficiency severely impaired NKT cell development. In contrast to SAP deficiency, SFR deficiency caused no apparent defect in follicular helper T (TFH) cell differentiation. Intriguingly, the deletion of SFRs completely rescued the severe defect in TFH cell generation caused by SAP deficiency, whereas SFR deletion had a minimal effect on the defective NKT cell development in SAP-deficient mice. These findings suggest that SAP-dependent activating SFR signaling is essential for NKT cell selection; however, SFR signaling is inhibitory in SAP-deficient TFH cells. Thus, our current study revises our understanding of the mechanisms underlying T cell defects in patients with XLP. PMID:28049627

Cross-talk between iNKT cells and monocytes triggers an atheroprotective immune response in SLE patients with asymptomatic plaque.

PubMed

Smith, Edward; Croca, Sara; Waddington, Kirsty E; Sofat, Reecha; Griffin, Maura; Nicolaides, Andrew; Isenberg, David A; Torra, Ines Pineda; Rahman, Anisur; Jury, Elizabeth C

2016-12-02

Accelerated atherosclerosis is a complication of the autoimmune rheumatic disease systemic lupus erythematosus (SLE). We questioned the role played by invariant natural killer T (iNKT) cells in this process because they not only are defective in autoimmunity but also promote atherosclerosis in response to CD1d-mediated lipid antigen presentation. iNKT cells from SLE patients with asymptomatic plaque (SLE-P) had increased proliferation and interleukin-4 production compared with those from SLE patients with no plaque. The anti-inflammatory iNKT cell phenotype was associated with dyslipidemia and was driven by altered monocyte phospholipid expression and CD1d-mediated cross-talk between iNKT cells and monocytes but not B cells. Healthy iNKT cells differentiated in the presence of healthy monocytes and SLE-P serum polarized macrophages toward an anti-inflammatory M2 phenotype. Conversely, patients with clinical cardiovascular disease had unresponsive iNKT cells and increased proinflammatory monocytes. iNKT cell function could link immune responses, lipids, and cardiovascular disease in SLE patients and, together with serum lipid taxonomy, help predict preclinical atherosclerosis in SLE patients. Copyright © 2016, American Association for the Advancement of Science.

Non‐glycosidic compounds can stimulate both human and mouse iNKT cells

PubMed Central

Jukes, John‐Paul; Gileadi, Uzi; Ghadbane, Hemza; Yu, Ting‐Fong; Shepherd, Dawn; Cox, Liam R.; Besra, Gurdyal S.

2016-01-01

Invariant natural killer T (iNKT) cells recognize CD1d/glycolipid complexes and upon activation with synthetic agonists display immunostimulatory properties. We have previously described that the non‐glycosidic CD1d‐binding lipid, threitolceramide (ThrCer) activates murine and human iNKT cells. Here, we show that incorporating the headgroup of ThrCer into a conformationally more restricted 6‐ or 7‐membered ring results in significantly more potent non‐glycosidic analogs. In particular, ThrCer 6 was found to promote strong anti‐tumor responses and to induce a more prolonged stimulation of iNKT cells than does the canonical α‐galactosylceramide (α‐GalCer), achieving an enhanced T‐cell response at lower concentrations compared with α‐GalCer both in vitro, using human iNKT‐cell lines and in vivo, using C57BL/6 mice. Collectively, these studies describe novel non‐glycosidic ThrCer‐based analogs that have improved potency in iNKT‐cell activation compared with that of α‐GalCer, and are clinically relevant iNKT‐cell agonists. PMID:26873393

Downregulation of BTLA on NKT Cells Promotes Tumor Immune Control in a Mouse Model of Mammary Carcinoma

PubMed Central

Sekar, Divya; Govene, Luisa; del Río, María-Luisa; Sirait-Fischer, Evelyn; Fink, Annika F.

2018-01-01

Natural Killer T cells (NKT cells) are emerging as critical regulators of pro- and anti-tumor immunity, both at baseline and in therapeutic settings. While type I NKT cells can promote anti-tumor immunity, their activity in the tumor microenvironment may be limited by negative regulators such as inhibitory immune checkpoints. We observed dominant expression of B- and T-lymphocyte attenuator (BTLA) on type I NKT cells in polyoma middle T oncogene-driven (PyMT) murine autochthonous mammary tumors. Other immune checkpoint receptors, such as programmed cell death 1 (PD-1) were equally distributed among T cell populations. Interference with BTLA using neutralizing antibodies limited tumor growth and pulmonary metastasis in the PyMT model in a therapeutic setting, correlating with an increase in type I NKT cells and expression of cytotoxic marker genes. While therapeutic application of an anti-PD-1 antibody increased the number of CD8+ cytotoxic T cells and elevated IL-12 expression, tumor control was not established. Expression of ZBTB16, the lineage-determining transcription factor of type I NKT cells, was correlated with a favorable patient prognosis in the METABRIC dataset, and BTLA levels were instrumental to further distinguish prognosis in patents with high ZBTB16 expression. Taken together, these data support a role of BTLA on type I NKT cells in limiting anti-tumor immunity. PMID:29518903

Downregulation of BTLA on NKT Cells Promotes Tumor Immune Control in a Mouse Model of Mammary Carcinoma.

PubMed

Sekar, Divya; Govene, Luisa; Del Río, María-Luisa; Sirait-Fischer, Evelyn; Fink, Annika F; Brüne, Bernhard; Rodriguez-Barbosa, José I; Weigert, Andreas

2018-03-07

Natural Killer T cells (NKT cells) are emerging as critical regulators of pro- and anti-tumor immunity, both at baseline and in therapeutic settings. While type I NKT cells can promote anti-tumor immunity, their activity in the tumor microenvironment may be limited by negative regulators such as inhibitory immune checkpoints. We observed dominant expression of B- and T-lymphocyte attenuator (BTLA) on type I NKT cells in polyoma middle T oncogene-driven (PyMT) murine autochthonous mammary tumors. Other immune checkpoint receptors, such as programmed cell death 1 (PD-1) were equally distributed among T cell populations. Interference with BTLA using neutralizing antibodies limited tumor growth and pulmonary metastasis in the PyMT model in a therapeutic setting, correlating with an increase in type I NKT cells and expression of cytotoxic marker genes. While therapeutic application of an anti-PD-1 antibody increased the number of CD8+ cytotoxic T cells and elevated IL-12 expression, tumor control was not established. Expression of ZBTB16, the lineage-determining transcription factor of type I NKT cells, was correlated with a favorable patient prognosis in the METABRIC dataset, and BTLA levels were instrumental to further distinguish prognosis in patents with high ZBTB16 expression. Taken together, these data support a role of BTLA on type I NKT cells in limiting anti-tumor immunity.

Abnormalities in iNKT cells are associated with impaired ability of monocytes to produce IL-10 and suppress T-cell proliferation in sarcoidosis.

PubMed

Crawshaw, Anjali; Kendrick, Yvonne R; McMichael, Andrew J; Ho, Ling-Pei

2014-07-01

Sarcoidosis is a multisystem granulomatous disorder characterized by marked T-cell expansion of T helper 1 (Th1) cells. The cause of T-cell overactivity is unknown. We hypothesized that interleukin-10 (IL-10) production by a yet undefined cell type might be defective, resulting in loss of regulation of T-cell activity. Focusing on IL-10-producing monocytes, we first showed that monocytes isolated from the peripheral blood of corticosteroid-naïve sarcoidosis patients (n = 51) produced less IL-10 compared to controls, and were less able to suppress T-cell proliferation. In addition, monocytic IL-10 production correlated negatively with disease activity score. As invariant natural killer T (iNKT) cells are known to both interact with monocytes and be reduced in sarcoidosis patients, we then asked whether iNKT-specific defects might be responsible for this reduced IL-10 production. We found that greater numbers of circulating iNKT cells was associated with higher IL-10 production. Moreover, iNKT cells enhanced monocytic IL-10 production in vitro. Defective IL-10 production and T-cell suppression by sarcoidosis monocytes could be restored following their coculture with iNKT cells, in a CD1d- and cell contact-dependent process. We suggest that reduced iNKT-cell numbers in sarcoidosis may lead to impaired monocytic IL-10 production and unchecked T-cell expansion in sarcoidosis. These findings provide fresh insight into the mechanism of sarcoidosis disease, and interaction between iNKT cells and monocytes. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Activation of murine invariant NKT cells promotes susceptibility to candidiasis by IL-10 induced modulation of phagocyte antifungal activity.

PubMed

Haraguchi, Norihiro; Kikuchi, Norihiro; Morishima, Yuko; Matsuyama, Masashi; Sakurai, Hirofumi; Shibuya, Akira; Shibuya, Kazuko; Taniguchi, Masaru; Ishii, Yukio

2016-07-01

Invariant NKT (iNKT) cells play an important role in a variety of antimicrobial immune responses due to their ability to produce high levels of immune-modulating cytokines. Here, we investigated the role of iNKT cells in host defense against candidiasis using Jα18-deficient mice (Jα18(-/-) ), which lack iNKT cells. Jα18(-/-) mice were more resistant to the development of lethal candidiasis than wild-type (WT) mice. In contrast, treatment of WT mice with the iNKT cell activating ligand α-galactosylceramide markedly enhanced their mortality after infection with Candida albicans. Serum IL-10 levels were significantly elevated in WT mice in response to infection with C. albicans. Futhermore, IL-10 production increased after in vitro coculture of peritoneal macrophages with iNKT cells and C. albicans. The numbers of peritoneal macrophages, the production of IL-1β and IL-18, and caspase-1 activity were also significantly elevated in Jα18(-/-) mice after infection with C. albicans. The adoptive transfer of iNKT cells or exogenous administration of IL-10 into Jα18(-/-) reversed susceptibility to candidiasis to the level of WT mice. These results suggest that activation of iNKT cells increases the initial severity of C. albicans infection, most likely mediated by IL-10 induced modulation of macrophage antifungal activity. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of age and latent CMV infection on CD8+ CD56+ T cells (NKT-like) frequency and functionality.

PubMed

Hassouneh, Fakhri; Campos, Carmen; López-Sejas, Nelson; Alonso, Corona; Tarazona, Raquel; Solana, Rafael; Pera, Alejandra

2016-09-01

Changes in the T cell pool caused by CMV infection have been proposed to contribute to immunosenescence, but it has been postulated that CMV can also have some beneficial effects in young individuals improving the immune response to other pathogens. T cells expressing CD56 (NKT-like cells) are cytotoxic effector cells with a significant role in the immune response against cancer. We have studied how age and latent CMV infection affect the frequency of NKT-like cells (CD8+ CD56+ T cells) and their response to Staphylococcal Enterotoxin B (SEB) in the context of CMV and ageing. NKT-like cell percentage increases with the combination of both CMV and age. The response to SEB and the polyfunctional index of NKT-like cells also increase with age in CMV-seropositive individuals. In young individuals, CMV infection induces a shift on the polyfunctional profile of CD8+ CD56- T cells not observed on the NKT-like cells response. NKT-like cells expressing CD57 are expanded in CMV-seropositive individuals and are more polyfunctional than their CD57-  counterpart. In addition CD57- NKT-like cells are more polyfunctional than CD8+ CD56- CD57- T cells. The results support that the expansion of polyfunctional NKT-cells may have a beneficial effect on the immune response against pathogens. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum

PubMed Central

Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G.; Garrote, José A.; Arranz, Eduardo

2015-01-01

Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum. PMID:26529008

Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum.

PubMed

Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G; Garrote, José A; Arranz, Eduardo

2015-10-30

Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum.

The immunoregulatory role of type I and type II NKT cells in cancer and other diseases

PubMed Central

Terabe, Masaki; Berzofsky, Jay A.

2014-01-01

NKT cells are CD1d-restricted T cells that recognize lipid antigens. They also have been shown to play critical roles in the regulation of immune responses. In the immune responses against tumors, two subsets of NKT cells, type I and type II, play opposing roles and cross-regulate each other. As members of both the innate and adaptive immune systems, which form a network of multiple components, they also interact with other immune components. Here we discuss the function of NKT cells in tumor immunity and their interaction with other regulatory cells, especially CD4+CD25+Foxp3+ regulatory T cells. PMID:24384834

Critical role for invariant chain in CD1d-mediated selection and maturation of Vα14-invariant NKT cells.

PubMed

Sillé, Fenna C M; Martin, Constance; Jayaraman, Pushpa; Rothchild, Alissa; Besra, Gurdyal S; Behar, Samuel M; Boes, Marianne

2011-09-30

The development and maturation of Vα14 invariant (i)NKT cells in mice requires CD1d-mediated lipid antigen presentation in the thymus and the periphery. Cortical thymocytes mediate positive selection, while professional APCs are involved in thymic negative selection and in terminal maturation of iNKT cells in the periphery. CD1d requires entry in the endosomal pathway to allow antigen acquisition for assembly as lipid/CD1d complexes for display to iNKT cells. This process involves tyrosine-based sorting motifs in the CD1d cytoplasmic tail and invariant chain (Ii) that CD1d associates with in the endoplasmic reticulum. The function of Ii in iNKT cell thymic development and peripheral maturation had not been fully understood. Using mice deficient in Ii and the Ii-processing enzyme cathepsin S (catS), we addressed this question. Ii(-/-) mice but not catS(-/-) mice developed significantly fewer iNKT cells in thymus, that were less mature as measured by CD44 and NK1.1 expression. Ii(-/-) mice but not catS(-/-) mice developed fewer Vβ7(+) cells in their iNKT TCR repertoire than WT counterparts, indicative of a change in endogenous glycolipid antigen/CD1d-mediated iNKT cell selection. Finally, using a Mycobacterium tuberculosis infection model in macrophages, we show that iNKT developed in Ii(-/-) but not catS(-/-) mice have defective effector function. Our data support a role for professional APCs expressing Ii, but no role for catS in the thymic development and peripheral terminal maturation of iNKT cells. Copyright © 2011 Elsevier B.V. All rights reserved.

The Extended Family of CD1d-Restricted NKT Cells: Sifting through a Mixed Bag of TCRs, Antigens, and Functions

PubMed Central

Macho-Fernandez, Elodie; Brigl, Manfred

2015-01-01

Natural killer T (NKT) cells comprise a family of specialized T cells that recognize lipid antigens presented by CD1d. Based on their T cell receptor (TCR) usage and antigen specificities, CD1d-restricted NKT cells have been divided into two main subsets: type I NKT cells that use a canonical invariant TCR α-chain and recognize α-galactosylceramide (α-GalCer), and type II NKT cells that use a more diverse αβ TCR repertoire and do not recognize α-GalCer. In addition, α-GalCer-reactive NKT cells that use non-canonical αβ TCRs and CD1d-restricted T cells that use γδ or δ/αβ TCRs have recently been identified, revealing further diversity among CD1d-restricted T cells. Importantly, in addition to their distinct antigen specificities, functional differences are beginning to emerge between the different members of the CD1d-restricted T cell family. In this review, while using type I NKT cells as comparison, we will focus on type II NKT cells and the other non-invariant CD1d-restricted T cell subsets, and discuss our current understanding of the antigens they recognize, the formation of stimulatory CD1d/antigen complexes, the modes of TCR-mediated antigen recognition, and the mechanisms and consequences of their activation that underlie their function in antimicrobial responses, anti-tumor immunity, and autoimmunity. PMID:26284062

Enhanced tumor metastasis in response to blockade of the chemokine receptor CXCR6 is overcome by NKT cell activation.

PubMed

Cullen, Robyn; Germanov, Elitza; Shimaoka, Takeshi; Johnston, Brent

2009-11-01

Invariant NKT (iNKT) cells can induce potent antitumor responses in vivo. However, the mechanisms that regulate the effects of iNKT cells are unclear. The chemokine receptor CXCR6, and its ligand CXCL16, have been shown to play critical roles in iNKT cell homeostasis and activation. Thus we investigated the role of CXCR6 in protection against experimental metastasis of B16-F10 melanoma (B16) and Lewis lung carcinoma (LLC) cells to the liver and lungs. Wild-type and CXCR6(-/-) mice exhibited no differences in tumor cell metastasis to the lungs. However, metastasis of LLC and B16 tumor cells to the liver was enhanced in CXCR6(-/-) mice. Liver metastasis was also increased in wild-type mice treated with a CXCL16 neutralizing Ab. As Ab treatments did not alter iNKT cell numbers, this implicates a direct role for CXCR6/CXCL16 in regulating antitumor immunity. Cytokine induction was significantly attenuated in CXCR6(-/-) mice upon systemic iNKT cell activation with the glycolipid Ags alpha-galactosylceramide (alpha-GalCer), alpha-C-GalCer (a Th1 polarizing derivative), or OCH (a Th2 polarizing derivative). Despite differences in the levels of cytokine production, liver and lung metastasis were inhibited significantly in both wild-type and CXCR6(-/-) mice treated with glycolipids. Single doses of alpha-GalCer, alpha-C-GalCer, or OCH were sufficient to prevent liver metastasis and subsequent doses failed to elicit optimal cytokine responses. Our findings implicate a role for CXCR6 in natural immunosurveillance against liver metastasis. However, CXCR6 deficiency could be overcome by systemic iNKT cell activation, demonstrating that even suboptimal iNKT cell activation can protect against metastasis.

Activation of iNKT cells by a distinct constituent of the endogenous glucosylceramide fraction

PubMed Central

Brennan, Patrick J.; Tatituri, Raju V. V.; Heiss, Christian; Watts, Gerald F. M.; Hsu, Fong-Fu; Veerapen, Natacha; Cox, Liam R.; Azadi, Parastoo; Besra, Gurdyal S.; Brenner, Michael B.

2014-01-01

Invariant natural killer T (iNKT) cells are a specialized T-cell subset that recognizes lipids as antigens, contributing to immune responses in diverse disease processes. Experimental data suggests that iNKT cells can recognize both microbial and endogenous lipid antigens. Several candidate endogenous lipid antigens have been proposed, although the contextual role of specific antigens during immune responses remains largely unknown. We have previously reported that mammalian glucosylceramides (GlcCers) activate iNKT cells. GlcCers are found in most mammalian tissues, and exist in variable molecular forms that differ mainly in N-acyl fatty acid chain use. In this report, we purified, characterized, and tested the GlcCer fractions from multiple animal species. Although activity was broadly identified in these GlcCer fractions from mammalian sources, we also found activity properties that could not be reconciled by differences in fatty acid chain use. Enzymatic digestion of β-GlcCer and a chromatographic separation method demonstrated that the activity in the GlcCer fraction was limited to a rare component of this fraction, and was not contained within the bulk of β-GlcCer molecular species. Our data suggest that a minor lipid species that copurifies with β-GlcCer in mammals functions as a lipid self antigen for iNKT cells. PMID:25197085

Interleukin-4-dependent innate collaboration between iNKT cells and B-1 B cells controls adaptative contact sensitivity

PubMed Central

Campos, Regis A; Szczepanik, Marian; Itakura, Atsuko; Lisbonne, Mariette; Dey, Neelendu; Leite-de-Moraes, Maria C; Askenase, Philip W

2006-01-01

We showed that hepatic Vα14+ invariant natural killer T (iNKT) cells, via their rapid interleukin (IL)-4 production, activate B-1 cells to initiate contact sensitivity (CS). This innate collaboration was absent in IL-4–/– and signal transducer and activator of transcription (STAT)-6–/– mice and was inhibited by anti-IL-4 treatment. These mice have defective CS because they fail to locally recruit the sensitized effector T cells of acquired immunity. Their CS is reconstituted by transfer of downstream-acting 1-day immune B-1 cells from wild-type mice. Responses were not reconstituted with B-1 cells from IL-4 receptor-α–/– or STAT-6–/– mice, nor by IL-4 treatment of B cell-deficient mice at immunization. Finally, IL-4 was preferentially and transiently produced by hepatic iNKT cells within 7 min after sensitization to mediate collaboration between innate-like iNKT cells and the B-1 B cells that participate in the recruitment of effector T cells in vivo. PMID:16556268

Allelic Variation of Ets1 Does Not Contribute to NK and NKT Cell Deficiencies in Type 1 Diabetes Susceptible NOD Mice

PubMed Central

Jordan, Margaret A.; Poulton, Lynn D.; Fletcher, Julie M.; Baxter, Alan G.

2009-01-01

The NOD mouse is a well characterized model of type 1 diabetes that shares several of the characteristics of Ets1-deficient targeted mutant mice, viz: defects in TCR allelic exclusion, susceptibility to a lupus like disease characterized by IgM and IgG autoantibodies and immune complex-mediated glomerulonephritis, and deficiencies of NK and NKT cells. Here, we sought evidence for allelic variation of Ets1 in mice contributing to the NK and NKT cell phenotypes of the NOD strain. ETS1 expression in NK and NKT cells was reduced in NOD mice, compared to C57BL/6 mice. Although NKT cells numbers were significantly correlated with ETS1 expression in both strains, NKT cell numbers were not linked to the Ets1 gene in a first backcross from NOD to C57BL/6 mice. These results indicate that allelic variation of Ets1 did not contribute to variation in NKT cell numbers in these mice. It remains possible that a third factor not linked to the Ets1 locus controls both ETS1 expression and subsequently NK and NKT cell phenotypes. PMID:19806240

Amide Analogues of CD1d Agonists Modulate iNKT-Cell-Mediated Cytokine Production

PubMed Central

2012-01-01

Invariant natural killer T (iNKT) cells are restricted by the non-polymorphic MHC class I-like protein, CD1d, and activated following presentation of lipid antigens bound to CD1d molecules. The prototypical iNKT cell agonist is α-galactosyl ceramide (α-GalCer). CD1d-mediated activation of iNKT cells by this molecule results in the rapid secretion of a range of pro-inflammatory (Th1) and regulatory (Th2) cytokines. Polarization of the cytokine response can be achieved by modifying the structure of the glycolipid, which opens up the possibility of using CD1d agonists as therapeutic agents for a range of diseases. Analysis of crystal structures of the T-cell receptor−α-GalCer–CD1d complex led us to postulate that amide isosteres of known CD1d agonists should modulate the cytokine response profile upon iNKT-cell activation. To this end, we describe the synthesis and biological activity of amide analogues of α-GalCer and its non-glycosidic analogue threitol ceramide (ThrCer). All of the analogues were found to stimulate murine and human iNKT cells by CD1d-mediated presentation to varying degrees; however, the thioamide and carbamate analogues of ThrCer were of particular interest in that they elicited a strongly polarized cytokine response (more interferon-gamma (IFN-γ), no interleukin-4 (IL-4)) in mice. While the ThrCer-carbamate analogue was shown to transactivate natural killer (NK) cells, a mechanism that has been used to account for the preferential production of IFN-γ by other CD1d agonists, this pathway does not account for the polarized cytokine response observed for the thioamide analogue. PMID:22324848

Human Cytomegalovirus (HCMV) US2 Protein Interacts with Human CD1d (hCD1d) and Down-Regulates Invariant NKT (iNKT) Cell Activity

PubMed Central

Han, Jihye; Rho, Seung Bae; Lee, Jae Yeon; Bae, Joonbeom; Park, Se Ho; Lee, Suk Jun; Lee, Sang Yeol; Ahn, Curie; Kim, Jae Young; Chun, Taehoon

2013-01-01

To avoid host immune surveillance, human cytomegalovirus (HCMV) encoded endoplasmic reticulum (ER)-membrane glycoprotein US2, which interferes with antigen presenting mechanism of major histocompatibility complex (MHC) class Ia and class II molecules. However, not many attempts have been made to study the effect of HCMV US2 on the expression of MHC class Ib molecules. In this study, we examined the effect of HCMV US2 on the expression and function of human CD1d (hCD1d), which presents glycolipid antigens to invariant NKT (iNKT) cells. Our results clearly showed that the physiological interaction between ER lumenal domain of HCMV US2 and α3 domain of hCD1d was observed within ER. Compared with mature form of hCD1d, immature form of hCD1d is more susceptible to ubiquitin-dependent proteasomal degradation mediated by HCMV US2. Moreover, the ectopic expression of HCMV US2 leads to the down-modulation of iNKT cell activity without significant change of hCD1d expression. These results will advance our understanding of the function of HCMV US2 in immune evasive mechanisms against anti-viral immunity of iNKT cells. PMID:24213674

A mutation within the SH2 domain of slp-76 regulates the tissue distribution and cytokine production of iNKT cells in mice.

PubMed

Danzer, Claudia; Koller, Anna; Baier, Julia; Arnold, Harald; Giessler, Claudia; Opoka, Robert; Schmidt, Stephanie; Willers, Maike; Mihai, Sidonia; Parsch, Hans; Wirtz, Stefan; Daniel, Christoph; Reinhold, Annegret; Engelmann, Swen; Kliche, Stefanie; Bogdan, Christian; Hoebe, Kasper; Mattner, Jochen

2016-09-01

TCR ligation is critical for the selection, activation, and integrin expression of T lymphocytes. Here, we explored the role of the TCR adaptor protein slp-76 on iNKT-cell biology. Compared to B6 controls, slp-76(ace/ace) mice carrying a missense mutation (Thr428Ile) within the SH2-domain of slp-76 showed an increase in iNKT cells in the thymus and lymph nodes, but a decrease in iNKT cells in spleens and livers, along with reduced ADAP expression and cytokine response. A comparable reduction in iNKT cells was observed in the livers and spleens of ADAP-deficient mice. Like ADAP(-/-) iNKT cells, slp-76(ace/ace) iNKT cells were characterized by enhanced CD11b expression, correlating with an impaired induction of the TCR immediate-early gene Nur77 and a decreased adhesion to ICAM-1. Furthermore, CD11b-intrinsic effects inhibited cytokine release, concanavalin A-mediated inflammation, and iNKT-cell accumulation in the liver. Unlike B6 and ADAP(-/-) mice, the expression of the transcription factors Id3 and PLZF was reduced, whereas NP-1-expression was enhanced in slp-76(ace/ace) mice. Blockade of NP-1 decreased the recovery of iNKT cells from peripheral lymph nodes, identifying NP-1 as an iNKT-cell-specific adhesion factor. Thus, slp-76 contributes to the regulation of the tissue distribution, PLZF, and cytokine expression of iNKT cells via ADAP-dependent and -independent mechanisms. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Immunosuppressive myeloid-derived suppressor cells can be converted into immunogenic APCs with the help of activated NKT cells: an alternative cell-based antitumor vaccine.

PubMed

Ko, Hyun-Jeong; Lee, Jung-Mi; Kim, Yeon-Jeong; Kim, Yun-Sun; Lee, Kyoo-A; Kang, Chang-Yuil

2009-02-15

Myeloid-derived suppressor cells (MDSCs), which are known to be accumulated in the blood, spleen, and bone marrow of tumor-bearing mice and cancer patients, were tested as APCs for a cellular vaccine because they have phenotypical similarity with inflammatory monocytes and may be differentiated from the same precursors as monocytes. Although MDSCs have immunosuppressive properties, in vivo transferred MDSCs, which present tumor Ag and NKT cell ligand (alpha-galactosylceramide), significantly prolonged survival time in metastatic tumor-bearing mice in a CD8(+) cell-, NK cell-, and NKT cell-dependent manner vs a CD4(+) T cell- and host dendritic cell-independent manner. Major concerns about using MDSCs as APCs in a vaccine are their suppression of CTLs and their induction of Foxp3(+) regulatory T cells. However, alpha-galactosylceramide-loaded MDSCs did not suppress CD4(+) and CD8(+) T cells and allowed for the generation of Ag-specific CTL immunity without increasing the generation of regulatory T cells. Furthermore, stimulation with activated NKT cells induced changes on MDSCs in phenotypical or maturation markers, including CD11b, CD11c, and CD86. Taken together, these findings suggest that NKT cells facilitate the conversion of immunosuppressive MDSCs into immunogenic APCs, eliciting successful antitumor immunity and providing the basis for alternative cell-based vaccines.

CD1d-Expressing Breast Cancer Cells Modulate NKT Cell-Mediated Antitumor Immunity in a Murine Model of Breast Cancer Metastasis

PubMed Central

Hix, Laura M.; Shi, Yihui H.; Brutkiewicz, Randy R.; Stein, Paul L.; Wang, Chyung-Ru; Zhang, Ming

2011-01-01

Background Tumor tolerance and immune suppression remain formidable obstacles to the efficacy of immunotherapies that harness the immune system to eradicate breast cancer. A novel syngeneic mouse model of breast cancer metastasis was developed in our lab to investigate mechanisms of immune regulation of breast cancer. Comparative analysis of low-metastatic vs. highly metastatic tumor cells isolated from these mice revealed several important genetic alterations related to immune control of cancer, including a significant downregulation of cd1d1 in the highly metastatic tumor cells. The cd1d1 gene in mice encodes the MHC class I-like molecule CD1d, which presents glycolipid antigens to a specialized subset of T cells known as natural killer T (NKT) cells. We hypothesize that breast cancer cells, through downregulation of CD1d and subsequent evasion of NKT-mediated antitumor immunity, gain increased potential for metastatic tumor progression. Methodology/Principal Findings In this study, we demonstrate in a mouse model of breast cancer metastasis that tumor downregulation of CD1d inhibits iNKT-mediated antitumor immunity and promotes metastatic breast cancer progression in a CD1d-dependent manner in vitro and in vivo. Using NKT-deficient transgenic mouse models, we demonstrate important differences between type I and type II NKT cells in their ability to regulate antitumor immunity of CD1d-expressing breast tumors. Conclusions/Significance The results of this study emphasize the importance of determining the CD1d expression status of the tumor when tailoring NKT-based immunotherapies for the prevention and treatment of metastatic breast cancer. PMID:21695190

Metabolic regulator Fnip1 is crucial for iNKT lymphocyte development

PubMed Central

Park, Heon; Tsang, Mark; Iritani, Brian M.; Bevan, Michael J.

2014-01-01

Folliculin-interacting protein 1 (Fnip1) is an adaptor protein that physically interacts with AMPK, an energy-sensing kinase that stimulates mitochondrial biogenesis and autophagy in response to low ATP, while turning off energy consumption mediated by mammalian target of rapamycin. Previous studies with Fnip1-null mice revealed that Fnip1 is essential for pre–B-cell development. Here we report a critical role of Fnip1 in invariant natural killer T (iNKT) cell development. Thymic iNKT development in Fnip1−/− mice was arrested at stage 2 (NK1.1−CD44+) but development of CD4, CD8, γδ T-cell, and NK cell lineages proceeded normally. Enforced expression of a Vα14Jα18 iNKT TCR transgene or loss of the proapoptotic protein Bim did not rescue iNKT cell maturation in Fnip1−/− mice. Whereas most known essential transcription factors for iNKT cell development were represented normally, Fnip1−/− iNKT cells failed to down-regulate Promyelocytic leukemia zinc finger compared with their WT counterparts. Moreover, Fnip1−/− iNKT cells contained hyperactive mTOR and reduced mitochondrial number despite lower ATP levels, resulting in increased sensitivity to apoptosis. These results indicate that Fnip1 is vital for iNKT cell development by maintaining metabolic homeostasis in response to metabolic stress. PMID:24785297

A brief review of clinical trials involving manipulation of invariant NKT cells as a promising approach in future cancer therapies

PubMed Central

Bojarska-Junak, Agnieszka; Roliński, Jacek

2017-01-01

In the recent years researchers have put a lot of emphasis on the possible immunotherapeutic strategies able to target tumors. Many studies have proven that the key role in recognition and eradication of cancer cells, both for mice and humans, is being conducted by the invariant natural killer T-cells (NKT). This small subpopulation of lymphocytes can kill other cells, either directly or indirectly, through the natural killer cells’ (NK) activation. They can also swiftly release cytokines, causing the involvement of elements of the innate and acquired immune system. With the discovery of α-galactosylceramide (α-GalCer) – the first known agonist for iNKT cells – and its later subsequent analogs, it became possible to effectively stimulate iNKT cells, hence to keep control over the tumor progression. This article refers to the current knowledge concerning iNKT cells and the most important aspects of their antitumor activity. It also highlights the clinical trials that aim at increasing the amount of iNKT cells in general and in the microenvironment of the tumor. For sure, the iNKT-based immunotherapeutic approach holds a great potential and is highly probable to become a part of the cancer immunotherapy in the future. PMID:28860937

AHR-Enhancing γδ T Cells Develop in Normal Untreated Mice and Fail to Produce IL-4/13, Unlike TH2 Cells and NKT Cells1

PubMed Central

Jin, Niyun; Roark, Christina L.; Miyahara, Nobuaki; Taube, Christian; Aydintug, M. Kemal; Wands, JM; Huang, Yafei; Hahn, Youn-Soo; Gelfand, Erwin W.; O’Brien, Rebecca L.; Born, Willi K.

2008-01-01

Allergic airway hyperresponsiveness (AHR) in OVA-sensitized and challenged mice, mediated by allergen-specific Th2 cells and Th2-like iNKT cells, develops under the influence of enhancing and inhibitory γδ T cells. The AHR-enhancing cells belong to the Vγ1+ γδ T cell subset, cells that are capable of increasing IL-5 and IL-13 levels in the airways in a manner like Th2 cells. They also synergize with iNKT cells in mediating AHR. However, unlike Th2 cells, the AHR-enhancers arise in untreated mice, and we show here that they exhibit their functional bias already as thymocytes, at an HSAhi maturational stage. In further contrast to Th2 cells and also unlike iNKT cells, they could not be stimulated to produce IL-4 and IL-13, consistent with their synergistic dependence on iNKT cells in mediating AHR. Mice deficient in IFN-γ, TNFRp75 or IL-4 did not produce these AHR-enhancing γδ T cells, but in the absence of IFN-γ, their spontaneous development was restored by adoptive transfer of IFN-γ competent dendritic cells from untreated donors. Intra-peritoneal injection of OVA/alum restored development of the AHR-enhancers in all of the mutant strains, indicating that the enhancers still can be induced when they fail to develop spontaneously, and that they themselves need not express TNFRp75, IFN-γ or IL-4 in order to exert their function. We conclude that both the development and the cytokine potential of the AHR-enhancing γδ T cells differs critically from that of Th2 cells and NKT cells, despite similar influences of these cell populations on AHR. PMID:19201853

Accumulation of invariant NKT cells with increased IFN-γ production in persistent high-risk HPV-infected high-grade cervical intraepithelial neoplasia.

PubMed

Hu, Ting; Yang, Pei; Zhu, Hongmei; Chen, Xinlian; Xie, Xiaoyan; Yang, Mei; Liu, Shanling; Wang, He

2015-04-02

Persistent high-risk human papillomavirus (HR-HPV) infection has been implicated in the development of high-grade cervical intraepithelial neoplasia (CIN) and cervical cancer. Invariant natural killer T (iNKT) cells produce large amounts of cytokines to regulate immune responses. However, the role of iNKT cells in human persistent HPV-infected cervical tissues is unknown. In our study, 201 patients with diagnoses ranging from normal ectocervical tissue to CINIII from June 2010 to May 2012 were enrolled. HPV DNA and HPV types were detected using the hybrid capture-2 HPV DNA test. Flow cytometry was used to investigate iNKT and CD3+ T cell infiltration into cervical tissues. Real-time quantitative reverse transcription-polymerase chain reaction was used to study IFN-γ expression and immunohistochemistry was used to determine CD3+ T cell distribution. A significant increase in iNKT cells was observed in HPV-positive cervical tissues (p < 0.05), especially in CINII-III (p < 0.01). IFN-γ expression was also increased in HPV-positive cervical tissues (p < 0.05). CD3+ T cells were detected among both epithelium and stromal layers in cervical tissues, and the percentage of CD3+ T cells in HPV-positive cervical tissues was similar to that in HPV-negative cervical tissues (p > 0.05). The iNKT cell aggregation in cervical tissues during the progression from HPV infection to CIN indicates that iNKT cells might play an important role in suppressing immunity. IFN-γ expression could also be related to the HPV infection status. Preventing the accumulation or functioning of iNKT cells in cervical tissues may be a viable method to prevent the development of CIN. The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2521874671514142.

Identification of an IL-17–producing NK1.1neg iNKT cell population involved in airway neutrophilia

PubMed Central

Michel, Marie-Laure; Keller, Alexandre Castro; Paget, Christophe; Fujio, Masakazu; Trottein, François; Savage, Paul B.; Wong, Chi-Huey; Schneider, Elke; Dy, Michel; Leite-de-Moraes, Maria C.

2007-01-01

Invariant natural killer T (iNKT) cells are an important source of both T helper type 1 (Th1) and Th2 cytokines, through which they can exert beneficial, as well as deleterious, effects in a variety of inflammatory diseases. This functional heterogeneity raises the question of how far phenotypically distinct subpopulations are responsible for such contrasting activities. In this study, we identify a particular set of iNKT cells that lack the NK1.1 marker (NK1.1neg) and secrete high amounts of interleukin (IL)-17 and low levels of interferon (IFN)-γ and IL-4. NK1.1neg iNKT cells produce IL-17 upon synthetic (α-galactosylceramide [α-GalCer] or PBS-57), as well as natural (lipopolysaccharides or glycolipids derived from Sphingomonas wittichii and Borrelia burgdorferi), ligand stimulation. NK1.1neg iNKT cells are more frequent in the lung, which is consistent with a role in the natural immunity to inhaled antigens. Indeed, airway neutrophilia induced by α-GalCer or lipopolysaccharide instillation was significantly reduced in iNKT-cell–deficient Jα18−/− mice, which produced significantly less IL-17 in their bronchoalveolar lavage fluid than wild-type controls. Furthermore, airway neutrophilia was abolished by a single treatment with neutralizing monoclonal antibody against IL-17 before α-GalCer administration. Collectively, our findings reveal that NK1.1neg iNKT lymphocytes represent a new population of IL-17–producing cells that can contribute to neutrophil recruitment through preferential IL-17 secretion. PMID:17470641

IL-30 (IL27p28) alleviates sepsis via modulation of cytokine profiles produced by NKT cells

PubMed Central

Yan, Jun; Mitra, Abhisek; Hu, Jiemiao; Cutrera, Jeffery J; Xia, Xueqing; Doetschman, Thomas; Gagea, Mihai; Mishra, Lopa; Li, Shulin

2016-01-01

Background & Aims Sepsis is an acute systemic inflammatory response to infection associated with high patient mortality (28-40%). We hypothesized that interleukin (IL)-30, a novel cytokine protecting mice against liver injury resulted from inflammation, would generate a protective effect against systemic inflammation and sepsis-induced death. Methods Sepsis was induced by lipopolysaccharide (LPS) or cecal ligation and puncture (CLP). The inhibitory effects of IL-30 on septic inflammation and associated therapeutic effects were determined in wild-type, IL-30 (p28)−/−, IL10−/−, and CD1d−/− mice. Results Mice treated with pIL30 gene therapy or recombinant IL-30 protein (rIL30) were protected from LPS-induced septic shock or CLP-induced polymicrobial sepsis and showed markedly less liver damage and lymphocyte apoptosis than control septic mice. The resulting reduction in mortality was mediated through attenuation of the systemic pro-inflammatory response and augmentation of bacterial clearance. Mice lacking IL-30 were more sensitive to LPS-induced sepsis. Natural killer–like T cells (NKT) produced much higher levels of IL-10 and lower levels of interferon–gamma and tumor necrosis factor–alpha in IL-30–treated septic mice than in control septic mice. Likewise, deficiency in IL-10 or NKT cells abolished the protective role of IL-30 against sepsis. Furthermore, IL-30 induced IL-10 production in purified and LPS-stimulated NKT cells. Blocking IL-6R or gp130 inhibited IL-30 mediated IL-10 production. Conclusions IL-30 is important in modulating production of NKT cytokines and subsequent NKT cell–mediated immune regulation of other cells. Therefore, IL-30 has a role in prevention and treatment of sepsis via modulation of cytokine production by NKT. PMID:26767500

Human iNKT cells induce IL-1β secretion by peripheral blood monocytes via a P2X7-independent pathway

PubMed Central

Felley, Laura E.; Sharma, Akshat; Theisen, Erin; Romero-Masters, James C.; Sauer, John-Demian; Gumperz, Jenny E.

2016-01-01

The cytokine IL-1β plays a central role in inflammatory responses that are initiated by microbial challenges, as well as in those that are due to endogenous processes (often called “sterile” inflammation). IL-1β secretion that occurs independently of microbial stimulation is typically associated with the presence of endogenous alarmins, such as extracellular ATP (an indicator of cytopathic damage). Here we show that IL-2 activated human iNKT cells stimulate the secretion of IL-1β protein by human peripheral blood monocytes in a manner that requires neither the presence of microbial compounds nor signaling through the extracellular ATP receptor P2X7. Monocyte IL-1β production was specifically induced by iNKT cells, since similarly activated polyclonal autologous T cells did not have this effect. Secretion of IL-1β protein occurred rapidly (within 3-4 hours), and required cell contact between the iNKT cells and monocytes. Similar to IL-1β production induced by TLR stimulation, the iNKT-induced pathway appeared to entail a two-step process involving NFκB signaling and IL1B gene transcription, as well as assembly of the NLRP3 inflammasome and activation of caspase 1. However, in contrast to the classical inflammasome-mediated pathway of IL-1β production, activation of monocytes via P2X7 was dispensable for iNKT-induced IL-1β secretion and potassium efflux was not required. Moreover, the iNKT-induced effect involved caspase 8 activity, yet induced little monocyte death. These results suggest that IL-2 activated human iNKT cells induce monocytes to produce IL-1β through a distinctive pathway that does not require the presence of microbial danger signals or alarmins associated with cytopathic damage. PMID:27534556

Multiple layers of transcriptional regulation by PLZF in NKT-cell development.

PubMed

Mao, Ai-Ping; Constantinides, Michael G; Mathew, Rebecca; Zuo, Zhixiang; Chen, Xiaoting; Weirauch, Matthew T; Bendelac, Albert

2016-07-05

The transcription factor PLZF [promyelocytic leukemia zinc finger, encoded by zinc finger BTB domain containing 16 (Zbtb16)] is induced during the development of innate and innate-like lymphocytes to direct their acquisition of a T-helper effector program, but the molecular mechanisms involved are poorly understood. Using biotinylation-based ChIP-seq and microarray analysis of both natural killer T (NKT) cells and PLZF-transgenic thymocytes, we identified several layers of regulation of the innate-like NKT effector program. First, PLZF bound and regulated genes encoding cytokine receptors as well as homing and adhesion receptors; second, PLZF bound and activated T-helper-specific transcription factor genes that in turn control T-helper-specific programs; finally, PLZF bound and suppressed the transcription of Bach2, a potent general repressor of effector differentiation in naive T cells. These findings reveal the multilayered architecture of the transcriptional program recruited by PLZF and elucidate how a single transcription factor can drive the developmental acquisition of a broad effector program.

iNKT-CELL ACTIVATION INDUCES LATE PRETERM BIRTH THAT IS ATTENUATED BY ROSIGLITAZONE1

PubMed Central

St Louis, Derek; Romero, Roberto; Plazyo, Olesya; Arenas-Hernandez, Marcia; Panaitescu, Bogdan; Xu, Yi; Milovic, Tatjana; Xu, Zhonghui; Bhatti, Gaurav; Qing-Sheng, Mi; Drewlo, Sascha; Tarca, Adi L.; Hassan, Sonia S.; Gomez-Lopez, Nardhy

2015-01-01

Preterm birth (PTB) is a leading cause of neonatal morbidity and mortality; however, its non-infection-related mechanisms are poorly understood. Herein, we show that the expansion of activated CD1d-restricted invariant NKT (iNKT) cells in the third trimester by administration of α-galactosylceramide (α-GalCer) induces late PTB and neonatal mortality. In vivo imaging revealed that fetuses from mice that underwent α-GalCer-induced late PTB had bradycardia and died shortly after delivery. Yet, administration of α-GalCer in the second trimester did not cause pregnancy loss. PPARγ activation, through rosiglitazone treatment, reduced the rate of α-GalCer-induced late PTB and improved neonatal survival. Administration of α-GalCer in the third trimester suppressed PPARγ activation as shown by the down-regulation of Fabp4 and Fatp4 in myometrial and decidual tissues, respectively; this suppression was rescued by rosiglitazone treatment. Administration of α-GalCer in the third trimester induced an increase in the activation of conventional CD4+ T cells in myometrial tissues and the infiltration of activated macrophages, neutrophils and mature DCs to myometrial and/or decidual tissues. All of these effects were blunted after rosiglitazone treatment. Administration of α-GalCer also up-regulated the expression of inflammatory genes at the maternal-fetal interface and systemically, and rosiglitazone treatment partially attenuated these responses. Finally, an increased infiltration of activated iNKT-like cells in human decidual tissues is associated with non-infection-related preterm labor/birth. Collectively, these results demonstrate that iNKT-cell activation in vivo leads to late PTB by initiating innate and adaptive immune responses and suggest that the PPARγ pathway has potential as a target for prevention of this syndrome. PMID:26740111

CRISPR-Mediated Triple Knockout of SLAMF1, SLAMF5 and SLAMF6 Supports Positive Signaling Roles in NKT Cell Development.

PubMed

Huang, Bonnie; Gomez-Rodriguez, Julio; Preite, Silvia; Garrett, Lisa J; Harper, Ursula L; Schwartzberg, Pamela L

2016-01-01

The SLAM family receptors contribute to diverse aspects of lymphocyte biology and signal via the small adaptor molecule SAP. Mutations affecting SAP lead to X-linked lymphoproliferative syndrome Type 1, a severe immunodysregulation characterized by fulminant mononucleosis, dysgammaglobulinemia, and lymphoproliferation/lymphomas. Patients and mice having mutations affecting SAP also lack germinal centers due to a defect in T:B cell interactions and are devoid of invariant NKT (iNKT) cells. However, which and how SLAM family members contribute to these phenotypes remains uncertain. Three SLAM family members: SLAMF1, SLAMF5 and SLAMF6, are highly expressed on T follicular helper cells and germinal center B cells. SLAMF1 and SLAMF6 are also implicated in iNKT development. Although individual receptor knockout mice have limited iNKT and germinal center phenotypes compared to SAP knockout mice, the generation of multi-receptor knockout mice has been challenging, due to the genomic linkage of the genes encoding SLAM family members. Here, we used Cas9/CRISPR-based mutagenesis to generate mutations simultaneously in Slamf1, Slamf5 and Slamf6. Genetic disruption of all three receptors in triple-knockout mice (TKO) did not grossly affect conventional T or B cell development and led to mild defects in germinal center formation post-immunization. However, the TKO worsened defects in iNKT cells development seen in SLAMF6 single gene-targeted mice, supporting data on positive signaling and potential redundancy between these receptors.

Critical role of dendritic cell-derived IL-27 in antitumor immunity through regulating the recruitment and activation of NK and NKT cells.

PubMed

Wei, Jun; Xia, Siyuan; Sun, Huayan; Zhang, Song; Wang, Jingya; Zhao, Huiyuan; Wu, Xiaoli; Chen, Xi; Hao, Jianlei; Zhou, Xinglong; Zhu, Zhengmao; Gao, Xiang; Gao, Jian-xin; Wang, Puyue; Wu, Zhenzhou; Zhao, Liqing; Yin, Zhinan

2013-07-01

Critical roles of IL-27 in autoimmune diseases and infections have been reported; however, the contribution of endogenous IL-27 to tumor progression remains elusive. In this study, by using IL-27p28 conditional knockout mice, we demonstrate that IL-27 is critical in protective immune response against methyl-cholanthrene-induced fibrosarcoma and transplanted B16 melanoma, and dendritic cells (DCs) are the primary source. DC-derived IL-27 is required for shaping tumor microenvironment by inducing CXCL-10 expression in myeloid-derived suppressor cells and regulating IL-12 production from DCs, which lead to the recruitment and activation of NK and NKT cells resulting in immunological control of tumors. Indeed, reconstitution of IL-27 or CXCL-10 in tumor site significantly inhibits tumor growth and restores the number and activation of NK and NKT cells. In summary, our study identifies a previous unknown critical role of DC-derived IL-27 in NK and NKT cell-dependent antitumor immunity through shaping tumor microenvironment, and sheds light on developing novel therapeutic approaches based on IL-27.

Protection Against Type 1 Diabetes Upon Coxsackievirus B4 Infection and iNKT-Cell Stimulation

PubMed Central

Ghazarian, Liana; Diana, Julien; Beaudoin, Lucie; Larsson, Pär G.; Puri, Raj K.; van Rooijen, Nico; Flodström-Tullberg, Malin; Lehuen, Agnès

2013-01-01

Invariant natural killer T (iNKT) cells belong to the innate immune system and exercise a dual role as potent regulators of autoimmunity and participate in responses against different pathogens. They have been shown to prevent type 1 diabetes development and to promote antiviral responses. Many studies in the implication of environmental factors on the etiology of type 1 diabetes have suggested a link between enteroviral infections and the development of this disease. This study of the pancreatropic enterovirus Coxsackievirus B4 (CVB4) shows that although infection accelerated type 1 diabetes development in a subset of proinsulin 2–deficient NOD mice, the activation of iNKT cells by a specific agonist, α-galactosylceramide, at the time of infection inhibited the disease. Diabetes development was associated with the infiltration of pancreatic islets by inflammatory macrophages, producing high levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α and activation of anti-islet T cells. On the contrary, macrophages infiltrating the islets after CVB4 infection and iNKT-cell stimulation expressed a number of suppressive enzymes, among which indoleamine 2,3-dioxygenase was sufficient to inhibit anti-islet T-cell response and to prevent diabetes. This study highlights the critical interaction between virus and the immune system in the acceleration or prevention of type 1 diabetes. PMID:23894189

CD205-TLR9-IL-12 axis contributes to CpG-induced oversensitive liver injury in HBsAg transgenic mice by promoting the interaction of NKT cells with Kupffer cells.

PubMed

Hou, Xin; Hao, Xiaolei; Zheng, Meijuan; Xu, Congfei; Wang, Jun; Zhou, Rongbin; Tian, Zhigang

2017-08-01

Gut-derived bacterial products contribute to liver inflammation and injury during chronic hepatitis B virus infection; however, the underlying mechanisms remain obscure. In this study, hepatitis B surface antigen transgenic (HBs-Tg) mice and their wild-type (WT) control C57BL/6 mice were injected with CpG-oligodeoxynucleotides (ODNs) to mimic the translocation of gut microbial products into the systemic circulation. We found that, compared with the WT mice, the HBs-Tg mice were oversensitive to CpG-ODN-induced liver injury, which was dependent on natural killer T (NKT) cells. CpG-ODN injection enhanced the expression of Fas ligand (FasL) on NKT cells. In addition, hepatocytes from the HBs-Tg mice expressed higher levels of Fas than did those from the WT mice, which was further augmented by CpG-ODN. Interaction of Fas and FasL was involved in the cytotoxicity of NKT cells against hepatocytes in the HBs-Tg mice. Moreover, Kupffer cells in the HBs-Tg mice expressed higher levels of CD205 and produced greater amounts of interleukin (IL)-12 than did those in the WT mice. Finally, the depletion of Kupffer cells, neutralization of IL-12 or specific silencing of CD205 on Kupffer cells significantly inhibited CpG-ODN-induced liver injury and NKT activation in the HBs-Tg mice. Our data suggest that CD205-expressing Kupffer cells respond to CpG-ODNs and subsequently release IL-12 to promote NKT cell activation. Activated NKT cells induce liver damage through the Fas signaling pathway in HBs-Tg mice.

CD205-TLR9-IL-12 axis contributes to CpG-induced oversensitive liver injury in HBsAg transgenic mice by promoting the interaction of NKT cells with Kupffer cells

PubMed Central

Hou, Xin; Hao, Xiaolei; Zheng, Meijuan; Xu, Congfei; Wang, Jun; Zhou, Rongbin; Tian, Zhigang

2017-01-01

Gut-derived bacterial products contribute to liver inflammation and injury during chronic hepatitis B virus infection; however, the underlying mechanisms remain obscure. In this study, hepatitis B surface antigen transgenic (HBs-Tg) mice and their wild-type (WT) control C57BL/6 mice were injected with CpG-oligodeoxynucleotides (ODNs) to mimic the translocation of gut microbial products into the systemic circulation. We found that, compared with the WT mice, the HBs-Tg mice were oversensitive to CpG-ODN-induced liver injury, which was dependent on natural killer T (NKT) cells. CpG-ODN injection enhanced the expression of Fas ligand (FasL) on NKT cells. In addition, hepatocytes from the HBs-Tg mice expressed higher levels of Fas than did those from the WT mice, which was further augmented by CpG-ODN. Interaction of Fas and FasL was involved in the cytotoxicity of NKT cells against hepatocytes in the HBs-Tg mice. Moreover, Kupffer cells in the HBs-Tg mice expressed higher levels of CD205 and produced greater amounts of interleukin (IL)-12 than did those in the WT mice. Finally, the depletion of Kupffer cells, neutralization of IL-12 or specific silencing of CD205 on Kupffer cells significantly inhibited CpG-ODN-induced liver injury and NKT activation in the HBs-Tg mice. Our data suggest that CD205-expressing Kupffer cells respond to CpG-ODNs and subsequently release IL-12 to promote NKT cell activation. Activated NKT cells induce liver damage through the Fas signaling pathway in HBs-Tg mice. PMID:27041637

Airway Hyperresponsiveness through Synergy of γδ T Cells and NKT Cells1

PubMed Central

Jin, Niyun; Miyahara, Nobuaki; Roark, Christina L.; French, Jena D.; Aydintug, M. Kemal; Matsuda, Jennifer L.; Gapin, Laurent; O'Brien, Rebecca L.; Gelfand, Erwin W.; Born, Willi K.

2015-01-01

Mice sensitized and challenged with OVA were used to investigate the role of innate T cells in the development of allergic airway hyperresponsiveness (AHR). AHR, but not eosinophilic airway inflammation, was induced in T cell-deficient mice by small numbers of cotransferred γδ T cells and invariant NKT cells, whereas either cell type alone was not effective. Only Vγ1+Vδ5+ γδ T cells enhanced AHR. Surprisingly, OVA-specific αβ T cells were not required, revealing a pathway of AHR development mediated entirely by innate T cells. The data suggest that lymphocytic synergism, which is key to the Ag-specific adaptive immune response, is also intrinsic to T cell-dependent innate responses. PMID:17709511

CD1d expression by hepatocytes is a main restriction element for intrahepatic T-cell recognition.

PubMed

Agrati, C; Martini, F; Nisii, C; Oliva, A; D'Offizi, G; Narciso, P; Nardacci, R; Piacentini, M; Dieli, F; Pucillo, L P; Poccia, F

2005-01-01

The liver has specific mechanisms to protect itself from infectious agents and to avoid autoimmunity, indicating an important role of the hepatic tissues in antigen presentation and tolerance induction. Since intrahepatic lymphocytes may contribute to the innate immunity and to the liver pathology, it is of interest to analyze the expression of antigen presenting molecules and of the related T cell recognition in liver, and how these change in relation to different diseases. We analyzed the expression of MHC class I, and of CD1-a, -b, -c, and -d proteins on liver tissues from patients with different hepatic diseases. Moreover, in the same patients we studied the intrahepatic and peripheral NKT cell recognition of alpha-galactosyl ceramide antigen in the context of CD1d. Unlike in other tissues, classical MHC class I molecules were poorly expressed in the hepatic compartment, suggesting that inflamed hepatocytes may trigger weak MHC-restricted T cell responses. Nevertheless, we observed a prevalent expression of HLA class I-like CD1d isoform on the hepatocyte surface, indicating that CD1d is the main restriction element in the liver. In patients with viral hepatitis, the intrahepatic CD1d expression parallels the recruitment of CD56+Valpha24Vbeta11+ NKT cells in the liver which recognize CD1d presenting glycolipids such as alpha-galactosyl ceramide, suggesting that the intrahepatic T cell immunity may focus on glycolipid antigens.

Quantitative and qualitative iNKT repertoire associations with disease susceptibility and outcome in macaque tuberculosis infection.

PubMed

Chancellor, Andrew; White, Andrew; Tocheva, Anna S; Fenn, Joe R; Dennis, Mike; Tezera, Liku; Singhania, Akul; Elliott, Tim; Tebruegge, Marc; Elkington, Paul; Gadola, Stephan; Sharpe, Sally; Mansour, Salah

2017-07-01

Correlates of immune protection that reliably predict vaccine efficacy against Mycobacterium tuberculosis (Mtb) infection are urgently needed. Invariant NKT cells (iNKTs) are CD1d-dependent innate T cells that augment host antimicrobial immunity through production of cytokines, including interferon (IFN)-γ and tumour necrosis factor (TNF)-α. We determined peripheral blood iNKT numbers, their proliferative responses and iNKT subset proportions after in vitro antigen expansion by α-galactosylceramide (αGC) in a large cohort of mycobacteria-naïve non-human primates, and macaques from Bacillus Calmette-Guerin (BCG) vaccine and Mtb challenge studies. Animals studied included four genetically distinct groups of macaques within cynomolgus and rhesus species that differ in their susceptibility to Mtb infection. We demonstrate significant differences in ex vivo iNKT frequency between groups, which trends towards an association with susceptibility to Mtb, but no significant difference in overall iNKT proliferative responses. Susceptible animals exhibited a skewed CD4 + /CD8 + iNKT subset ratio in comparison to more Mtb-resistant groups. Correlation of iNKT subsets post BCG vaccination with clinical disease manifestations following Mtb challenge in the Chinese cynomolgus and Indian rhesus macaques identified a consistent trend linking increased CD8 + iNKTs with favourable disease outcome. Finally, a similar iNKT profile was conferred by BCG vaccination in rhesus macaques. Our study provides the first detailed characterisation of iNKT cells in macaque tuberculosis infection, suggesting that iNKT repertoire differences may impact on disease outcome, which warrants further investigation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Influence of In Vitro IL-2 or IL-15 Alone or in Combination with Hsp 70 Derived 14-Mer Peptide (TKD) on the Expression of NK Cell Activatory and Inhibitory Receptors on Peripheral Blood T Cells, B Cells and NKT Cells.

PubMed

Hromadnikova, Ilona; Li, Shuang; Kotlabova, Katerina; Dickinson, Anne M

2016-01-01

Previous studies from Multhoff and colleagues reported that plasma membrane Hsp70 acts as a tumour-specific recognition structure for activated NK cells, and that the incubation of NK cells with Hsp70 and/or a 14-mer peptide derived from the N-terminal sequence of Hsp70 (TKDNNLLGRFELSG, TKD, aa 450-463) plus a low dose of IL-2 triggers NK cell proliferation and migration, and their capacity to kill cancer cells expressing membrane Hsp70. Herein, we have used flow cytometry to determine the influence of in vitro stimulation of peripheral blood mononuclear cells from healthy individuals with IL-2 or IL-15, either alone or in combination with TKD peptide on the cell surface expression of CD94, NK cell activatory receptors (CD16, NK2D, NKG2C, NKp30, NKp44, NKp46, NKp80, KIR2DL4, DNAM-1 and LAMP1) and NK cell inhibitory receptors (NKG2A, KIR2DL2/L3, LIR1/ILT-2 and NKR-P1A) by CD3+CD56+ (NKT), CD3+CD4+, CD3+CD8+ and CD19+ populations. NKG2D, DNAM-1, LAMP1 and NKR-P1A expression was upregulated after the stimulation with IL-2 or IL-15 alone or in combination with TKD in NKT, CD8+ T cells and B cells. CD94 was upregulated in NKT and CD8+ T cells. Concurrently, an increase in a number of CD8+ T cells expressing LIR1/ILT-2 and CD4+ T cells positive for NKR-P1A was observed. The proportion of CD8+ T cells that expressed NKG2D was higher after IL-2/TKD treatment, when compared with IL-2 treatment alone. In comparison with IL-15 alone, IL-15/TKD treatment increased the proportion of NKT cells that were positive for CD94, LAMP1 and NKRP-1A. The more potent effect of IL-15/TKD on cell surface expression of NKG2D, LIR1/ILT-2 and NKRP-1A was observed in B cells compared with IL-15 alone. However, this increase was not of statistical significance. IL-2/TKD induced significant upregulation of LAMP1 in CD8+ T cells compared with IL-2 alone. Besides NK cells, other immunocompetent cells present within the fraction of peripheral blood mononuclear cells were influenced by the treatment

Unusual Suspects in the Development of Obesity-Induced Inflammation and Insulin Resistance: NK cells, iNKT cells, and ILCs.

PubMed

Bonamichi, Beatriz Dal Santo Francisco; Lee, Jongsoon

2017-08-01

The notion that obesity-induced inflammation mediates the development of insulin resistance in animal models and humans has been gaining strong support. It has also been shown that immune cells in local tissues, in particular in visceral adipose tissue, play a major role in the regulation of obesity-induced inflammation. Specifically, obesity increases the numbers and activation of proinflammatory immune cells, including M1 macrophages, neutrophils, Th1 CD4 T cells, and CD8 T cells, while simultaneously suppressing anti-inflammatory cells such as M2 macrophages, CD4 regulatory T cells, regulatory B cells, and eosinophils. Recently, however, new cell types have been shown to participate in the development of obesity-induced inflammation and insulin resistance. Some of these cell types also appear to regulate obesity. These cells are natural killer (NK) cells and innate lymphoid cells (ILCs), which are closely related, and invariant natural killer T (iNKT) cells. It should be noted that, although iNKT cells resemble NK cells in name, they are actually a completely different cell type in terms of their development and functions in immunity and metabolism. In this review, we will focus on the roles that these relatively new players in the metabolism field play in obesity-induced insulin resistance and the regulation of obesity. Copyright © 2017 Korean Diabetes Association.

Comparative, general pharmacology of SDZ NKT 343, a novel, selective NK1 receptor antagonist

PubMed Central

Walpole, C S J; Brown, M C S; James, I F; Campbell, E A; McIntyre, P; Docherty, R; Ko, S; Hedley, L; Ewan, S; Buchheit, K-H; Urban, L A

1998-01-01

The in vitro and in vivo pharmacology of SDZ NKT 343 (2-nitrophenyl-carbamoyl-(S)-prolyl-(S)-3-(2-naphthyl)alanyl-N-benzyl-N-methylamide), a novel tachykinin NK1 receptor antagonist was investigated.SDZ NKT 343 inhibited [3H]-substance P binding to the human NK1 receptor in transfected Cos-7 cell membranes (IC50=0.62±0.11 nM). In comparison, in the same assay Ki values for FK888, CP 99,994, SR 140,333 and RPR 100,893 were 2.13±0.04 nM, 0.96±0.20 nM, 0.15±0.06 nM and 1.77±0.41 nM, respectively. SDZ NKT 343 showed a markedly lower affinity at rat NK1 receptors in whole forebrain membranes (IC50=451±139 nM).SDZ NKT 343 caused an increase in EC50 as well as reduction in the number of binding sites (Bmax) determined for [3H]-substance P, suggesting a non-competitive interaction at the human NK1 receptor. SDZ NKT 343 also caused a reduction in the maximum elevation of [Ca2+]i evoked by substance P (SP) in human U373MG cells and depressed the maximum [Sar9]SP sulphone-induced contraction of the guinea-pig isolated ileum. The antagonism of SP effects on U373MG cells by SDZ NKT 343 was reversible.SDZ NKT 343 showed weak affinity to human NK2 and NK3 receptors in transfected Cos-7 cells (Ki of 0.52±0.04 μM and 3.4±1.2 μM, respectively). SDZ NKT 343 was inactive in a broad array of binding assays including the bradykinin B2 receptor the histamine H1 receptor, opiate receptors and adrenoceptors. SDZ NKT 343 only weakly inhibited the voltage-activated Ca2+ and Na+currents in guinea-pig dorsal root ganglion neurones. The enantiomer of SDZ NKT 343, (R,R)-SDZ NKT 343 was about 1000 times less active at human NK1 receptors expressed in Cos-7 cell membranes.Contractions of the guinea-pig ileum by [Sar9]SP sulphone were inhibited by SDZ NKT 343 in a concentration-dependent manner, with an IC50=1.60±0.94 nM, while the enantiomer (R,R)-SDZ NKT 343 was 100 times less active (IC50=162±26 nM). In comparison, in the same assay IC50 values for other NK1

CD1d-dependent expansion of NKT follicular helper cells in vivo and in vitro is a product of cellular proliferation and differentiation.

PubMed

Rampuria, Pragya; Lang, Mark L

2015-05-01

NKT follicular helper cells (NKTfh cells) are a recently discovered functional subset of CD1d-restricted NKT cells. Given the potential for NKTfh cells to promote specific antibody responses and germinal center reactions, there is much interest in determining the conditions under which NKTfh cells proliferate and/or differentiate in vivo and in vitro. We confirm that NKTfh cells expressing the canonical semi-invariant Vα14 TCR were CXCR5(+)/ICOS(+)/PD-1(+)/Bcl6(+) and increased in number following administration of the CD1d-binding glycolipid α-galactosylceramide (α-GC) to C57Bl/6 mice. We show that the α-GC-stimulated increase in NKTfh cells was CD1d-dependent since the effect was diminished by reduced CD1d expression. In vivo and in vitro treatment with α-GC, singly or in combination with IL-2, showed that NKTfh cells increased in number to a greater extent than total NKT cells, but proliferation was near-identical in both populations. Acquisition of the NKTfh phenotype from an adoptively transferred PD-1-depleted cell population was also evident, showing that peripheral NKT cells differentiated into NKTfh cells. Therefore, the α-GC-stimulated, CD1d-dependent increase in peripheral NKTfh cells is a result of cellular proliferation and differentiation. These findings advance our understanding of the immune response following immunization with CD1d-binding glycolipids. © The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Involvement of the iNKT Cell Pathway Is Associated With Early-Onset Eosinophilic Esophagitis and Response to Allergen Avoidance Therapy

PubMed Central

Lexmond, Willem S.; Neves, Joana F.; Nurko, Samuel; Olszak, Torsten; Exley, Mark A.; Blumberg, Richard S.; Fiebiger, Edda

2014-01-01

OBJECTIVES Recent experimental evidence suggests that environmental microbial factors early in life determine susceptibility to allergic diseases through inappropriate chemotaxis and local activation of CD1d-restricted, invariant chain natural killer T (iNKT) cells. In this study, we analyzed the involvement of these pathways in pediatric patients with eosinophilic esophagitis (EoE) before and after dietary allergen elimination. METHODS mRNA expression levels of components of the C-X-C motif chemokine ligand 16 (CXCL16)–iNKT–CD1d axis were compared in esophageal biopsies from EoE patients vs. normal or inflammatory controls and before and after treatment. RESULTS CXCL16, iNKT cell–associated cell marker Vα24, and CD1d were significantly upregulated in esophageal biopsies from EoE patients and correlated with the expression of inflammatory mediators associated with allergy. Upregulation of each of these factors was significantly more pronounced in patients aged < 6 years at diagnosis, and this early-onset EoE subpopulation was characterized by a more prominent food allergic disease phenotype in a cohort-wide analysis. Successful, but not unsuccessful, treatment of early-onset EoE patients with dietary elimination of instigating allergens led to reduction in infiltrating iNKT cells and complete normalization of mRNA expression levels of CXCL16 and CD1d. CONCLUSIONS Our observations place iNKT cells at the center of allergic inflammation associated with EoE, which could have profound implications for our understanding, treatment and prevention of this and other human allergic diseases. PMID:24513807

Adjuvant effects of invariant NKT cell ligand potentiates the innate and adaptive immunity to an inactivated H1N1 swine influenza virus vaccine in pigs.

PubMed

Dwivedi, Varun; Manickam, Cordelia; Dhakal, Santosh; Binjawadagi, Basavaraj; Ouyang, Kang; Hiremath, Jagadish; Khatri, Mahesh; Hague, Jacquelyn Gervay; Lee, Chang Won; Renukaradhya, Gourapura J

2016-04-15

Pigs are considered as the source of some of the emerging human flu viruses. Inactivated swine influenza virus (SwIV) vaccine has been in use in the US swine herds, but it failed to control the flu outbreaks. The main reason has been attributed to lack of induction of strong local mucosal immunity in the respiratory tract. Invariant natural killer T (iNKT) cell is a unique T cell subset, and activation of iNKT cell using its ligand α-Galactosylceramide (α-GalCer) has been shown to potentiate the cross-protective immunity to inactivated influenza virus vaccine candidates in mice. Recently, we discovered iNKT cell in pig and demonstrated its activation using α-GalCer. In this study, we evaluated the efficacy of an inactivated H1N1 SwIV coadministered with α-GalCer intranasally against a homologous viral challenge. Our results demonstrated the potent adjuvant effects of α-GalCer in potentiating both innate and adaptive immune responses to SwIV Ags in the lungs of pigs, which resulted in reduction in the lung viral load by 3 logs compared to without adjuvant. Immunologically, in the lungs of pigs vaccinated with α-GalCer an increased virus specific IgA response, IFN-α secretion and NK cell-cytotoxicity was observed. In addition, iNKT cell-stimulation enhanced the secretion of Th1 cytokines (IFN-γ and IL-12) and reduced the production of immunosuppressive cytokines (IL-10 and TGF-β) in the lungs of pigs⋅ In conclusion, we demonstrated for the first time iNKT cell adjuvant effects in pigs to SwIV Ags through augmenting the innate and adaptive immune responses in the respiratory tract. Copyright © 2016 Elsevier B.V. All rights reserved.

Functional CD1d and/or NKT cell invariant chain transcript in horse, pig, African elephant and guinea pig, but not in ruminants.

PubMed

Looringh van Beeck, Frank A; Reinink, Peter; Hermsen, Roel; Zajonc, Dirk M; Laven, Marielle J; Fun, Axel; Troskie, Milana; Schoemaker, Nico J; Morar, Darshana; Lenstra, Johannes A; Vervelde, Lonneke; Rutten, Victor P M G; van Eden, Willem; Van Rhijn, Ildiko

2009-04-01

CD1d-restricted invariant natural killer T cells (NKT cells) have been well characterized in humans and mice, but it is unknown whether they are present in other species. Here we describe the invariant TCR alpha chain and the full length CD1d transcript of pig and horse. Molecular modeling predicts that porcine (po) invariant TCR alpha chain/poCD1d/alpha-GalCer and equine (eq) invariant TCR alpha chain/eqCD1d/alpha-GalCer form complexes that are highly homologous to the human complex. Since a prerequisite for the presence of NKT cells is the expression of CD1d protein, we performed searches for CD1D genes and CD1d transcripts in multiple species. Previously, cattle and guinea pig have been suggested to lack CD1D genes. The CD1D genes of European taurine cattle (Bos taurus) are known to be pseudogenes because of disrupting mutations in the start codon and in the donor splice site of the first intron. Here we show that the same mutations are found in six other ruminants: African buffalo, sheep, bushbuck, bongo, N'Dama cattle, and roe deer. In contrast, intact CD1d transcripts were found in guinea pig, African elephant, horse, rabbit, and pig. Despite the discovery of a highly homologous NKT/CD1d system in pig and horse, our data suggest that functional CD1D and CD1d-restricted NKT cells are not universally present in mammals.

CD1d-restricted immunoglobulin G formation to GPI-anchored antigens mediated by NKT cells.

PubMed

Schofield, L; McConville, M J; Hansen, D; Campbell, A S; Fraser-Reid, B; Grusby, M J; Tachado, S D

1999-01-08

Immunoglobulin G (IgG) responses require major histocompatibility complex (MHC)-restricted recognition of peptide fragments by conventional CD4(+) helper T cells. Immunoglobulin G responses to glycosylphosphatidylinositol (GPI)- anchored protein antigens, however, were found to be regulated in part through CD1d-restricted recognition of the GPI moiety by thymus-dependent, interleukin-4-producing CD4(+), natural killer cell antigen 1.1 [(NK1.1)+] helper T cells. The CD1-NKT cell pathway regulated immunogobulin G responses to the GPI-anchored surface antigens of Plasmodium and Trypanosoma and may be a general mechanism for rapid, MHC-unrestricted antibody responses to diverse pathogens.

[Treatment outcome and prognosis of autologous hematopoietic stem cell transplantation combined with high dose radiotherapy/chemotherapy in 22 patients with nasal NK/T cell lymphoma].

PubMed

Cui, Xiu-Zhen; Wang, Hua-Qing; Liu, Xian-Ming; Zhang, Hui-Lai; Li, Wei

2007-09-01

To analyze the outcome and prognosis of autologous hematopoietic stem cell transplantation (AHSCT) combined with high dose radiotherapy/chemotherapy in 22 patients with nasal NK/T cell lymphoma. From July 1992 to December 2005, 22 patients with nasal NK/T cell lymphoma were diagnosed pathologically. Immunophenotyping was performed in 13 cases. The patients were classified by Ann Arbor staging system and international prognosis index (IPI). The patients received cycles of chemotherapy every other two weeks or combined with radiotherapy for remission induction, followed high dose radiotherapy/chemotherapy, combined with autologous peripheral blood stem cell transplantation (APBSCT), or autologous bone-marrow transplantation (ABMT). Patients were given complementary radiotherapy after transplantation if they did not have it before. Twelve patients of IPI 3 -4 received consolidation chemotherapy, and one of them received the second transplantation. The median follow-up duration was 64 (12 - 168) months. The 5 and 8-year overall survivals (OS) were 79.3% and 64.1%, and disease free survivals (DFS) were 36.4% and 27.3%, respectively. The 5-year OS were as follows: for stage I - II and III - IV disease were 90.0% and 70.0% (P = 0. 041); for patients without and with B symptom were 100.0% and 70.7% (P = 0.045); and for IPI 1 - 2 and 3 - 4 were 100.0% and 60.0% (P = 0.035), respectively. Multivariate analysis by COX regression revealed that disease stage, B symptom and IPI were independent prognostic factors. AHSCT combined with high dose radiotherapy/chemotherapy is an effective treatment for patients with poor prognosis nasal NK/T cell lymphoma.

Synthesis of truncated analogues of the iNKT cell agonist, α-galactosyl ceramide (KRN7000), and their biological evaluation

PubMed Central

Veerapen, Natacha; Reddington, Faye; Salio, Mariolina; Cerundolo, Vincenzo; Besra, Gurdyal S.

2011-01-01

Stimulation of iNKT cells by α-galactosyl ceramide (α-GalCer), also known as KRN7000, and its truncated analogue OCH induces both Th1- and Th2-cytokines, with OCH inducing a Th2-cytokine bias. Skewing of the iNKT cells’ response towards either a Th1- or Th2-cytokine profile offers potential therapeutic benefits. The length of both the acyl and the sphingosine chains in α-galactosyl ceramides is known to influence the cytokine release profile. We have synthesized analogues of α-GalCer with truncated sphingosine chains for biological evaluation, with particular emphasis on the Th1/Th2 distribution. Starting from a common precursor, d-lyxose, the sphingosine derivatives were synthesised via a straightforward Wittig condensation. PMID:21145749

ID2 collaborates with ID3 to suppress iNKT and innate-like tumors1

PubMed Central

Li, Jia; Roy, Sumedha; Kim, Young-Mi; Li, Shibo; Zhang, Baojun; Love, Cassandra; Reddy, Anupama; Rajagopalan, Deepthi; Dave, Sandeep; Diehl, Anna Mae; Zhuang, Yuan

2017-01-01

Inhibitor of DNA binding (ID) proteins, including ID1-4, are transcriptional regulators involved in promoting cell proliferation and survival in various cell types. Although upregulation of Id proteins has been associated with a broad spectrum of tumors, recent studies have identified that ID3 plays a tumor suppressor role in the development of Burkitt’s lymphoma in humans and Hepatosplenic T cell lymphomas in mice. Here, we report rapid lymphoma development in Id2/Id3 double knockout (L-DKO) mice caused by unchecked expansion of either invariant Natural Killer T (iNKT) cells, or a unique subset of innate-like, CD1d-independent T cells. These populations started expansion in neonatal mice and, upon malignant transformation, caused fatality at age between 3–11 months. The malignant cells also gave rise to lymphomas upon transfer to Rag-deficient and wild-type hosts, reaffirming their inherent tumorigenic potential. Microarray analysis revealed a significantly modified program in these neonatal iNKT cells that ultimately led to their malignant transformation. The lymphoma cells demonstrated chromosome instability, along with upregulation of several different signaling pathways, including the cytokine-cytokine receptor interaction pathway, which can promote their expansion and migration. Dysregulation of genes with reported driver mutations and the NF-kB pathway were found to be shared between L-DKO lymphomas and human NKT tumors. Our work identifies a distinct premalignant state and multiple tumoriogenic pathways caused by loss function of ID2 and ID3. Thus, conditional deletion of Id2 and Id3 in developing T cells establishes a unique animal model for iNKT and relevant innate-like lymphomas. PMID:28258199

Steroid Resistant CD8+CD28null NKT-Like Pro-inflammatory Cytotoxic Cells in Chronic Obstructive Pulmonary Disease.

PubMed

Hodge, Greg; Hodge, Sandra

2016-01-01

Corticosteroid resistance is a major barrier to effective treatment in chronic obstructive pulmonary disease (COPD), and failure to suppress systemic inflammation in these patients may result in increased comorbidity. Although much of the research to date has focused on the role of macrophages and neutrophils involved in inflammation in the airways in COPD, recent evidence suggests that CD8 + T cells may be central regulators of the inflammatory network in this disease. CD8 + cytotoxic pro-inflammatory T cells have been shown to be increased in the peripheral blood and airways in patients with COPD, whereas smokers that have not progressed to COPD only show an increase in the lungs. Although the mechanisms underlying steroid resistance in these lymphocytes is largely unknown, new research has identified a role for cytotoxic pro-inflammatory CD8 + T-cells and CD8 + natural killer T-like (NKT-like) cells. Increased numbers of these cells and their significant loss of the co-stimulatory molecule CD28 have been shown in COPD, consistent with findings in the elderly and in clinical conditions involving chronic activation of the immune system. In COPD, these senescent cells expressed increased levels of the cytotoxic mediators, perforin and granzyme b, and the pro-inflammatory cytokines, IFNγ and TNFα. They also demonstrated increased cytotoxicity toward lung epithelial cells and importantly were resistant to immunosuppression by corticosteroids compared with their CD28 + counterparts. Further research has shown these cells evade the immunosuppressive effects of steroids via multiple mechanisms. This mini review will focus on cytotoxic pro-inflammatory CD8 + CD28 null NKT-like cells involved in COPD and novel approaches to reverse steroid resistance in these cells.

Steroid Resistant CD8+CD28null NKT-Like Pro-inflammatory Cytotoxic Cells in Chronic Obstructive Pulmonary Disease

PubMed Central

Hodge, Greg; Hodge, Sandra

2016-01-01

Corticosteroid resistance is a major barrier to effective treatment in chronic obstructive pulmonary disease (COPD), and failure to suppress systemic inflammation in these patients may result in increased comorbidity. Although much of the research to date has focused on the role of macrophages and neutrophils involved in inflammation in the airways in COPD, recent evidence suggests that CD8+ T cells may be central regulators of the inflammatory network in this disease. CD8+ cytotoxic pro-inflammatory T cells have been shown to be increased in the peripheral blood and airways in patients with COPD, whereas smokers that have not progressed to COPD only show an increase in the lungs. Although the mechanisms underlying steroid resistance in these lymphocytes is largely unknown, new research has identified a role for cytotoxic pro-inflammatory CD8+ T-cells and CD8+ natural killer T-like (NKT-like) cells. Increased numbers of these cells and their significant loss of the co-stimulatory molecule CD28 have been shown in COPD, consistent with findings in the elderly and in clinical conditions involving chronic activation of the immune system. In COPD, these senescent cells expressed increased levels of the cytotoxic mediators, perforin and granzyme b, and the pro-inflammatory cytokines, IFNγ and TNFα. They also demonstrated increased cytotoxicity toward lung epithelial cells and importantly were resistant to immunosuppression by corticosteroids compared with their CD28+ counterparts. Further research has shown these cells evade the immunosuppressive effects of steroids via multiple mechanisms. This mini review will focus on cytotoxic pro-inflammatory CD8+CD28null NKT-like cells involved in COPD and novel approaches to reverse steroid resistance in these cells. PMID:28066427

Natural Killer T Cells in Cancer Immunotherapy

PubMed Central

Nair, Shiny; Dhodapkar, Madhav V.

2017-01-01

Natural killer T (NKT) cells are specialized CD1d-restricted T cells that recognize lipid antigens. Following stimulation, NKT cells lead to downstream activation of both innate and adaptive immune cells in the tumor microenvironment. This has impelled the development of NKT cell-targeted immunotherapies for treating cancer. In this review, we provide a brief overview of the stimulatory and regulatory functions of NKT cells in tumor immunity as well as highlight preclinical and clinical studies based on NKT cells. Finally, we discuss future perspectives to better harness the potential of NKT cells for cancer therapy. PMID:29018445

IL30 (IL27p28) attenuates liver fibrosis through inducing NKG2D-Rae1 interaction between NKT and activated hepatic stellate cells

PubMed Central

Mitra, Abhisek; Satelli, Arun; Yan, Jun; Xueqing, Xia; Gagea, Mihai; Hunter, Christopher A.; Mishra, Lopa; Li, Shulin

2014-01-01

Chronic hepatic diseases such as cirrhosis, hepatocellular carcinoma and virus mediated immunopathogenic infections are affecting billions of people worldwide. These diseases commonly initiate with fibrosis. Owing to the various side effects of anti-fibrotic therapy and the difficulty of diagnosing asymptomatic patients, suitable medication remains a major concern. To overcome this drawback, the use of cytokine-based sustained therapy might be a suitable alternative with minimal side effects. Here, we studied the therapeutic efficacy and potential mechanisms of IL30 as anti-fibrosis therapy in murine liver fibrosis models. Carbon tetrachloride (CCl4) mixed with corn oil at a ratio 1:3 was injected intraperitoneally (IP) 1µl/gm body weight twice per week for 1 month or 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) 0.1% (wt/wt) Purnima 5015 Chow was fed for 3 weeks to induce liver fibrosis. Either control vector (pCtr) or pIL30 was injected hydrodynamically once per week. A significant decrease in collagen deposition and reduced expression of α-smooth muscle Actin (αSMA) protein indicated that IL30–based gene therapy dramatically reduced bridging fibrosis that was induced by CCl4 or DDC. Immunophenotyping and knockout studies showed that IL30 recruits NKT cells to the liver to decrease activated hepatic stellate cells (HSCs) significantly and ameliorate liver fibrosis. Both flow cytometric and antibody mediated neutralization studies showed NKT cells alleviate liver fibrosis in an NKG2D dependent manner. Furthermore, chronic treatment with CCl4 showed inducible surface expression of the NKG2D ligand Rae1 on activated HSCs as compared to quiescent ones. Taken together, our results show that highly target specific liver NKT cells selectively remove activated HSCs via an NKG2D-Rae1 interaction to ameliorate liver fibrosis after IL30 treatment. PMID:25351459

Inhibition of T Helper Cell Type 2 Cell Differentiation and Immunoglobulin E Response by Ligand-Activated Vα14 Natural Killer T Cells

PubMed Central

Cui, Junqing; Watanabe, Naohiro; Kawano, Tetsu; Yamashita, Masakatsu; Kamata, Tohru; Shimizu, Chiori; Kimura, Motoko; Shimizu, Eiko; Koike, Jyunzo; Koseki, Haruhiko; Tanaka, Yujiro; Taniguchi, Masaru; Nakayama, Toshinori

1999-01-01

Murine Vα14 natural killer T (NKT) cells are thought to play a crucial role in various immune responses, including infectious, allergic, and autoimmune diseases. Because Vα14 NKT cells produce large amounts of both interleukin (IL)-4 and interferon (IFN)-γ upon in vivo stimulation with a specific ligand, α-galactosylceramide (α-GalCer), or after treatment with anti-CD3 antibody, a regulatory role on helper T (Th) cell differentiation has been proposed for these cells. However, the identity of the cytokine produced by Vα14 NKT cells that play a dominant role on the Th cell differentiation still remains controversial. Here, we demonstrate by using Vα14 NKT-deficient mice that Vα14 NKT cells are dispensable for the induction of antigen-specific immunoglobulin (Ig)E responses induced by ovalbumin immunization or Nippostrongylus brasiliensis infection. However, upon in vivo activation with α-GalCer, Vα14 NKT cells are found to suppress antigen-specific IgE production. The suppression appeared to be IgE specific, and was not detected in either Vα14 NKT– or IFN-γ–deficient mice. Consistent with these results, we also found that ligand-activated Vα14 NKT cells inhibited Th2 cell differentiation in an in vitro induction culture system. Thus, it is likely that activated Vα14 NKT cells exert a potent inhibitory effect on Th2 cell differentiation and subsequent IgE production by producing a large amount of IFN-γ. In marked contrast, our studies have revealed that IL-4 produced by Vα14 NKT cells has only a minor effect on Th2 cell differentiation. PMID:10499917

CXCL16-positive dendritic cells enhance invariant natural killer T cell-dependent IFNγ production and tumor control

PubMed Central

Veinotte, Linnea; Gebremeskel, Simon; Johnston, Brent

2016-01-01

ABSTRACT Crosstalk interactions between dendritic cells (DCs) and invariant natural killer T (iNKT) cells are important in regulating antitumor responses elicited by glycolipid antigens. iNKT cells constitutively express the chemokine receptor CXCR6, while cytokine-activated DCs upregulate the transmembrane chemokine ligand, CXCL16. This study examined the co-stimulatory role of CXCR6/CXCL16 interactions in glycolipid-dependent iNKT cell activation and tumor control. Spleen and liver DCs in wild-type mice, but not iNKT cell deficient (Jα18−/−) mice, transiently upregulated surface CXCL16 following in vivo administration of the glycolipid antigen α-galactosylceramide. Recombinant CXCL16 did not directly induce iNKT cell activation in vitro but enhanced interferon (IFN)-γ production when mouse or human iNKT cells were stimulated with plate-bound anti-CD3. Compared with glycolipid-loaded CXCL16neg DCs, CXCL16hi DCs induced higher levels of IFNγ production in iNKT cell cultures and following adoptive transfer in vivo. The number of IFNγ+ iNKT cells and expansion of T-bet+ iNKT cells were reduced in vivo when CXCL16−/− DCs were used to activate iNKT cells. Enhanced IFNγ production in vivo was not dependent on CXCR6 expression on natural killer (NK) cells. Adoptive transfer of glycolipid-loaded CXCL16hi DCs provided superior protection against tumor metastasis compared to CXCL16neg DC transfers. Similarly, wild-type DCs provided superior protection against metastasis compared with CXCL16−/− DCs. These experiments implicate an important role for CXCR6/CXCL16 interactions in regulating iNKT cell IFNγ production and tumor control. The selective use of CXCL16hi DCs in adoptive transfer immunotherapies may prove useful for enhancing T helper (Th) type 1 responses and clinical outcomes in cancer patients. PMID:27471636

Regulatory T cells decrease invariant natural killer T cell-mediated pregnancy loss in mice.

PubMed

Li, L; Tu, J; Jiang, Y; Zhou, J; Schust, D J

2017-05-01

Pregnancy loss is the commonest complication of pregnancy. The causes of pregnancy loss are poorly understood. It has been reported that stimulation of invariant natural killer T (iNKT) cells using α-galactosylceramide (αGC) induces pregnancy loss in mice. Here we investigated the mechanisms, especially the role of regulatory T (Treg) cells, in iNKT cell-mediated pregnancy loss. We found that injection of αGC rapidly induced fetal resorption, activated decidual iNKT cells, decreased the percentage of decidual Treg cells and their interleukin (IL)-10 and transforming growth factor (TGF)-β production, and upregulated the levels of interferon (IFN)-γ, tumor necrosis factor-α, IL-4, and IL-10 in serum. Adoptive transfer of iNKT cells from wild-type (WT) and IL-4 -/- mice but not IFN-γ -/- mice into αGC-treated iNKT cell-deficient Jα18 -/- mice restored αGC-induced pregnancy loss. Adoptive transfer of Treg cells downregulated α-GC-induced pregnancy loss in WT mice. Finally, co-culture with αGC-stimulated decidual iNKT cells decreased the production of IL-10 and TGF-β in decidual Treg cells and inhibited their suppressive activity. These findings suggest that activation of iNKT cells induces pregnancy loss in mice in an IFN-γ-dependent manner. In addition, inhibition of the function of decidual Treg cells has an important role in iNKT cell-mediated pregnancy loss.

Immunotherapeutic strategies targeting Natural killer T cell responses in cancer

PubMed Central

Shissler, Susannah C.; Bollino, Dominique R.; Tiper, Irina V.; Bates, Joshua; Derakhshandeh, Roshanak; Webb, Tonya J.

2017-01-01

Natural killer T (NKT) cells are a unique subset of lymphocytes that bridge the innate and adaptive immune system. NKT cells possess a classic αβ T-cell receptor (TCR) that is able to recognize self and foreign glycolipid antigens presented by the nonclassical class I major histocompatibility complex (MHC) molecule, CD1d. Type I NKT cells (referred to as invariant NKT cells) express a semi-invariant Vα14Jα18 TCR in mice and Vα24Jα18 TCR in humans. Type II NKT cells are CD1d-restricted T cells that express a more diverse set of TCR α chains. The two types of NKT cells often exert opposing effects especially in tumor immunity, where Type II cells generally suppress tumor immunity while Type I NKT cells can enhance antitumor immune responses. In this review, we focus on the role of NKT cells in cancer. We discuss their effector and suppressive functions, as well as describe preclinical and clinical studies utilizing therapeutic strategies focused on harnessing their potent anti-tumor effector functions, and conclude with a discussion on potential next steps for the utilization of NKT cell targeted therapies for the treatment of cancer. PMID:27393665

Circulating natural killer T cells in patients with asthma.

PubMed

Ikegami, Yasuhiko; Yokoyama, Akihito; Haruta, Yoshinori; Hiyama, Keiko; Kohno, Nobuoki

2004-01-01

Recent studies suggest that therapies targeted at depletion or limiting of natural killer (NK) T cells may be a possible strategy for the treatment of asthma. In the present study, we measured the number of circulating V alpha24+ NKT cells in 32 asthmatic patients and compared these patients with 29 nonatopic healthy controls. We investigated the relationships between NKT cell number and clinical variables such as the number of eosinophils, the circulating level of IgE, and the severity of asthma. In addition, we also investigated the ability of NKT cells to proliferate in response to alpha-galactosyl ceramide (alpha-GalCer) in vitro. The V alpha24+ NKT cell counts of asthmatic patients were significantly lower than those of healthy controls. There were no significant differences observed in asthmatic patients among the subgroups in terms of atopic status and severity. There was no significant correlation between the number of NKT cells and clinical variables. The proliferative response to alpha-GalCer of the patients and controls was not significantly different, indicating no intrinsic proliferative defect of NKT cells in asthma. These results suggest that the number of circulating NKT cells was already decreased in patients with asthma. Further study, such as the evaluation of lung NKT cells, will be needed to determine the role of NKT cells in patients with asthma.

Dendritic cells rapidly undergo apoptosis in vitro following culture with activated CD4+ Vα24 natural killer T cells expressing CD40L

PubMed Central

Nieda, M; Kikuchi, A; Nicol, A; Koezuka, Y; Ando, Y; Ishihara, S; Lapteva, N; Yabe, T; Tokunaga, K; Tadokoro, K; Juji, T

2001-01-01

Human Vα24 natural killer T (Vα24NKT) cells are activated by α-glycosylceramide-pulsed dendritic cells (DCs) in a CD1d-dependent and T-cell receptor-mediated manner. There are two major subpopulations of Vα24NKT cells, CD4– CD8– Vα24NKT and CD4+ Vα24NKT cells. We have recently shown that activated CD4– CD8– Vα24NKT cells have cytotoxic activity against DCs, but knowledge of the molecules responsible for cytotoxicity of Vα24NKT cells is currently limited. We aimed to investigate whether CD4+ Vα24NKT cells also have cytotoxic activity against DCs and to determine the mechanisms underlying any observed cytotoxic activity. We demonstrated that activated CD4+ Vα24NKT cells [CD40 ligand (CD40L) -positive] have cytotoxic activity against DCs (strongly CD40-positive), but not against monocytes (weakly CD40-positive) or phytohaemagglutinin blast T cells (CD40-negative), and that apoptosis of DCs significantly contributes to the observed cytotoxicity. The apoptosis of DCs following culture with activated CD4+ Vα24NKT cells, but not with resting CD4+ Vα24NKT cells (CD40L-negative), was partially inhibited by anti-CD40L mAb. Direct ligation of CD40 on the DCs by the anti-CD40 antibody also induced apoptosis of DCs. Our results suggest that CD40–CD40L interaction plays an important role in the induction of apoptosis of DCs following culture with activated CD4+ Vα24NKT cells. The apoptosis of DCs from normal donors, triggered by the CD40–CD40L interaction, may contribute to the homeostatic regulation of the normal human immune system, preventing the interminable activation of activated CD4+ Vα24NKT cells by virtue of apoptosis of DCs. PMID:11260318

Natural killer T cells in health and disease

PubMed Central

Wu, Lan; Van Kaer, Luc

2013-01-01

Natural killer T (NKT) cells are a subset of T lymphocytes that share surface markers and functional characteristics with both conventional T lymphocytes and natural killer cells. Most NKT cells express a semiinvariant T cell receptor that reacts with glycolipid antigens presented by the major histocompatibility complex class I-related protein CD1d on the surface of antigen-presenting cells. NKT cells become activated during a variety of infections and inflammatory conditions, rapidly producing large amounts of immunomodulatory cytokines. NKT cells can influence the activation state and functional properties of multiple other cell types in the immune system and, thus, modulate immune responses against infectious agents, autoantigens, tumors, tissue grafts and allergens. One attractive aspect of NKT cells is that their immunomodulatory activities can be readily harnessed with cognate glycolipid antigens, such as the marine sponge-derived glycosphingolipid alpha-galactosylceramide. These properties of NKT cells are being exploited for therapeutic intervention to prevent or treat cancer, infections, and autoimmune and inflammatory diseases. PMID:21196373

Lipid-Antigen Presentation by CD1d+ B Cells Is Essential for the Maintenance of Invariant Natural Killer T Cells

PubMed Central

Bosma, Anneleen; Abdel-Gadir, Azza; Isenberg, David A.; Jury, Elizabeth C.; Mauri, Claudia

2012-01-01

Summary B cells perform many immunological functions, including presenting lipid antigen to CD1d-restricted invariant natural killer T (iNKT) cells, known to contribute to maintaining tolerance in autoimmunity. Patients with systemic lupus erythematous (SLE) display dysregulated B cell responses and reduced peripheral iNKT cell frequencies. The significance of these defects and how they relate to SLE pathogenesis remain elusive. We report that B cells are essential for iNKT cell expansion and activation in healthy donors but fail to exert a similar effect in SLE patients. Defective B cell-mediated stimulation of iNKT cells in SLE patients was associated with altered CD1d recycling, a defect recapitulated in B cells from healthy donors after stimulation with interferon-α (IFN-α) and anti-immunoglobulin (Ig). iNKT cell number and function were restored in SLE patients responding to anti-CD20 treatment upon normalization of CD1d expression exclusively in repopulated immature B cells. We propose that healthy B cells are pivotal for iNKT cell homeostasis. PMID:22406267

Immature Renal Dendritic Cells Recruit Regulatory CXCR6+ Invariant Natural Killer T Cells to Attenuate Crescentic GN

PubMed Central

Riedel, Jan-Hendrik; Paust, Hans-Joachim; Turner, Jan-Eric; Tittel, André P.; Krebs, Christian; Disteldorf, Erik; Wegscheid, Claudia; Tiegs, Gisa; Velden, Joachim; Mittrücker, Hans-Willi; Garbi, Natalio; Stahl, Rolf A.K.; Steinmetz, Oliver M.; Kurts, Christian

2012-01-01

Immature renal dendritic cells (DCs) are protective early in murine crescentic GN, but the mechanisms underlying this protection are unknown. Here, depletion of DCs reduced the recruitment of invariant natural killer T (iNKT) cells, which attenuate GN, into the kidney in the early stage of experimental crescentic GN. More than 90% of renal iNKT cells expressed the chemokine receptor CXCR6, and renal DCs produced high amounts of the cognate ligand CXCL16 early after induction of nephritis, suggesting that renal DC-derived CXCL16 might attract protective CXCR6+ iNKT cells. Consistent with this finding, CXCR6-deficient mice exhibited less iNKT cell recruitment and developed nephritis that was more severe, similar to the aggravated nephritis observed in mice depleted of immature DCs. Finally, adoptive transfer of CXCR6-competent NKT cells ameliorated nephritis. Taken together, these results suggest an immunoprotective mechanism involving immature DCs, CXCL16, CXCR6, and regulatory iNKT cells, which might stimulate the development of new therapeutic strategies for GN. PMID:23138484

Immature renal dendritic cells recruit regulatory CXCR6(+) invariant natural killer T cells to attenuate crescentic GN.

PubMed

Riedel, Jan-Hendrik; Paust, Hans-Joachim; Turner, Jan-Eric; Tittel, André P; Krebs, Christian; Disteldorf, Erik; Wegscheid, Claudia; Tiegs, Gisa; Velden, Joachim; Mittrücker, Hans-Willi; Garbi, Natalio; Stahl, Rolf A K; Steinmetz, Oliver M; Kurts, Christian; Panzer, Ulf

2012-12-01

Immature renal dendritic cells (DCs) are protective early in murine crescentic GN, but the mechanisms underlying this protection are unknown. Here, depletion of DCs reduced the recruitment of invariant natural killer T (iNKT) cells, which attenuate GN, into the kidney in the early stage of experimental crescentic GN. More than 90% of renal iNKT cells expressed the chemokine receptor CXCR6, and renal DCs produced high amounts of the cognate ligand CXCL16 early after induction of nephritis, suggesting that renal DC-derived CXCL16 might attract protective CXCR6(+) iNKT cells. Consistent with this finding, CXCR6-deficient mice exhibited less iNKT cell recruitment and developed nephritis that was more severe, similar to the aggravated nephritis observed in mice depleted of immature DCs. Finally, adoptive transfer of CXCR6-competent NKT cells ameliorated nephritis. Taken together, these results suggest an immunoprotective mechanism involving immature DCs, CXCL16, CXCR6, and regulatory iNKT cells, which might stimulate the development of new therapeutic strategies for GN.

Sequence analysis of Epstein-Barr virus (EBV) early genes BARF1 and BHRF1 in NK/T cell lymphoma from Northern China.

PubMed

Sun, Lingling; Che, Kui; Zhao, Zhenzhen; Liu, Song; Xing, Xiaoming; Luo, Bing

2015-09-04

NK/T cell lymphoma is an aggressive lymphoma almost always associated with EBV. BamHI-A rightward open reading frame 1 (BARF1) and BamHI-H rightward open reading frame 1 (BHRF1) are two EBV early genes, which may be involved in the oncogenicity of EBV. It has been found that V29A strains, a BARF1 mutant subtype, showed higher prevalence in NPC, which may suggest the association between this variation and nasopharyngeal carcinoma (NPC). To characterize the sequence variation patterns of the Epstein-Barr virus (EBV) early genes and to elucidate their association with NK/T cell lymphoma, we analyzed the sequences of BARF1 and BHRF1 in EBV-positive NK/T cell lymphoma samples from Northern China. In situ hybridization (ISH) performed for EBV-encoded small RNA1 (EBER1) with specific digoxigenin-labeled probes was used to select the EBV positive lymphoma samples. Nested-polymerase chain reaction (nested-PCR) and DNA sequence analysis technique were used to obtain the sequences of BARF1 and BHRF1. The polymorphisms of these two genes were classified according to the signature changes and compared with the known corresponding EBV gene variation data. Two major subtypes of BARF1 gene, designated as B95-8 and V29A subtype, were identified. B95-8 subtype was the dominant subtype. The V29A subtype had one consistent amino acid change at amino acid residue 29 (V → A). Compared with B95-8, AA change at 88 (L → V) of BHRF1 was found in the majority of the isolates, and AA79 (V → L) mutation in a few isolates. Functional domains of BARF1 and BHRF1 were highly conserved. The distributions of BARF1 and BHRF1 subtypes had no significant differences among different EBV-associated malignancies and healthy donors. The sequences of BARF1 and BHRF1 are highly conserved which may contribute to maintain the biological function of these two genes. There is no evidence that particular EBV substrains of BARF1 or BHRF1 is region-restricted or disease-specific.

New Directions for Natural Killer T Cells in the Immunotherapy of Cancer

PubMed Central

Teyton, Luc

2017-01-01

Natural killer T (NKT) cells have been placed at the interface between innate and adaptive immunity by a long series of experiments that convincingly showed that beyond cytokine secretion and NK cell recruitment, NKT cells were coordinating dendritic cell and B cell maturation through direct membrane contacts and initiate productive responses. As such, NKT cells are the cellular adjuvant of many immune reactions and have functions that go much beyond what their name encapsulates. In addition, the initial discovery of the ligands of NKT cells is deeply linked to cancer biology and therapy. However, for a host of reasons, animal models in which agonists of NKT cells were used did not translate well to human cancers. A systematic reassessment of NKT cells role in tumorigenesis, especially spontaneous one, is now accessible using single cell analysis technologies both in mouse and man, and should be taken advantage of. Similarly, the migration, localization, phenotype of NKT cells following induced expansion after injection of an agonist can be examined at the single cell level. This technological revolution will help evaluate where and how NKT cells can be used in cancer. PMID:29209309

Pollution of mycological surfaces in hospital emergency departments correlates positively with blood NKT CD3+ 16+ 56+ and negatively with CD4+ cell levels of their staff

PubMed Central

Suska, Milena; Kiepura, Anna; Winnicka, Izabela; Leszczyński, Paweł; Bielawska-Drózd, Agata; Cieślik, Piotr; Kubiak, Leszek; Depczyńska, Daria; Brewczyńska, Aleksandra; Skopińska-Różewska, Ewa; Kocik, Janusz

2016-01-01

The aim of the present study was the assessment of the putative influence of yeast and filamentous fungi in healthcare and control (office) workplaces (10 of each kind) on immune system competence measured by NK (natural killer), CD4+, and NKT (natural killer T lymphocyte) cell levels in the blood of the personnel employed at these workplaces. Imprints from floors and walls were collected in winter. The blood was taken in spring the following year, from 40 men, 26 to 53 years old, healthcare workers of hospital emergency departments (HED), who had been working for at least five years in their current positions, and from 36 corresponding controls, working in control offices. Evaluation of blood leukocyte subpopulations was done by flow cytometry. The qualitative analysis of the surface samples revealed a prevalence of strains belonging to Aspergillus spp. and Penicillium spp. genus. There was no statistically significant difference between the level of NKT; however, the percentage of NK cells was lower in the blood of HED workers than in the blood of offices personnel. Spearman analysis revealed the existence of positive correlation (r = 0.4677, p = 0.002) between the total CFU/25 cm2 obtained by imprinting method from walls and floors of HED and the percentage of NKT (CD3+16+56+) lymphocytes collected from the blood of their personnel, and negative correlation (r = –0. 3688, p = 0.019) between this parameter of fungal pollution and the percentage of CD4+ lymphocytes in the blood of HED staff. No other correlations were found. PMID:27095925

Activation of CD1d-restricted T cells protects NOD mice from developing diabetes by regulating dendritic cell subsets

PubMed Central

Naumov, Yuri N.; Bahjat, Keith S.; Gausling, Rudolph; Abraham, Roshini; Exley, Mark A.; Koezuka, Yasuhiko; Balk, Steven B.; Strominger, Jack L.; Clare-Salzer, Michael; Wilson, S. Brian

2001-01-01

CD1d-restricted invariant NKT (iNKT) cells are immunoregulatory cells whose loss exacerbates diabetes in nonobese diabetic (NOD) female mice. Here, we show that the relative numbers of iNKT cells from the pancreatic islets of NOD mice decrease at the time of conversion from peri-insulitis to invasive insulitis and diabetes. Conversely, NOD male mice who have a low incidence of diabetes showed an increased frequency of iNKT cells. Moreover, administration of α-galactosylceramide, a potent activating ligand presented by CD1d, ameliorated the development of diabetes in NOD female mice and resulted in the accumulation of iNKT cells and myeloid dendritic cells (DC) in pancreatic lymph nodes (PLN), but not in inguinal lymph nodes. Strikingly, injection of NOD female mice with myeloid DC isolated from the PLN, but not those from the inguinal lymph nodes, completely prevented diabetes. Thus, the immunoregulatory role of iNKT cells is manifested by the recruitment of tolerogenic myeloid DC to the PLN and the inhibition of ongoing autoimmune inflammation. PMID:11707602

Dendritic cells combined with tumor cells and α-galactosylceramide induce a potent, therapeutic and NK-cell dependent antitumor immunity in B cell lymphoma.

PubMed

Escribà-Garcia, Laura; Alvarez-Fernández, Carmen; Tellez-Gabriel, Marta; Sierra, Jorge; Briones, Javier

2017-05-26

Invariant natural killer T (iNKT) cells are a small population of lymphocytes with unique specificity for glycolipid antigens presented by non-polymorphic CD1d receptor on dendritic cells (DCs). iNKT cells play a central role in tumor immunology since they are implicated in the coordination of innate and adaptive immune responses. These cells can be activated with the prototypic lipid α-galactosylceramide (α-GalCer), stimulating interferon gamma (IFN-γ) production and cytokine secretion, which contribute to the enhancement of T cell activation. We evaluated the antitumor effect of a combination of dendritic cells (DCs) and tumor cells with the iNKT cell agonist α-GalCer in a therapeutic model of B cell lymphoma. iNKT, NK and T cell phenotype was determined by flow cytometry. Serum cytokines were analyzed by Luminex technology. Significant differences between survival curves were assessed by the log-rank test. For all other data, Mann-Whitney test was used to analyze the differences between groups. This vaccine induced a potent (100% survival), long-lasting and tumor-specific antitumor immune response, that was associated with an increase of both Th1 cytokines and IFN-γ secreting iNKT cells (4.59 ± 0.41% vs. 0.92 ± 0.12% in control group; p = 0.01) and T cells (CD4 IFN-γ + : 3.75 ± 0.59% vs. 0.66 ± 0.18% p = 0.02; CD8 IFN-γ + : 10.61 ± 0.84% vs. 0.47 ± 0.03% p = 0.002). Importantly, natural killer (NK) cells played a critical role in the antitumor effect observed after vaccination. This study provides clinically relevant data for the development of iNKT-cell based immunotherapy treatments for patients with B cell malignancies.

Natural Killer T Cells: An Ecological Evolutionary Developmental Biology Perspective

PubMed Central

Kumar, Amrendra; Suryadevara, Naveenchandra; Hill, Timothy M.; Bezbradica, Jelena S.; Van Kaer, Luc; Joyce, Sebastian

2017-01-01

Type I natural killer T (NKT) cells are innate-like T lymphocytes that recognize glycolipid antigens presented by the MHC class I-like protein CD1d. Agonistic activation of NKT cells leads to rapid pro-inflammatory and immune modulatory cytokine and chemokine responses. This property of NKT cells, in conjunction with their interactions with antigen-presenting cells, controls downstream innate and adaptive immune responses against cancers and infectious diseases, as well as in several inflammatory disorders. NKT cell properties are acquired during development in the thymus and by interactions with the host microbial consortium in the gut, the nature of which can be influenced by NKT cells. This latter property, together with the role of the host microbiota in cancer therapy, necessitates a new perspective. Hence, this review provides an initial approach to understanding NKT cells from an ecological evolutionary developmental biology (eco-evo-devo) perspective. PMID:29312339

Natural Killer T Cells: An Ecological Evolutionary Developmental Biology Perspective.

PubMed

Kumar, Amrendra; Suryadevara, Naveenchandra; Hill, Timothy M; Bezbradica, Jelena S; Van Kaer, Luc; Joyce, Sebastian

2017-01-01

Type I natural killer T (NKT) cells are innate-like T lymphocytes that recognize glycolipid antigens presented by the MHC class I-like protein CD1d. Agonistic activation of NKT cells leads to rapid pro-inflammatory and immune modulatory cytokine and chemokine responses. This property of NKT cells, in conjunction with their interactions with antigen-presenting cells, controls downstream innate and adaptive immune responses against cancers and infectious diseases, as well as in several inflammatory disorders. NKT cell properties are acquired during development in the thymus and by interactions with the host microbial consortium in the gut, the nature of which can be influenced by NKT cells. This latter property, together with the role of the host microbiota in cancer therapy, necessitates a new perspective. Hence, this review provides an initial approach to understanding NKT cells from an ecological evolutionary developmental biology (eco-evo-devo) perspective.

Innate and cytokine-driven signals, rather than microbial antigens, dominate in natural killer T cell activation during microbial infection.

PubMed

Brigl, Manfred; Tatituri, Raju V V; Watts, Gerald F M; Bhowruth, Veemal; Leadbetter, Elizabeth A; Barton, Nathaniel; Cohen, Nadia R; Hsu, Fong-Fu; Besra, Gurdyal S; Brenner, Michael B

2011-06-06

Invariant natural killer T cells (iNKT cells) are critical for host defense against a variety of microbial pathogens. However, the central question of how iNKT cells are activated by microbes has not been fully explained. The example of adaptive MHC-restricted T cells, studies using synthetic pharmacological α-galactosylceramides, and the recent discovery of microbial iNKT cell ligands have all suggested that recognition of foreign lipid antigens is the main driver for iNKT cell activation during infection. However, when we compared the role of microbial antigens versus innate cytokine-driven mechanisms, we found that iNKT cell interferon-γ production after in vitro stimulation or infection with diverse bacteria overwhelmingly depended on toll-like receptor-driven IL-12. Importantly, activation of iNKT cells in vivo during infection with Sphingomonas yanoikuyae or Streptococcus pneumoniae, pathogens which are known to express iNKT cell antigens and which require iNKT cells for effective protection, also predominantly depended on IL-12. Constitutive expression of high levels of IL-12 receptor by iNKT cells enabled instant IL-12-induced STAT4 activation, demonstrating that among T cells, iNKT cells are uniquely equipped for immediate, cytokine-driven activation. These findings reveal that innate and cytokine-driven signals, rather than cognate microbial antigen, dominate in iNKT cell activation during microbial infections.

Different subsets of natural killer T cells may vary in their roles in health and disease

PubMed Central

Kumar, Vipin; Delovitch, Terry L

2014-01-01

Natural killer T cells (NKT) can regulate innate and adaptive immune responses. Type I and type II NKT cell subsets recognize different lipid antigens presented by CD1d, an MHC class-I-like molecule. Most type I NKT cells express a semi-invariant T-cell receptor (TCR), but a major subset of type II NKT cells reactive to a self antigen sulphatide use an oligoclonal TCR. Whereas TCR-α dominates CD1d-lipid recognition by type I NKT cells, TCR-α and TCR-β contribute equally to CD1d-lipid recognition by type II NKT cells. These variable modes of NKT cell recognition of lipid–CD1d complexes activate a host of cytokine-dependent responses that can either exacerbate or protect from disease. Recent studies of chronic inflammatory and autoimmune diseases have led to a hypothesis that: (i) although type I NKT cells can promote pathogenic and regulatory responses, they are more frequently pathogenic, and (ii) type II NKT cells are predominantly inhibitory and protective from such responses and diseases. This review focuses on a further test of this hypothesis by the use of recently developed techniques, intravital imaging and mass cytometry, to analyse the molecular and cellular dynamics of type I and type II NKT cell antigen-presenting cell motility, interaction, activation and immunoregulation that promote immune responses leading to health versus disease outcomes. PMID:24428389

Turned on by danger: activation of CD1d-restricted invariant natural killer T cells

PubMed Central

Lawson, Victoria

2012-01-01

CD1d-restricted invariant natural killer T (iNKT) cells bear characteristics of innate and adaptive lymphocytes, which allow them to bridge the two halves of the immune response and play roles in many disease settings. Recent work has characterized precisely how their activation is initiated and regulated. Novel antigens from important pathogens have been identified, as has an abundant self-antigen, β-glucopyranosylcaramide, capable of mediating an iNKT-cell response. Studies of the iNKT T-cell receptor (TCR)–antigen–CD1d complex show how docking between CD1d–antigen and iNKT TCR is highly conserved, and how small sequence differences in the TCR establish intrinsic variation in iNKT TCR affinity. The sequence of the TCR CDR3β loop determines iNKT TCR affinity for ligand–CD1d, independent of ligand identity. CD1d ligands can promote T helper type 1 (Th1) or Th2 biased cytokine responses, depending on the composition of their lipid tails. Ligands loaded into CD1d on the cell surface promote Th2 responses, whereas ligands with long hydrophobic tails are loaded endosomally and promote Th1 responses. This information is informing the design of synthetic iNKT-cell antigens. The iNKT cells may be activated by exogenous antigen, or by a combination of dendritic cell-derived interleukin-12 and iNKT TCR–self-antigen–CD1d engagement. The iNKT-cell activation is further modulated by recent foreign or self-antigen encounter. Activation of dendritic cells through pattern recognition receptors alters their antigen presentation and cytokine production, strongly influencing iNKT-cell activation. In a range of bacterial infections, dendritic cell-dependent innate activation of iNKT cells through interleukin-12 is the dominant influence on their activity. PMID:22734667

Isthmin 1 Is a Secreted Protein Expressed in Skin, Mucosal Tissues, and NK, NKT, and Th17 Cells

PubMed Central

Valle-Rios, Ricardo; Maravillas-Montero, José L.; Burkhardt, Amanda M.; Martinez, Cynthia; Buhren, Bettina Alexandra; Homey, Bernhard; Gerber, Peter Arne; Robinson, Octavio; Hevezi, Peter

2014-01-01

Using a comprehensive microarray database of human gene expression, we identified that in mammals, a secreted protein known as isthmin 1 (ISM1) is expressed in skin, mucosal tissues, and selected lymphocyte populations. ISM1 was originally identified in Xenopus brain during development, and it encodes a predicted ∼50-kDa protein containing a signal peptide, a thrombospondin domain, and an adhesion-associated domain. We confirmed the pattern of expression of ISM1 in both human and mouse tissues. ISM1 is expressed by DX5+ lung lymphocytes that include NK and NKT-like cells, and is also expressed by some CD4+ T cells upon activation but its expression increases significantly when CD4+ T cells were polarized to the Th17 lineage in vitro. The presence of IFN-γ during CD4+ T cell polarization inhibits ISM1 expression. Given that ISM1 has been reported to have anti-angiogenic properties, these observations suggest that ISM1 is a mediator of lymphocyte effector functions and may participate in both innate and acquired immune responses. PMID:24956034

Isthmin 1 is a secreted protein expressed in skin, mucosal tissues, and NK, NKT, and th17 cells.

PubMed

Valle-Rios, Ricardo; Maravillas-Montero, José L; Burkhardt, Amanda M; Martinez, Cynthia; Buhren, Bettina Alexandra; Homey, Bernhard; Gerber, Peter Arne; Robinson, Octavio; Hevezi, Peter; Zlotnik, Albert

2014-10-01

Using a comprehensive microarray database of human gene expression, we identified that in mammals, a secreted protein known as isthmin 1 (ISM1) is expressed in skin, mucosal tissues, and selected lymphocyte populations. ISM1 was originally identified in Xenopus brain during development, and it encodes a predicted ∼50-kDa protein containing a signal peptide, a thrombospondin domain, and an adhesion-associated domain. We confirmed the pattern of expression of ISM1 in both human and mouse tissues. ISM1 is expressed by DX5(+) lung lymphocytes that include NK and NKT-like cells, and is also expressed by some CD4(+) T cells upon activation but its expression increases significantly when CD4(+) T cells were polarized to the Th17 lineage in vitro. The presence of IFN-γ during CD4(+) T cell polarization inhibits ISM1 expression. Given that ISM1 has been reported to have anti-angiogenic properties, these observations suggest that ISM1 is a mediator of lymphocyte effector functions and may participate in both innate and acquired immune responses.

Innate-like behavior of human invariant natural killer T cells during herpes simplex virus infection.

PubMed

Novakova, Lucie; Nevoralova, Zuzana; Novak, Jan

2012-01-01

Invariant natural killer T (iNKT) cells, CD1d restricted T cells, are involved in the immune responses against various infection agents. Here we describe their behavior during reactivation of human herpes simplex virus (HSV). iNKT cells exhibit only discrete changes, which however, reached statistically significant level due to the relatively large patient group. Higher percentage of iNKT cells express NKG2D. iNKT cells down-regulate NKG2A in a subset of patients. Finally, iNKT cells enhance their capacity to produce TNF-α. Our data suggests that iNKT cells are involved in the immune response against HSV and contribute mainly to its early, innate phase. Copyright © 2012 Elsevier Inc. All rights reserved.

Different subsets of natural killer T cells may vary in their roles in health and disease.

PubMed

Kumar, Vipin; Delovitch, Terry L

2014-07-01

Natural killer T cells (NKT) can regulate innate and adaptive immune responses. Type I and type II NKT cell subsets recognize different lipid antigens presented by CD1d, an MHC class-I-like molecule. Most type I NKT cells express a semi-invariant T-cell receptor (TCR), but a major subset of type II NKT cells reactive to a self antigen sulphatide use an oligoclonal TCR. Whereas TCR-α dominates CD1d-lipid recognition by type I NKT cells, TCR-α and TCR-β contribute equally to CD1d-lipid recognition by type II NKT cells. These variable modes of NKT cell recognition of lipid-CD1d complexes activate a host of cytokine-dependent responses that can either exacerbate or protect from disease. Recent studies of chronic inflammatory and autoimmune diseases have led to a hypothesis that: (i) although type I NKT cells can promote pathogenic and regulatory responses, they are more frequently pathogenic, and (ii) type II NKT cells are predominantly inhibitory and protective from such responses and diseases. This review focuses on a further test of this hypothesis by the use of recently developed techniques, intravital imaging and mass cytometry, to analyse the molecular and cellular dynamics of type I and type II NKT cell antigen-presenting cell motility, interaction, activation and immunoregulation that promote immune responses leading to health versus disease outcomes. © 2014 John Wiley & Sons Ltd.

A Case Report of NK-Cell Lymphoproliferative Disease With a Wide Involvement of Digestive Tract Develop Into Epstein-Barr Virus Associated NK/T Cell Lymphoma in an Immunocompetent Patient.

PubMed

Chen, Haotian; Zhang, Yu; Jiang, Zhinong; Zhou, Wei; Cao, Qian

2016-03-01

Epstein-Barr virus (EBV) plays an important role in various diseases. EBV-associated lymphoproliferative disease (LPD) is a rare disease with a canceration tendency. It is difficult to differentiate LPD with involvement of digestive tract from Crohn disease due to similar clinical and endoscopic manifestations. We present a case report of multiple ulcers with esophagus, small bowel and the entire colon involved, proved to be NK-Cell LPD, developed into EBV-associated NK/T Cell lymphoma, in an immunocompetent man who was initially misdiagnosed as Crohn disease.This report underscores that intestinal ulcers should be cautiously diagnosed, for it sometimes could be a precancerous lesion.

Invariant natural killer T cells act as an extravascular cytotoxic barrier for joint-invading Lyme Borrelia.

PubMed

Lee, Woo-Yong; Sanz, Maria-Jesus; Wong, Connie H Y; Hardy, Pierre-Olivier; Salman-Dilgimen, Aydan; Moriarty, Tara J; Chaconas, George; Marques, Adriana; Krawetz, Roman; Mody, Christopher H; Kubes, Paul

2014-09-23

CXCR6-GFP(+) cells, which encompass 70% invariant natural killer T cells (iNKT cells), have been found primarily patrolling inside blood vessels in the liver. Although the iNKT cells fail to interact with live pathogens, they do respond to bacterial glycolipids presented by CD1d on liver macrophage that have caught the microbe. In contrast, in this study using dual laser multichannel spinning-disk intravital microscopy of joints, the CXCR6-GFP, which also made up 60-70% iNKT cells, were not found in the vasculature but rather closely apposed to and surrounding the outside of blood vessels, and to a lesser extent throughout the extravascular space. These iNKT cells also differed in behavior, responding rapidly and directly to joint-homing pathogens like Borrelia burgdorferi, which causes Lyme disease. These iNKT cells interacted with B. burgdorferi at the vessel wall and disrupted dissemination attempts by these microbes into joints. Successful penetrance of B. burgdorferi out of the vasculature and into the joint tissue was met by a lethal attack by extravascular iNKT cells through a granzyme-dependent pathway, an observation also made in vitro for iNKT cells from joint but not liver or spleen. These results suggest a novel, critical extravascular iNKT cell immune surveillance in joints that functions as a cytotoxic barrier and explains a large increase in pathogen burden of B. burgdorferi in the joint of iNKT cell-deficient mice, and perhaps the greater susceptibility of humans to this pathogen because of fewer iNKT cells in human joints.

Deficiencies of Circulating Mucosal-associated Invariant T Cells and Natural Killer T Cells in Patients with Multiple Trauma.

PubMed

Jo, Young Goun; Choi, Hyun Jung; Kim, Jung Chul; Cho, Young Nan; Kang, Jeong Hwa; Jin, Hye Mi; Kee, Seung Jung; Park, Yong Wook

2017-05-01

Mucosal-associated invariant T (MAIT) cells and natural killer T (NKT) cells are known to play important roles in autoimmunity, infectious diseases and cancers. However, little is known about the roles of these invariant T cells in multiple trauma. The purposes of this study were to examine MAIT and NKT cell levels in patients with multiple trauma and to investigate potential relationships between these cell levels and clinical parameters. The study cohort was composed of 14 patients with multiple trauma and 22 non-injured healthy controls (HCs). Circulating MAIT and NKT cell levels in the peripheral blood were measured by flow cytometry. The severity of injury was categorised according to the scoring systems, such as Acute Physiology and Chronic Health Evaluation (APACHE) II score, Simplified Acute Physiology Score (SAPS) II, and Injury Severity Score (ISS). Circulating MAIT and NKT cell numbers were significantly lower in multiple trauma patients than in HCs. Linear regression analysis showed that circulating MAIT cell numbers were significantly correlated with age, APACHE II, SAPS II, ISS category, hemoglobin, and platelet count. NKT cell numbers in the peripheral blood were found to be significantly correlated with APACHE II, SAPS II, and ISS category. This study shows numerical deficiencies of circulating MAIT cells and NKT cells in multiple trauma. In addition, these invariant T cell deficiencies were found to be associated with disease severity. These findings provide important information for predicting the prognosis of multiple trauma. © 2017 The Korean Academy of Medical Sciences.

Third-party CD4+ invariant natural killer T cells protect from murine GVHD lethality

PubMed Central

Schneidawind, Dominik; Baker, Jeanette; Pierini, Antonio; Buechele, Corina; Luong, Richard H.; Meyer, Everett H.

2015-01-01

Graft-versus-host disease (GVHD) is driven by extensive activation and proliferation of alloreactive donor T cells causing significant morbidity and mortality following allogeneic hematopoietic cell transplantation (HCT). Invariant natural killer T (iNKT) cells are a potent immunoregulatory T-cell subset in both humans and mice. Here, we explored the role of adoptively transferred third-party CD4+ iNKT cells for protection from lethal GVHD in a murine model of allogeneic HCT across major histocompatibility barriers. We found that low numbers of CD4+ iNKT cells from third-party mice resulted in a significant survival benefit with retained graft-versus-tumor effects. In vivo expansion of alloreactive T cells was diminished while displaying a T helper cell 2-biased phenotype. Notably, CD4+ iNKT cells from third-party mice were as protective as CD4+ iNKT cells from donor mice although third-party CD4+ iNKT cells were rejected early after allogeneic HCT. Adoptive transfer of third-party CD4+ iNKT cells resulted in a robust expansion of donor CD4+CD25+FoxP3+ regulatory T cells (Tregs) that were required for protection from lethal GVHD. However, in vivo depletion of myeloid-derived suppressor cells abrogated both Treg expansion and protection from lethal GVHD. Despite the fact that iNKT cells are a rare cell population, the almost unlimited third-party availability and feasibility of in vitro expansion provide the basis for clinical translation. PMID:25795920

Invariant Natural Killer T Cells are Reduced in Hereditary Hemochromatosis Patients.

PubMed

Maia, M L; Pereira, C S; Melo, G; Pinheiro, I; Exley, M A; Porto, G; Macedo, M F

2015-01-01

Invariant natural killer T (iNKT) cells are CD1d restricted-T cells that react to lipid antigens. iNKT cells were shown to be important in infection, autoimmunity and tumor surveillance. Alterations in the number and function of these cells were described in several pathological conditions including autoimmune and/or liver diseases. CD1d is critical for antigen presentation to iNKT cells, and its expression is increased in liver diseases. The liver is the major organ affected in Hereditary Hemochromatosis (HH), an autosomal recessive disorder caused by excessive iron absorption. Herein, we describe the study of iNKT cells of HH patients. Twenty-eight HH patients and 24 control subjects from Santo António Hospital, Porto, were included in this study. Patient's iron biochemical parameters (serum transferrin saturation and ferritin levels) and the liver function marker alanine transaminase (ALT) were determined at the time of study. Peripheral blood iNKT cells were analyzed by flow cytometry using an anti-CD3 antibody and the CD1d tetramer loaded with PBS57. We found a decrease in the percentage and number of circulating iNKT cells from HH patients when compared with control population independently of age. iNKT cell defects were more pronounced in untreated patients, relating with serum ferritin and transferrin saturation levels. No correlation was found with ALT, a marker of active liver dysfunction. Altogether, our results demonstrate that HH patients have reduced numbers of iNKT cells and that these are influenced by iron overload.

P02.03INCREASED COUNTS OF NK AND NKT CELLS ARE ASSOCIATED WITH PROLONGED SURVIVAL IN PRIMARY GLIOBLASTOMA PATIENTS TREATED WITH DENDRITIC CELL IMMUNOTHERAPY IN COMBINATION WITH RADIO- AND CHEMO-THERAPY

PubMed Central

Pellegatta, S.; Eoli, M.; Cantini, G.; Anghileri, E.; Antozzi, C.; Frigerio, S.; Bruzzone, M.; Pollo, B.; Parati, E.; Finocchiaro, G.

2014-01-01

Two clinical studies, DENDR1 and DENDR2 including, respectively, the treatment of first diagnosis and recurrent glioblastoma (GB) patients with dendritic cells (DCs) loaded with autologous tumor lysate are currently active at Istituto Neurologico Besta, Milan. Our first results obtained on a group of recurrent GB patients demonstrated that the response of NK cells correlates with significantly prolonged survival. Here we provide results of the interim analysis on 22 patients affected by primary GB. Patients with post-surgery volume ≤10 cc underwent leukapheresis before radiotherapy and chemotherapy with temozolomide (TMZ). Three intradermal injections of mature DC were done before adjuvant chemotherapy. The subsequent 4 injections were performed 17 ± 3 days after adjuvant TMZ. MRI, clinical and immunological follow-up were performed every 2 months. The median age at surgery was 54.5 years (28-69). RT-TMZ induced significant lymphopenia (<1000 lymphocytes/microl) in 17/22 patients (77.2%). Patients with >1000 lymphocytes/microl (5/22) before first vaccination had shorter PFS than others (p < 0.005). Peripheral Blood Lymphocytes (PBLs) were analyzed by flow cytometry to identify CD8+ T cells, NK and NKT cells before and after DC vaccines. The ratio of vaccination/baseline frequencies and counts (V/B ratio) of all of the immunological parameters for each patient was calculated, and the median of all of the observations used as the cut off value to separate patients. V/B ratio was correlated with the progression free survival (PFS) of each patient. Increased V/B ratio for NK cells and in particular NKT cells, but not for CD8 T lymphocytes, was significantly associated with prolonged PFS (median PFS 14 vs 8.0 mo, p = 0.01; 15.0 vs 8.0 mo, respectively). Interferon (IFN)-γ in PBLs was significantly higher in patients with PFS12 (p < 0.02), increasing immediately after the second vaccination as evaluated by real time-PCR. No changes in the expression levels of IFN

Invariant natural killer T-cell control of type 1 diabetes: a dendritic cell genetic decision of a silver bullet or Russian roulette.

PubMed

Driver, John P; Scheuplein, Felix; Chen, Yi-Guang; Grier, Alexandra E; Wilson, S Brian; Serreze, David V

2010-02-01

In part, activation of invariant natural killer T (iNKT)-cells with the superagonist alpha-galactosylceramide (alpha-GalCer) inhibits the development of T-cell-mediated autoimmune type 1 diabetes in NOD mice by inducing the downstream differentiation of antigen-presenting dendritic cells (DCs) to an immunotolerogenic state. However, in other systems iNKT-cell activation has an adjuvant-like effect that enhances rather than suppresses various immunological responses. Thus, we tested whether in some circumstances genetic variation would enable activated iNKT-cells to support rather than inhibit type 1 diabetes development. We tested whether iNKT-conditioned DCs in NOD mice and a major histocompatibility complex-matched C57BL/6 (B6) background congenic stock differed in capacity to inhibit type 1 diabetes induced by the adoptive transfer of pathogenic AI4 CD8 T-cells. Unlike those of NOD origin, iNKT-conditioned DCs in the B6 background stock matured to a state that actually supported rather than inhibited AI4 T-cell-induced type 1 diabetes. The induction of a differing activity pattern of T-cell costimulatory molecules varying in capacity to override programmed death-ligand-1 inhibitory effects contributes to the respective ability of iNKT-conditioned DCs in NOD and B6 background mice to inhibit or support type 1 diabetes development. Genetic differences inherent to both iNKT-cells and DCs contribute to their varying interactions in NOD and B6.H2(g7) mice. This great variability in the interactions between iNKT-cells and DCs in two inbred mouse strains should raise a cautionary note about considering manipulation of this axis as a potential type 1 diabetes prevention therapy in genetically heterogeneous humans.

Enhanced cytotoxic function of natural killer and natural killer T-like cells associated with decreased CD94 (Kp43) in the chronic obstructive pulmonary disease airway.

PubMed

Hodge, Greg; Mukaro, Violet; Holmes, Mark; Reynolds, Paul N; Hodge, Sandra

2013-02-01

Natural killer (NK) and natural killer T (NKT)-like cells represent a small but important proportion of effector lymphocytes that we have previously shown to be major sources of pro-inflammatory cytokines and granzymes. We hypothesized that these cells would be increased in the airway in chronic obstructive pulmonary disease (COPD), accompanied by reduced expression of the inhibitory receptor CD94 (Kp43) and increased expression of cytotoxic mediators granzyme B and perforin. We measured NK and NKT-like cells and their expression of CD94 in the blood of COPD patients (n = 71; 30 current and 41 ex-smokers), smokers (16) and healthy controls (25), and bronchoalveolar lavage fluid (BALF) from a cohort of subjects (19 controls, 12 smokers, 33 COPD). Activation was assessed by measuring CD69 in blood and the cytotoxic potential of NK cells by measuring granzymes A and B, and using a cytotoxicity assay in blood and BALF. In blood in COPD, there were no significant changes in the proportion of NK or NKT-like cells or expression of granzyme A or NK cytotoxic potential versus controls. There was, however, increased expression of granzyme B and decreased expression of CD94 by both cell types versus controls. The proportion of NK and NKT-like cells were increased in BALF in COPD, associated with increased NK cytotoxicity, increased expression of granzyme B and decreased expression of the inhibitory receptor CD94 by both cell types. Treatment strategies that target NK and NKT-like cells, their cytotoxicity and production of inflammatory mediators in the airway may improve COPD morbidity. © 2012 The Authors. Respirology © 2012 Asian Pacific Society of Respirology.

Characterization of human invariant natural killer T subsets in health and disease using a novel invariant natural killer T cell-clonotypic monoclonal antibody, 6B11

PubMed Central

Montoya, Carlos J; Pollard, David; Martinson, Jeffrey; Kumari, Kumud; Wasserfall, Clive; Mulder, Candice B; Rugeles, Maria T; Atkinson, Mark A; Landay, Alan L; Wilson, S Brian

2007-01-01

Identification of human CD1d-restricted T-cell receptor (TCR)-invariant natural killer T (iNKT) cells has been dependent on utilizing combinations of monoclonal antibodies or CD1d tetramers, which do not allow for the most specific analysis of this T-cell subpopulation. A novel monoclonal antibody (clone 6B11), specific for the invariant CDR3 loop of human canonical Vα24Jα18 TCR α chain, was developed and used to specifically characterize iNKT cells. In healthy individuals studied for up to 1 year, a wide but stable frequency of circulating iNKT cells (range: 0·01–0·92%) was observed, with no differences in frequency by gender. Four stable iNKT cell subsets were characterized in peripheral blood based on the expression of CD4 and CD8, with CD8+ iNKT cells being a phenotypic and functionally different subset from CD4+ and double negative iNKT cells; in particular, LAG-3 was preferentially expressed on CD8+ iNKT cells. In addition, a strong negative linear correlation between the frequency of total iNKT cells and percentage of the CD4+ subset was observed. In terms of their potential association with disease, patients at risk for type 1 diabetes had significantly expanded frequencies of double negative iNKT cells when compared to matched controls and first-degree relatives. Moreover, peripheral blood CD4+ iNKT cells were the highest producers of interleukin-4, while the production of interferon-γ and tumour necrosis factor-α was similar amongst all iNKT cell subsets. These differences in iNKT cell subsets suggest that in humans the relative ratio of iNKT cell subsets may influence susceptibility vs. resistance to immune-mediated diseases. PMID:17662044

Size of the population of CD4+ natural killer T cells in the liver is maintained without supply by the thymus during adult life

PubMed Central

Kameyama, Hitoshi; Kawamura, Toshihiko; Naito, Tetsuya; Bannai, Makoto; Shimamura, Kazuhiko; Hatakeyama, Katsuyoshi; Abo, Toru

2001-01-01

Given that there are few natural killer T (NKT) cells in the liver of athymic nude mice and in neonatally thymectomized mice, it is still controversial whether all NKT cells existing in the liver are supplied by the thymus or if some such cells develop in the liver. To determine whether or not NKT cells are consistently supplied from the thymus during adult life, thymectomy was conducted in mice at the age of 8 weeks. Interestingly, the proportion and number of CD4+ NKT cells increased or remained unchanged in the liver after adult thymectomy and this phenomenon continued for up to 6 months after thymectomy. The administration of α-galactosylceramide induced severe cytopenia (due to apoptosis) of CD4+ NKT cells in the liver on day 1, but subsequent expansion of these NKT cells occurred in thymectomized mice similar to the case in normal mice. However, in thymectomized mice given lethal irradiation (9·5 Gy) and subsequent bone marrow transfer, the population of CD4+ NKT cells no longer expanded in the liver, although that of CD8+ NKT cells did. These results suggest that thymic CD4+ NKT cells, or their progenitors, may migrate to the liver at a neonatal stage but are not supplied from the thymus in the adult stage under usual conditions. CD8+ NKT cells can be generated in the liver. PMID:11683952

Invariant Natural Killer T Cell Deficiency and Functional Impairment in Sleep Apnea: Links to Cancer Comorbidity.

PubMed

Gaoatswe, Gadintshware; Kent, Brian D; Corrigan, Michelle A; Nolan, Geraldine; Hogan, Andrew E; McNicholas, Walter T; O'Shea, Donal

2015-10-01

Emerging evidence links obstructive sleep apnea (OSA) with increased cancer incidence and mortality. Invariant natural killer T (iNKT) cells play an important role in cancer immunity. We hypothesized that patients with OSA have low number of circulating invariant natural killer T (iNKT) cells, which may also be functionally impaired. This study aims to evaluate the frequency of circulating iNKT cells in OSA. We evaluated the frequency of circulating iNKT cells by flow cytometry in 33 snorers being assessed for possible OSA. Using iNKT cell lines, we also evaluated the effect of exposure to hypoxia over 24 hours on apoptosis, cytotoxicity, and cytokine production. Teaching hospital based sleep unit and research laboratory. Thirty-three snorers were evaluated: 9 with no OSA (apnea-hypopnea frequency [AHI] < 5/h), 12 with mild-moderate OSA (AHI 5-30) and 12 with severe OSA (AHI > 30). Patients with severe OSA had considerably fewer iNKT cells (0.18%) compared to patients with mild-moderate (0.24%) or no OSA (0.35%), P = 0.0026. The frequency of iNKT cells correlated negatively with apnea-hypopnea index (r = -0.58, P = 0.001), oxygen desaturation index (r = -0.58, P = 0.0003), and SpO2% < 90% (r = -0.5407, P = 0.005). The frequency of iNKT cells increased following 12 months of nCPAP therapy (P = 0.015). Hypoxia resulted in increased apoptosis (P = 0.016) and impaired cytotoxicity (P = 0.035). Patients with obstructive sleep apnea (OSA) have significantly reduced levels of circulating invariant natural killer T (iNKT) cells and hypoxia leads to impaired iNKT cell function. These observations may partly explain the increased cancer risk reported in patients with OSA. © 2015 Associated Professional Sleep Societies, LLC.

Coevolution of MHC genes (LMP/TAP/class Ia; NKT-class Ib; NKp30-B7H6): Lessons from cold-blooded vertebrates

PubMed Central

Ohta, Yuko; Flajnik, Martin F.

2015-01-01

Summary Comparative immunology provides the long view of what is conserved across all vertebrate taxa versus what is specific to particular organisms or group of organisms. Regarding the major histocompatibility complex (MHC) and coevolution, three striking cases have been revealed in cold-blooded vertebrates: lineages of class Ia antigen-processing and -presenting genes, evolutionary conservation of NKT-class Ib recognition, and the ancient emergence of the natural cytotoxicity receptor NKp30 and its ligand B7H6. While coevolution of transporter associated with antigen processing (TAP) and class Ia has been documented in endothermic birds and two mammals, lineages of LMP7 are restricted to ectotherms. The unambiguous discovery of natural killer T (NKT) cells in Xenopus demonstrated that NKT cells are not restricted to mammals and are likely to have emerged at the same time in evolution as classical α/β and γ/δ T cells. NK cell receptors evolve at a rapid rate, and orthologues are nearly impossible to identify in different vertebrate classes. By contrast, we have detected NKp30 in all gnathostomes, except in species where it was lost. The recently discovered ligand of NKp30, B7H6, shows strong signs of coevolution with NKp30 throughout evolution, i.e. coincident loss or expansion of both genes in some species. NKp30 also offers an attractive IgSF candidate for the invasion of the RAG transposon, which is believed to have initiated T-cell receptor/immunoglobulin adaptive immunity. Besides reviewing these intriguing features of MHC evolution and coevolution, we offer suggestions for future studies and propose a model for the primordial or proto MHC. PMID:26284468

Natural Killer Dendritic Cells Enhance Immune Responses Elicited by α -Galactosylceramide-Stimulated Natural Killer T Cells.

PubMed

Lee, Sung Won; Park, Hyun Jung; Kim, Nayoung; Hong, Seokmann

2013-01-01

Natural killer dendritic cells (NKDCs) possess potent anti-tumor activity, but the cellular effect of NKDC interactions with other innate immune cells is unclear. In this study, we demonstrate that the interaction of NKDCs and natural killer T (NKT) cells is required for the anti-tumor immune responses that are elicited by α -galactosylceramide ( α -GC) in mice. The rapid and strong expression of interferon- γ by NKDCs after α -GC stimulation was dependent on NKT cells. Various NK and DC molecular markers and cytotoxic molecules were up-regulated following α -GC administration. This up-regulation could improve NKDC presentation of tumor antigens and increase cytotoxicity against tumor cells. NKDCs were required for the stimulation of DCs, NK cells, and NKT cells. The strong anti-tumor immune responses elicited by α -GC may be due to the down-regulation of regulatory T cells. Furthermore, the depletion of NKDCs dampened the tumor clearance mediated by α -GC-stimulated NKT cells in vivo. Taken together, these results indicate that complex interactions of innate immune cells might be required to achieve optimal anti-tumor immune responses during the early stages of tumorigenesis.

Expansion of highly activated invariant natural killer T cells with altered phenotype in acute dengue infection

PubMed Central

Kamaladasa, A.; Wickramasinghe, N.; Adikari, T. N.; Gomes, L.; Shyamali, N. L. A.; Salio, M.; Cerundolo, V.; Ogg, G. S.

2016-01-01

Summary Invariant natural killer T (iNKT) cells are capable of rapid activation and production of cytokines upon recognition of antigenic lipids presented by CD1d molecules. They have been shown to play a significant role in many viral infections and were observed to be highly activated in patients with acute dengue infection. In order to characterize further their role in dengue infection, we investigated the proportion of iNKT cells and their phenotype in adult patients with acute dengue infection. The functionality of iNKT cells in patients was investigated by both interferon (IFN)‐γ and interleukin (IL)−4 ex‐vivo enzyme‐linked immunospot (ELISPOT) assays following stimulation with alpha‐galactosyl‐ceramide (αGalCer). We found that circulating iNKT cell proportions were significantly higher (P = 0·03) in patients with acute dengue when compared to healthy individuals and were predominantly of the CD4+ subset. iNKT cells of patients with acute dengue had reduced proportions expressing CD8α and CD161 when compared to healthy individuals. The iNKT cells of patients were highly activated and iNKT activation correlated significantly with dengue virus‐specific immunoglobulin (Ig)G antibody levels. iNKT cells expressing Bcl‐6 (P = 0·0003) and both Bcl‐6 and inducible T cell co‐stimulator (ICOS) (P = 0·006) were increased significantly in patients when compared to healthy individuals. Therefore, our data suggest that in acute dengue infection there is an expansion of highly activated CD4+ iNKT cells, with reduced expression of CD161 markers. PMID:26874822

Invariant natural killer T cells ameliorate murine chronic GVHD by expanding donor regulatory T cells.

PubMed

Du, Jing; Paz, Katelyn; Thangavelu, Govindarajan; Schneidawind, Dominik; Baker, Jeanette; Flynn, Ryan; Duramad, Omar; Feser, Colby; Panoskaltsis-Mortari, Angela; Negrin, Robert S; Blazar, Bruce R

2017-06-08

Chronic graft-versus-host-disease (cGVHD) can cause multiorgan system disease, typically with autoimmune-like features, resulting in high mortality and morbidity caused by treatment limitations. Invariant natural killer T cells (iNKTs), a small population characterized by expression of a semi-invariant T-cell receptor, rapidly produce copious amounts of diverse cytokines on activation that exert potent immune regulatory function. Here, we show that iNKTs are significantly reduced in a cGVHD murine model that recapitulates several aspects of autoimmunity and organ fibrosis observed in patients with cGVHD. Low iNKT infused doses effectively prevented and, importantly, reversed established cGVHD, as did third-party iNKTs. iNKTs suppressed the autoimmune response by reducing the germinal center (GC) reaction, which was associated with an increase in total Tregs and follicular Tregs (Tfr) that control the GC reaction, along with pathogenic antibody production. Treg depletion during iNKT infusions completely abolished iNKT efficacy in treating cGVHD. iNKT cell interleukin 4 production and GC migration were critical to cGVHD reversal. In vivo stimulation of iNKT cells by α-galactosyl-ceramide was effective in both preventing and treating cGVHD. Together, this study demonstrates iNKT deficiency in cGVHD mice and highlights the key role of iNKTs in regulating cGVHD pathogenesis and as a potentially novel prophylactic and therapeutic option for patients with cGVHD. © 2017 by The American Society of Hematology.

T cell-recruiting triplebody 19-3-19 mediates serial lysis of malignant B-lymphoid cells by a single T cell

PubMed Central

Roskopf, Claudia C.; Schiller, Christian B.; Braciak, Todd A.; Kobold, Sebastian; Schubert, Ingo A.; Fey, Georg H.; Hopfner, Karl-Peter; Oduncu, Fuat S.

2014-01-01

Triplebody 19-3-19, an antibody-derived protein, carries three single chain fragment variable domains in tandem in a single polypeptide chain. 19-3-19 binds CD19-bearing lymphoid cells via its two distal domains and primary T cells via its CD3-targeting central domain in an antigen-specific manner. Here, malignant B-lymphoid cell lines and primary cells from patients with B cell malignancies were used as targets in cytotoxicity tests with pre-stimulated allogeneic T cells as effectors. 19-3-19 mediated up to 95% specific lysis of CD19-positive tumor cells and, at picomolar EC50 doses, had similar cytolytic potency as the clinically successful agent BlinatumomabTM. 19-3-19 activated resting T cells from healthy unrelated donors and mediated specific lysis of both autologous and allogeneic CD19-positive cells. 19-3-19 led to the elimination of 70% of CD19-positive target cells even with resting T cells as effectors at an effector-to-target cell ratio of 1 : 10. The molecule is therefore capable of mediating serial lysis of target cells by a single T cell. These results highlight that central domains capable of engaging different immune effectors can be incorporated into the triplebody format to provide more individualized therapy tailored to a patient’s specific immune status. PMID:25115385

Dendritic cell internalization of α-galactosylceramide from CD8 T cells induces potent antitumor CD8 T-cell responses.

PubMed

Choi, Dong Hoon; Kim, Kwang Soon; Yang, Se Hwan; Chung, Doo Hyun; Song, Boyeong; Sprent, Jonathan; Cho, Jae Ho; Sung, Young Chul

2011-12-15

Dendritic cells (DC) present α-galactosylceramide (αGalCer) to invariant T-cell receptor-expressing natural killer T cells (iNKT) activating these cells to secrete a variety of cytokines, which in turn results in DC maturation and activation of other cell types, including NK cells, B cells, and conventional T cells. In this study, we showed that αGalCer-pulsing of antigen-activated CD8 T cells before adoptive transfer to tumor-bearing mice caused a marked increase in donor T-cell proliferation, precursor frequency, and cytotoxic lymphocyte activity. This effect was interleukin (IL)-2 dependent and involved both natural killer T cells (NKT) and DCs, as mice lacking IL-2, NKTs, and DCs lacked any enhanced response to adoptively transferred αGalCer-loaded CD8 T cells. iNKT activation was mediated by transfer of αGalCer from the cell membrane of the donor CD8 T cells onto the αGalCer receptor CD1d which is present on host DCs. αGalCer transfer was increased by prior activation of the donor CD8 T cells and required AP-2-mediated endocytosis by host DCs. In addition, host iNKT cell activation led to strong IL-2 synthesis, thereby increasing expansion and differentiation of donor CD8 T cells. Transfer of these cells led to improved therapeutic efficacy against established solid tumors in mice. Thus, our findings illustrate how αGalCer loading of CD8 T cells after antigen activation in vitro may leverage the therapeutic potential of adoptive T-cell therapies.

Innate lymphoid cells and natural killer T cells in the gastrointestinal tract immune system.

PubMed

Montalvillo, Enrique; Garrote, José Antonio; Bernardo, David; Arranz, Eduardo

2014-05-01

The gastrointestinal tract is equipped with a highly specialized intrinsic immune system. However, the intestine is exposed to a high antigenic burden that requires a fast, nonspecific response -so-called innate immunity- to maintain homeostasis and protect the body from incoming pathogens. In the last decade multiple studies helped to unravel the particular developmental requirements and specific functions of the cells that play a role in innate immunity. In this review we shall focus on innate lymphoid cells, a newly discovered, heterogeneous set of cells that derive from an Id2-dependent lymphoid progenitor cell population. These cells have been categorized on the basis of the pattern of cytokines that they secrete, and the transcription factors that regulate their development and functions. Innate lymphoid cells play a role in the early response to pathogens, the anatomical contention of the commensal flora, and the maintenance of epithelial integrity.Amongst the various innate lymphoid cells we shall lay emphasis on a subpopulation with several peculiarities, namely that of natural killer T cells, a subset of T lymphocytes that express both T-cell and NK-cell receptors. The most numerous fraction of the NKT population are the so-called invariant NKT or iNKT cells. These iNKT cells have an invariant TCR and recognize the glycolipidic structures presented by the CD1d molecule, a homolog of class-I MHC molecules. Following activation they rapidly acquire cytotoxic activity and secrete both Th1 and Th2 cytokines, including IL-17. While their specific role is not yet established, iNKT cells take part in a great variety of intestinal immune responses ranging from oral tolerance to involvement in a number of gastrointestinal conditions.

Contribution of Invariant Natural Killer T Cells to Skin Wound Healing.

PubMed

Tanno, Hiromasa; Kawakami, Kazuyoshi; Ritsu, Masae; Kanno, Emi; Suzuki, Aiko; Kamimatsuno, Rina; Takagi, Naoyuki; Miyasaka, Tomomitsu; Ishii, Keiko; Imai, Yoshimichi; Maruyama, Ryoko; Tachi, Masahiro

2015-12-01

In the present study, we determined the contribution of invariant natural killer T (iNKT) cells to the skin wound healing process. In iNKT cell-deficient (Jα18KO) mice lacking iNKT cells, wound closure was significantly delayed compared with wild-type mice. Collagen deposition, expression of α-smooth muscle actin and CD31, and wound breaking strength were significantly attenuated in Jα18KO mice. The adoptive transfer of liver mononuclear cells from wild-type but not from Jα18KO or interferon (IFN)-γ gene-disrupted (IFN-γKO) mice resulted in the reversal of this impaired wound healing in Jα18KO mice. IFN-γ expression was induced in the wounded tissues, which was significantly decreased at 6, 12, and 24 hours, but increased on day 3 after wounding in Jα18KO mice. The main source of the late-phase IFN-γ production in Jα18KO mice were neutrophils rather than NK cells and T cells. Administration of α-galactosylceramide, an activator of iNKT cells, resulted in the acceleration of wound healing on day 3 in wild-type mice. This effect was not observed in IFN-γKO mice. These results indicate that iNKT cells play important roles in wound healing. The iNKT cell-induced IFN-γ production may regulate the wound healing process in the early phase. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Regulatory T cells and other lymphocyte subpopulations in patients with melanoma developing interferon-induced thyroiditis during high-dose interferon-α2b treatment.

PubMed

Soldevila, Berta; Alonso, Núria; Martínez-Arconada, Maria J; Granada, Maria L; Boada, Aram; Vallejos, Virginia; Fraile, Manuel; Fernández-Sanmartín, Marco A; Pujol-Borrell, Ricardo; Puig-Domingo, Manel; Sanmartí, Anna; Martínez-Cáceres, Eva M

2013-04-01

One of the side effects of interferon-alpha therapy is interferon-induced thyroiditis (IIT). The role of lymphocyte subpopulations in IIT melanoma patients remains to be defined. Our objective was to assess different peripheral blood lymphocyte subpopulations, mainly regulatory T cells (Tregs), in melanoma patients who developed IIT. From 30 melanoma patients receiving high-dose interferon (HDI)-alpha 2b (IFN-α2b) treatment, those who developed IIT (IIT patients) were selected and compared with patients who did not develop IIT (Co-MM) and healthy controls (Co-H). Peripheral blood mononuclear cells were obtained before treatment (BT), mid-treatment (MT), end of treatment (ET), 24 weeks post-treatment and at appearance of IIT (TT). Nine patients developed IIT (30%): four Hashimoto's thyroiditis and five destructive thyroiditis. An increase in Tregs was observed in both melanoma groups during HDI treatment. A decrease in CD3(+) , NKT lymphocyte subpopulations and Bcl2 expression on B cells was also observed in both groups. However, no changes were observed in the percentage of CD4(+) , CD8(+) , CD3(+) γδ(+) , CD19(+) , transitional B cells (CD24(high) CD38(high) CD19(+) CD27(-) ), natural killer (NK), invariant NKT (iNKT) lymphocytes and Th1/Th2 balance when BT was compared with ET. At TT, IIT patients had a higher Tregs percentage than Co-MM (P = 0·012) and Co-H (P = 0·004), a higher iNKT percentage than Co-MM (P = 0·011), a higher transitional B cells percentage than Co-H (P = 0·015), a lower CD3(+) percentage than Co-H (P = 0·001) and a lower Bcl2 expression on B cells than Co-H (P < 0·001). Our results point to the immunomodulatory effects of IFN-α on different lymphocyte subpopulations and a possible role of Tregs in melanoma patients who developed IIT. © 2012 Blackwell Publishing Ltd.

Chemokine receptor CXCR6-dependent hepatic NK T Cell accumulation promotes inflammation and liver fibrosis.

PubMed

Wehr, Alexander; Baeck, Christer; Heymann, Felix; Niemietz, Patricia Maria; Hammerich, Linda; Martin, Christian; Zimmermann, Henning W; Pack, Oliver; Gassler, Nikolaus; Hittatiya, Kanishka; Ludwig, Andreas; Luedde, Tom; Trautwein, Christian; Tacke, Frank

2013-05-15

Chronic liver injury characteristically results in hepatic inflammation, which represents a prerequisite for organ fibrosis. Although NKT cells are abundantly present in liver and involved in hepatic inflammation, molecular mechanisms of their recruitment in liver fibrosis remained elusive. We hypothesized that chemokine receptor CXCR6 and its ligand CXCL16 control NKT cell migration and functionality in liver fibrosis. In patients with chronic liver diseases (n = 58), CXCR6 and CXCL16 expression was intrahepatically upregulated compared with controls. In murine liver, Cxcl16 was strongly expressed by endothelium and macrophages, whereas lymphocyte populations (NKT, NK, CD4 T, CD8 T cells) expressed CXCR6. Intravital two-photon microscopy imaging of Cxcr6(+/gfp) and Cxcr6(gfp/gfp) mice and chemotaxis studies in vitro revealed that CXCR6 specifically controls hepatic NKT cell accumulation during the early response upon experimental liver damage. Hepatic invariant NKT cells expressed distinct proinflammatory cytokines including IFN-γ and IL-4 upon injury. CXCR6-deficient mice were protected from liver fibrosis progression in two independent experimental models. Macrophage infiltration and protein levels of inflammatory cytokines IFN-γ, TNF-α, and IL-4 were also reduced in fibrotic livers of Cxcr6(-/-) mice, corroborating that hepatic NKT cells provide essential cytokine signals perpetuating hepatic inflammation and fibrogenesis. Adoptive transfer of NKT cells, but not CD4 T cells, isolated from wild type livers restored hepatic fibrosis in Cxcr6(-/-) mice upon experimental steatohepatitis. Our results demonstrate that hepatic NKT cells accumulate CXCR6-dependent early upon injury, thereby accentuating the inflammatory response in the liver and promoting hepatic fibrogenesis. Interfering with CXCR6/CXCL16 might therefore bear therapeutic potential in liver fibrosis.

STAT3 is a critical cell-intrinsic regulator of human unconventional T cell numbers and function

PubMed Central

Wilson, Robert P.; Ives, Megan L.; Rao, Geetha; Lau, Anthony; Payne, Kathryn; Kobayashi, Masao; Arkwright, Peter D.; Peake, Jane; Wong, Melanie; Adelstein, Stephen; Smart, Joanne M.; French, Martyn A.; Fulcher, David A.; Picard, Capucine; Bustamante, Jacinta; Boisson-Dupuis, Stephanie; Gray, Paul; Stepensky, Polina; Warnatz, Klaus; Freeman, Alexandra F.; Rossjohn, Jamie; McCluskey, James; Holland, Steven M.; Casanova, Jean-Laurent; Uzel, Gulbu; Ma, Cindy S.

2015-01-01

Unconventional T cells such as γδ T cells, natural killer T cells (NKT cells) and mucosal-associated invariant T cells (MAIT cells) are a major component of the immune system; however, the cytokine signaling pathways that control their development and function in humans are unknown. Primary immunodeficiencies caused by single gene mutations provide a unique opportunity to investigate the role of specific molecules in regulating human lymphocyte development and function. We found that individuals with loss-of-function mutations in STAT3 had reduced numbers of peripheral blood MAIT and NKT but not γδ T cells. Analysis of STAT3 mosaic individuals revealed that this effect was cell intrinsic. Surprisingly, the residual STAT3-deficient MAIT cells expressed normal levels of the transcription factor RORγt. Despite this, they displayed a deficiency in secretion of IL-17A and IL-17F, but were able to secrete normal levels of cytokines such as IFNγ and TNF. The deficiency in MAIT and NKT cells in STAT3-deficient patients was mirrored by loss-of-function mutations in IL12RB1 and IL21R, respectively. Thus, these results reveal for the first time the essential role of STAT3 signaling downstream of IL-23R and IL-21R in controlling human MAIT and NKT cell numbers. PMID:25941256

Retinoic Acid Modulates Interferon-γ Production by Hepatic Natural Killer T Cells via Phosphatase 2A and the Extracellular Signal-Regulated Kinase Pathway

PubMed Central

Chang, Heng-Kwei

2015-01-01

Retinoic acid (RA), an active metabolite converted from vitamin A, plays an active role in immune function, such as defending against infections and immune regulation. Although RA affects various types of immune cells, including antigen-presenting cells, B lymphocytes, and T lymphocytes, whether it affects natural killer T (NKT) cells remain unknown. In this study, we found that RA decreased interferon (IFN)-γ production by activated NKT cells through T-cell receptor (TCR) and CD28. We also found that RA reduced extracellular signal-regulated kinase (ERK) phosphorylation, but increased phosphatase 2A (PP2A) activity in TCR/CD28-stimulated NKT cells. The increased PP2A activity, at least partly, contributed to the reduction of ERK phosphorylation. Since inhibition of ERK activation decreases IFN-γ production by TCR/CD28-stimulated NKT cells, RA may downregulate IFN-γ production by TCR/CD28-stimulated NKT cells through the PP2A-ERK pathway. Our results demonstrated a novel function of RA in modulating the IFN-γ expression by activated NKT cells. PMID:25343668

Invariant Natural Killer T Cells Are Pathogenic in the HLA-DR4-Transgenic Humanized Mouse Model of Toxic Shock Syndrome and Can Be Targeted to Reduce Morbidity.

PubMed

Szabo, Peter A; Rudak, Patrick T; Choi, Joshua; Xu, Stacey X; Schaub, Robert; Singh, Bhagirath; McCormick, John K; Haeryfar, S M Mansour

2017-03-01

During toxic shock syndrome (TSS), bacterial superantigens trigger a polyclonal T -cell response leading to a potentially catastrophic "cytokine storm". Whether innate-like invariant natural killer T (iNKT) cells, with remarkable immunomodulatory properties, participate in TSS is unclear. Using genetic and cell depletion approaches, we generated iNKT cell-deficient, superantigen-sensitive HLA-DR4-transgenic (DR4tg) mice, which were compared with their iNKT-sufficient counterparts for responsiveness to staphylococcal enterotoxin B (SEB). Both approaches indicate that iNKT cells are pathogenic in TSS. Importantly, treating DR4tg mice with a TH2-polarizing glycolipid agonist of iNKT cells reduced SEB-inflicted morbidity/mortality. Therefore, iNKT cells may constitute an attractive therapeutic target in superantigen-mediated illnesses. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Natural Killer T Cell Activation Protects Mice Against Experimental Autoimmune Encephalomyelitis

PubMed Central

Singh, Avneesh K.; Wilson, Michael T.; Hong, Seokmann; Olivares-Villagómez, Danyvid; Du, Caigan; Stanic, Aleksandar K.; Joyce, Sebastian; Sriram, Subramaniam; Koezuka, Yasuhiko; Van Kaer, Luc

2001-01-01

Experimental autoimmune encephalomyelitis (EAE) serves as a prototypic model for T cell–mediated autoimmunity. Vα14 natural killer T (NKT) cells are a subset of T lymphocytes that recognize glycolipid antigens presented by the nonpolymorphic major histocompatibility complex (MHC) class I–like protein CD1d. Here, we show that activation of Vα14 NKT cells by the glycosphingolipid α-galactosylceramide (α-GalCer) protects susceptible mice against EAE. β-GalCer, which binds CD1d but is not recognized by NKT cells, failed to protect mice against EAE. Furthermore, α-GalCer was unable to protect CD1d knockout (KO) mice against EAE, indicating the requirement for an intact CD1d antigen presentation pathway. Protection of disease conferred by α-GalCer correlated with its ability to suppress myelin antigen-specific Th1 responses and/or to promote myelin antigen-specific Th2 cell responses. α-GalCer was unable to protect IL-4 KO and IL-10 KO mice against EAE, indicating a critical role for both of these cytokines. Because recognition of α-GalCer by NKT cells is phylogenetically conserved, our findings have identified NKT cells as novel target cells for treatment of inflammatory diseases of the central nervous system. PMID:11748281

Critical Role for Very-Long Chain Sphingolipids in Invariant Natural Killer T Cell Development and Homeostasis.

PubMed

Saroha, Ashish; Pewzner-Jung, Yael; Ferreira, Natalia S; Sharma, Piyush; Jouan, Youenn; Kelly, Samuel L; Feldmesser, Ester; Merrill, Alfred H; Trottein, François; Paget, Christophe; Lang, Karl S; Futerman, Anthony H

2017-01-01

The role of sphingolipids (SLs) in the immune system has come under increasing scrutiny recently due to the emerging contributions that these important membrane components play in regulating a variety of immunological processes. The acyl chain length of SLs appears particularly critical in determining SL function. Here, we show a role for very-long acyl chain SLs (VLC-SLs) in invariant natural killer T ( i NKT) cell maturation in the thymus and homeostasis in the liver. Ceramide synthase 2-null mice, which lack VLC-SLs, were susceptible to a hepatotropic strain of lymphocytic choriomeningitis virus, which is due to a reduction in the number of i NKT cells. Bone marrow chimera experiments indicated that hematopoietic-derived VLC-SLs are essential for maturation of i NKT cells in the thymus, whereas parenchymal-derived VLC-SLs are crucial for i NKT cell survival and maintenance in the liver. Our findings suggest a critical role for VLC-SL in i NKT cell physiology.

Dual Modifications of α-Galactosylceramide Synergize to Promote Activation of Human Invariant Natural Killer T Cells and Stimulate Anti-tumor Immunity.

PubMed

Chennamadhavuni, Divya; Saavedra-Avila, Noemi Alejandra; Carreño, Leandro J; Guberman-Pfeffer, Matthew J; Arora, Pooja; Yongqing, Tang; Koay, Hui-Fern; Godfrey, Dale I; Keshipeddy, Santosh; Richardson, Stewart K; Sundararaj, Srinivasan; Lo, Jae Ho; Wen, Xiangshu; Gascón, José A; Yuan, Weiming; Rossjohn, Jamie; Le Nours, Jérôme; Porcelli, Steven A; Howell, Amy R

2018-05-17

Glycosylceramides that activate CD1d-restricted invariant natural killer T (iNKT) cells have potential therapeutic applications for augmenting immune responses against cancer and infections. Previous studies using mouse models identified sphinganine variants of α-galactosylceramide as promising iNKT cell activators that stimulate cytokine responses with a strongly proinflammatory bias. However, the activities of sphinganine variants in mice have generally not translated well to studies of human iNKT cell responses. Here, we show that strongly proinflammatory and anti-tumor iNKT cell responses were achieved in mice by a variant of α-galactosylceramide that combines a sphinganine base with a hydrocinnamoyl ester on C6″ of the sugar. Importantly, the activities observed with this variant were largely preserved for human iNKT cell responses. Structural and in silico modeling studies provided a mechanistic basis for these findings and suggested basic principles for capturing useful properties of sphinganine analogs of synthetic iNKT cell activators in the design of immunotherapeutic agents. Copyright © 2018 Elsevier Ltd. All rights reserved.

Testosterone Increases Susceptibility to Amebic Liver Abscess in Mice and Mediates Inhibition of IFNγ Secretion in Natural Killer T Cells

PubMed Central

Lotter, Hannelore; Helk, Elena; Bernin, Hannah; Jacobs, Thomas; Prehn, Cornelia; Adamski, Jerzy; González-Roldán, Nestor; Holst, Otto; Tannich, Egbert

2013-01-01

Amebic liver abscess (ALA), a parasitic disease due to infection with the protozoan Entamoeba histolytica, occurs age and gender dependent with strong preferences for adult males. Using a mouse model for ALA with a similar male bias for the disease, we have investigated the role of female and male sexual hormones and provide evidence for a strong contribution of testosterone. Removal of testosterone by orchiectomy significantly reduced sizes of abscesses in male mice, while substitution of testosterone increased development of ALA in female mice. Activation of natural killer T (NKT) cells, which are known to be important for the control of ALA, is influenced by testosterone. Specifically activated NKT cells isolated from female mice produce more IFNγ compared to NKT cells derived from male mice. This high level production of IFNγ in female derived NKT cells was inhibited by testosterone substitution, while the IFNγ production in male derived NKT cells was increased by orchiectomy. Gender dependent differences were not a result of differences in the total number of NKT cells, but a result of a higher activation potential for the CD4− NKT cell subpopulation in female mice. Taken together, we conclude that the hormone status of the host, in particular the testosterone level, determines susceptibility to ALA at least in a mouse model of the disease. PMID:23424637

Chimeric antigen receptor-engineered natural killer and natural killer T cells for cancer immunotherapy.

PubMed

Bollino, Dominique; Webb, Tonya J

2017-09-01

Natural killer (NK) cells of the innate immune system and natural killer T (NKT) cells, which have roles in both the innate and adaptive responses, are unique lymphocyte subsets that have similarities in their functions and phenotypes. Both cell types can rapidly respond to the presence of tumor cells and participate in immune surveillance and antitumor immune responses. This has incited interest in the development of novel cancer therapeutics based on NK and NKT cell manipulation. Chimeric antigen receptors (CARs), generated through the fusion of an antigen-binding region of a monoclonal antibody or other ligand to intracellular signaling domains, can enhance lymphocyte targeting and activation toward diverse malignancies. Most of the CAR studies have focused on their expression in T cells; however, the functional heterogeneity of CAR T cells limits their therapeutic potential and is associated with toxicity. CAR-modified NK and NKT cells are becoming more prevalent because they provide a method to direct these cells more specifically to target cancer cells, with less risk of adverse effects. This review will outline current NK and NKT cell CAR constructs and how they compare to conventional CAR T cells, and discuss future modifications that can be explored to advance adoptive cell transfer of NK and NKT cells. Copyright © 2017 Elsevier Inc. All rights reserved.

Bacterial lysate increases the percentage of natural killer T cells in peripheral blood and alleviates asthma in children.

PubMed

Lu, Yanming; Li, Yaqin; Xu, Lingyun; Xia, Min; Cao, Lanfang

2015-01-01

To assess the efficacy of conventional treatment combined with bacterial lysate [OM-85 Broncho-Vaxom (BV)] in the prevention of asthma in children as well as its influence on the number of natural killer T (NKT) cells and their cytokine production. Sixty children diagnosed with asthma were divided into either a BV-treated group (with oral OM-85 BV) or a conventional inhaled corticosteroid (ICS) group. The numbers of NKT cells and CD4+ NKT cells were measured in the peripheral blood by flow cytometry. The levels of IFN-γ, IL-4, and IL-10 after the blood cells had been cultured with an NKT cell agonist were detected by ELISA. After therapy, asthma attacks were significantly decreased compared with before therapy in both groups. However, after therapy, respiratory tract infections were reduced compared with before therapy in the BV-treated group only. Additionally, the frequency of asthma attacks and use of antibiotics in the BV-treated group were lower than in the ICS group. With BV treatment, the numbers of peripheral blood NKT cells and CD4+ NKT cells were higher after therapy than before therapy. After therapy, the ratio of IFN-γ/IL-4 and IL-10 levels were increased in the BV-treated group, whereas IL-4 was reduced in the BV-treated group compared with the ICS group. BV combined with conventional asthma treatment can prevent recurrent respiratory tract infections and suppress the severity of asthma attacks, possibly by altering the rates and cytokines of NKT cells. © 2015 S. Karger AG, Basel

Superoxide produced by Kupffer cells is an essential effector in concanavalin A-induced hepatitis in mice.

PubMed

Nakashima, Hiroyuki; Kinoshita, Manabu; Nakashima, Masahiro; Habu, Yoshiko; Shono, Satoshi; Uchida, Takefumi; Shinomiya, Nariyoshi; Seki, Shuhji

2008-12-01

Although concanavalin A (Con-A)-induced experimental hepatitis is thought to be induced by activated T cells, natural killer T (NKT) cells, and cytokines, precise mechanisms are still unknown. In the current study, we investigated the roles of Kupffer cells, NKT cells, FasL, tumor necrosis factor (TNF), and superoxide in Con-A hepatitis in C57BL/6 mice. Removal of Kupffer cells using gadolinium chloride (GdCl(3)) from the liver completely inhibited Con-A hepatitis, whereas increased serum TNF and IFN-gamma levels were not inhibited at all. Unexpectedly, anti-FasL antibody pretreatment did not inhibit Con-A hepatitis, whereas it inhibited hepatic injury induced by a synthetic ligand of NKT cells, alpha-galactosylceramide. Furthermore, GdCl(3) pretreatment changed neither the activation-induced down-regulation of NK1.1 antigens as well as T cell receptors of NKT cells nor the increased expression of the CD69 activation antigen of hepatic T cells. CD68(+) Kupffer cells greatly increased in proportion in the early phase after Con-A injection; this increase was abrogated by GdCl(3) pretreatment. Anti-TNF antibody (Ab) pretreatment did not inhibit the increase of Kupffer cells, but it effectively suppressed superoxide/reactive oxygen production from Kupffer cells and the resulting hepatic injury. Conversely, depletion of NKT cells in mice by NK1.1 Ab pretreatment did suppress both the increase of CD68(+) Kupffer cells and Con-A hepatitis. Consistently, the diminution of oxygen radicals produced by Kupffer cells by use of free radical scavengers greatly inhibited Con-A hepatitis without suppressing cytokine production. However, adoptive transfer experiments also indicate that a close interaction/cooperation of Kupffer cells with NKT cells is essential for Con-A hepatitis. Superoxide produced by Kupffer cells may be the essential effector in Con-A hepatitis, and TNF and NKT cells support their activation and superoxide production.

Human Invariant Natural Killer T cells possess immune-modulating functions during Aspergillus infection.

PubMed

Beitzen-Heineke, Antonia; Bouzani, Maria; Schmitt, Anna-Lena; Kurzai, Oliver; Hünniger, Kerstin; Einsele, Hermann; Loeffler, Juergen

2016-02-01

Aspergillus fumigatus is the most common cause for invasive fungal infections, a disease associated with high mortality in immune-compromised patients. CD1d-restricted invariant natural killer T (iNKT) cells compose a small subset of T cells known to impact the immune response toward various infectious pathogens. To investigate the role of human iNKT cells during A. fumigatus infection, we studied their activation as determined by CD69 expression and cytokine production in response to distinct fungal morphotypes in the presence of different CD1d(+) antigen presenting cells using flow cytometry and multiplex enzyme-linked immunosorbent assay (ELISA). Among CD1d(+) subpopulations, CD1d(+)CD1c(+) mDCs showed the highest potential to activate iNKT cells on a per cell basis. The presence of A. fumigatus decreased this effect of CD1d(+)CD1c(+) mDCs on iNKT cells and led to reduced secretion of TNF-α, G-CSF and RANTES. Production of other Th1 and Th2 cytokines was not affected by the fungus, suggesting an immune-modulating function for human iNKT cells during A. fumigatus infection. © The Author 2015. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Human Invariant Natural Killer T Cells Respond to Antigen-Presenting Cells Exposed to Lipids from Olea europaea Pollen.

PubMed

Abos Gracia, Beatriz; López Relaño, Juan; Revilla, Ana; Castro, Lourdes; Villalba, Mayte; Martín Adrados, Beatriz; Regueiro, Jose Ramon; Fernández-Malavé, Edgar; Martínez Naves, Eduardo; Gómez Del Moral, Manuel

2017-01-01

Allergic sensitization might be influenced by the lipids present in allergens, which can be recognized by natural killer T (NKT) cells on antigen-presenting cells (APCs). The aim of this study was to analyze the effect of olive pollen lipids in human APCs, including monocytes as well as monocyte-derived macrophages (Mϕ) and dendritic cells (DCs). Lipids were extracted from olive (Olea europaea) pollen grains. Invariant (i)NKT cells, monocytes, Mϕ, and DCs were obtained from buffy coats of healthy blood donors, and their cell phenotype was determined by flow cytometry. iNKT cytotoxicity was measured using a lactate dehydrogenase assay. Gene expression of CD1A and CD1D was performed by RT-PCR, and the production of IL-6, IL-10, IL-12, and TNF-α cytokines by monocytes, Mϕ, and DCs was measured by ELISA. Our results showed that monocytes and monocyte-derived Mϕ treated with olive pollen lipids strongly activate iNKT cells. We observed several phenotypic modifications in the APCs upon exposure to pollen-derived lipids. Both Mϕ and monocytes treated with olive pollen lipids showed an increase in CD1D gene expression, whereas upregulation of cell surface CD1d protein occurred only in Mϕ. Furthermore, DCs differentiated in the presence of human serum enhance their surface CD1d expression when exposed to olive pollen lipids. Finally, olive pollen lipids were able to stimulate the production of IL-6 but downregulated the production of lipopolysaccharide- induced IL-10 by Mϕ. Olive pollen lipids alter the phenotype of monocytes, Mϕ, and DCs, resulting in the activation of NKT cells, which have the potential to influence allergic immune responses. © 2017 S. Karger AG, Basel.

Natural killer T cell facilitated engraftment of rat skin but not islet xenografts in mice.

PubMed

Gordon, Ethel J; Kelkar, Vinaya

2009-01-01

We have studied cellular components required for xenograft survival mediated by anti-CD154 monoclonal antibody (mAb) and a transfusion of donor spleen cells and found that the elimination of CD4(+) but not CD8(+) cells significantly improves graft survival. A contribution of other cellular components, such as natural killer (NK) cells and natural killer T (NKT) cells, for costimulation blockade-induced xenograft survival has not been clearly defined. We therefore tested the hypothesis that NK or NKT cells would promote rat islet and skin xenograft acceptance in mice. Lewis rat islets or skin was transplanted into wild type B6 mice or into B6 mice that were Jalpha18(null), CD1(null), or beta2 microglobulin (beta2M)(null) NK 1.1 depleted, or perforin(null). Graft recipients were pretreated with an infusion of donor derived spleen cells and a brief course of anti-CD154 mAb treatments. Additional groups received mAb or cells only. We first observed that the depletion of NK1.1 cells does not significantly interfere with graft survival in C57BL/6 (B6) mice. We used NKT cell deficient B6 mice to test the hypothesis that NKT cells are involved in islet and skin xenograft survival in our model. These mice bear a null mutation in the gene for the Jalpha18 component of the T-cell receptor. The component is uniquely associated with NKT cells. We found no difference in islet xenograft survival between Jalpha18(null) and wild type B6 mice. In contrast, median skin graft survival appeared shorter in Jalpha18(null) recipients. These data imply a role for Jalpha18(+) NKT cells in skin xenograft survival in treated mice. In order to confirm this inference, we tested skin xenograft survival in B6 CD1(null) mice because NKT cells are CD1 restricted. Results of these trials demonstrate that the absence of CD1(+) cells adversely affects rat skin graft survival. An additional assay in beta2M(null) mice demonstrated a requirement for major histocompatibility complex (MHC) class I

Lysophospholipid presentation by CD1d and recognition by a human Natural Killer T-cell receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

López-Sagaseta, Jacinto; Sibener, Leah V.; Kung, Jennifer E.

2014-10-02

Invariant Natural Killer T (iNKT) cells use highly restricted {alpha}{beta} T cell receptors (TCRs) to probe the repertoire of lipids presented by CD1d molecules. Here, we describe our studies of lysophosphatidylcholine (LPC) presentation by human CD1d and its recognition by a native, LPC-specific iNKT TCR. Human CD1d presenting LPC adopts an altered conformation from that of CD1d presenting glycolipid antigens, with a shifted {alpha}1 helix resulting in an open A pocket. Binding of the iNKT TCR requires a 7-{angstrom} displacement of the LPC headgroup but stabilizes the CD1d-LPC complex in a closed conformation. The iNKT TCR CDR loop footprint onmore » CD1d-LPC is anchored by the conserved positioning of the CDR3{alpha} loop, whereas the remaining CDR loops are shifted, due in part to amino-acid differences in the CDR3{beta} and J{beta} segment used by this iNKT TCR. These findings provide insight into how lysophospholipids are presented by human CD1d molecules and how this complex is recognized by some, but not all, human iNKT cells.« less

Excessive interferon-α signaling in autoimmunity alters glycosphingolipid processing in B cells.

PubMed

Tan, Andy Hee-Meng; Sanny, Arleen; Ng, Sze-Wai; Ho, Ying-Swan; Basri, Nurhidayah; Lee, Alison Ping; Lam, Kong-Peng

2018-05-01

Excessive interferon-α (IFN-α) production by innate immune cells is a hallmark of autoimmune diseases. What other cell type secretes IFN-α and how IFN-α affects immune cell metabolism and homeostasis in autoimmunity are largely unclear. Here, we report that autoimmune B cells, arising from two different B cell-specific genetic lesions in mice, secrete IFN-α. In addition, IFN-α, found in abundance in autoimmunity, elicited profound changes in the B cell lipidome, increasing their expression of glycosphingolipids (GSLs) and leading to their CD1d-mediated depletion of iNKT cells in vitro and in vivo. IFN-α receptor blockade could reverse the loss of iNKT cells. Excessive stimulation of B cells with IFN-α altered the expression of enzymes that catalyze critical steps in GSL processing, increasing the expressions of glucosylceramide synthase (GCS) and globotrihexosylceramide synthase (Gb3S) but decreasing that of α-galactosidase A (α-galA). Inhibiting GCS or restoring α-galA expression prevented iNKT depletion by IFN-α-activated B cells. Taken together, our work indicated that excessive IFN-α perturbs GSL metabolism in B cells which in turn adversely affects iNKT homeostasis. Copyright © 2017 Elsevier Ltd. All rights reserved.

IL13Rα2 expression identifies tissue-resident IL-22 producing PLZF+ innate T cells in the human liver.

PubMed

Paquin-Proulx, Dominic; Greenspun, Benjamin C; Pasquet, Lise; Strunz, Benedikt; Aleman, Soo; Falconer, Karolin; Terabe, Masaki; Berzofsky, Jay A; Sandberg, Johan K; Melum, Espen; Nixon, Douglas F; Björkström, Niklas K

2018-04-20

Innate lymphocytes are selectively enriched in the liver where they have important roles in liver immunology. Murine studies have shown that type I NKT cells can promote liver inflammation whereas type II NKT cells have an anti-inflammatory role. In humans, type II NKT cells were found to accumulate in the gut during inflammation and IL13Rα2 was proposed as a marker for these cells. In the human liver, less is known about type I and II NKT cells. Here, we studied the phenotype and function of human liver T cells expressing IL13Rα2. We found that IL13Rα2 was expressed by around 1% of liver resident memory T cells but not on circulating T cells. In support of their innate-like T cell character, the IL13Rα2 + T cells had higher expression of PLZF compared to IL13Rα2 - T cells and possessed the capacity to produce IL-22. However, only a minority of human liver sulfatide-reactive type II NKT cells expressed IL13Rα2. Collectively, these findings suggest that IL13Rα2 identifies tissue-resident intrahepatic T cells with innate characteristics and the capacity to produce IL-22. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Generation of Novel Traj18-Deficient Mice Lacking Vα14 Natural Killer T Cells with an Undisturbed T Cell Receptor α-Chain Repertoire.

PubMed

Dashtsoodol, Nyambayar; Shigeura, Tomokuni; Ozawa, Ritsuko; Harada, Michishige; Kojo, Satoshi; Watanabe, Takashi; Koseki, Haruhiko; Nakayama, Manabu; Ohara, Osamu; Taniguchi, Masaru

2016-01-01

Invariant Vα14 natural killer T (NKT) cells, characterized by the expression of a single invariant T cell receptor (TCR) α chain encoded by rearranged Trav11 (Vα14)-Traj18 (Jα18) gene segments in mice, and TRAV10 (Vα24)-TRAJ18 (Jα18) in humans, mediate adjuvant effects to activate various effector cell types in both innate and adaptive immune systems that facilitates the potent antitumor effects. It was recently reported that the Jα18-deficient mouse described by our group in 1997 harbors perturbed TCRα repertoire, which raised concerns regarding the validity of some of the experimental conclusions that have been made using this mouse line. To resolve this concern, we generated a novel Traj18-deficient mouse line by specifically targeting the Traj18 gene segment using Cre-Lox approach. Here we showed the newly generated Traj18-deficient mouse has, apart from the absence of Traj18, an undisturbed TCRα chain repertoire by using next generation sequencing and by detecting normal generation of Vα19Jα33 expressing mucosal associated invariant T cells, whose development was abrogated in the originally described Jα18-KO mice. We also demonstrated here the definitive requirement for NKT cells in the protection against tumors and their potent adjuvant effects on antigen-specific CD8 T cells.

Cytokeratin 19 (KRT19) has a Role in the Reprogramming of Cancer Stem Cell-Like Cells to Less Aggressive and More Drug-Sensitive Cells.

PubMed

Saha, Subbroto Kumar; Kim, Kyeongseok; Yang, Gwang-Mo; Choi, Hye Yeon; Cho, Ssang-Goo

2018-05-09

Cytokeratin 19 ( KRT19 ) is a cytoplasmic intermediate filament protein, which is responsible for structural rigidity and multipurpose scaffolds. In several cancers, KRT19 is overexpressed and may play a crucial role in tumorigenic transformation. In our previous study, we revealed the role of KRT19 as signaling component which mediated Wnt/NOTCH crosstalk through NUMB transcription in breast cancer. Here, we investigated the function of KRT19 in cancer reprogramming and drug resistance in breast cancer cells. We found that expression of KRT19 was attenuated in several patients-derived breast cancer tissues and patients with a low expression of KRT19 were significantly correlated with poor prognosis in breast cancer patients. Consistently, highly aggressive and drug-resistant breast cancer patient-derived cancer stem cell-like cells (konkuk university-cancer stem cell-like cell (KU-CSLCs)) displayed higher expression of cancer stem cell (CSC) markers, including ALDH1 , CXCR4 , and CD133 , but a much lower expression of KRT19 than that is seen in highly aggressive triple negative breast cancer MDA-MB231 cells. Moreover, we revealed that the knockdown of KRT19 in MDA-MB231 cells led to an enhancement of cancer properties, such as cell proliferation, sphere formation, migration, and drug resistance, while the overexpression of KRT19 in KU-CSLCs resulted in the significant attenuation of cancer properties. KRT19 regulated cancer stem cell reprogramming by modulating the expression of cancer stem cell markers ( ALDH1 , CXCR4 , and CD133 ), as well as the phosphorylation of Src and GSK3β (Tyr216). Therefore, our data may imply that the modulation of KRT19 expression could be involved in cancer stem cell reprogramming and drug sensitivity, which might have clinical implications for cancer or cancer stem cell treatment.

Cytokeratin 19 (KRT19) has a Role in the Reprogramming of Cancer Stem Cell-Like Cells to Less Aggressive and More Drug-Sensitive Cells

PubMed Central

Kim, Kyeongseok; Yang, Gwang-Mo; Choi, Hye Yeon

2018-01-01

Cytokeratin 19 (KRT19) is a cytoplasmic intermediate filament protein, which is responsible for structural rigidity and multipurpose scaffolds. In several cancers, KRT19 is overexpressed and may play a crucial role in tumorigenic transformation. In our previous study, we revealed the role of KRT19 as signaling component which mediated Wnt/NOTCH crosstalk through NUMB transcription in breast cancer. Here, we investigated the function of KRT19 in cancer reprogramming and drug resistance in breast cancer cells. We found that expression of KRT19 was attenuated in several patients-derived breast cancer tissues and patients with a low expression of KRT19 were significantly correlated with poor prognosis in breast cancer patients. Consistently, highly aggressive and drug-resistant breast cancer patient-derived cancer stem cell-like cells (konkuk university-cancer stem cell-like cell (KU-CSLCs)) displayed higher expression of cancer stem cell (CSC) markers, including ALDH1, CXCR4, and CD133, but a much lower expression of KRT19 than that is seen in highly aggressive triple negative breast cancer MDA-MB231 cells. Moreover, we revealed that the knockdown of KRT19 in MDA-MB231 cells led to an enhancement of cancer properties, such as cell proliferation, sphere formation, migration, and drug resistance, while the overexpression of KRT19 in KU-CSLCs resulted in the significant attenuation of cancer properties. KRT19 regulated cancer stem cell reprogramming by modulating the expression of cancer stem cell markers (ALDH1, CXCR4, and CD133), as well as the phosphorylation of Src and GSK3β (Tyr216). Therefore, our data may imply that the modulation of KRT19 expression could be involved in cancer stem cell reprogramming and drug sensitivity, which might have clinical implications for cancer or cancer stem cell treatment. PMID:29747452

Regulatory T, natural killer T and γδ T cells in multiple sclerosis and chronic fatigue syndrome/myalgic encephalomyelitis: a comparison.

PubMed

Ramos, Sandra; Brenu, Ekua; Broadley, Simon; Kwiatek, Richard; Ng, Jennifer; Nguyen, Thao; Freeman, Susan; Staines, Donald; Marshall-Gradisnik, Sonya

2016-12-01

Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME), and Multiple Sclerosis (MS) may share some similarities in relation to reduced NK cell activity. It is likely that other cells such as regulatory T (Tregs), invariant Natural Killer T (iNKT) and gamma delta T (γδ T) cells may also be dysregulated in CFS/ME and MS. To evaluate and compare specific immune regulatory cells of patients with CFS/ME, patients with MS and healthy controls. Sixty three volunteers were included in this study: 24 were CFS/ME patients, 11 were MS patients and 27 were healthy controls. Blood samples were obtained from all participants for flow cytometry analysis of iNKT cells, Tregs and γδ T cell phenotypes. We observed a significant increase in Tregs in the CFS/ME group (p≤0.05) compared to the healthy control group. Total γδ and γδ2 T cells were significantly reduced in MS patients in comparison with the healthy control group. Conversely, CD4+iNKT percentage of iNKT, was significantly increased in the CFS/ME group compared with healthy controls and the double-negative iNKT percentage of iNKT significantly decreased compared with the healthy control group. This study has not identified any immunological disturbances that are common in both MS and CFS/ME patients. However, the differential expression of cell types between the conditions investigated suggests different pathways of disease. These differences need to be explored in further studies.

Thiopurine use associated with reduced B and natural killer cells in inflammatory bowel disease

PubMed Central

Lord, James D; Shows, Donna M

2017-01-01

AIM To identify which blood and mucosal lymphocyte populations are specifically depleted by thiopurine use in vivo. METHODS The thiopurines azathioprine and 6-mercaptopurine have been a mainstay of inflammatory bowel disease (IBD) therapy for decades, but their mechanism of action in vivo remains obscure. Although thiopurines are lymphotoxic at high doses, and have been reported to cause T cell apoptosis in vitro, their ability to control IBD at lower doses suggests that they may selectively deplete particular lymphocyte populations. Blood cells from 19 IBD patients on a thiopurine, 19 IBD patients not on a thiopurine, and 38 matched healthy control subjects were analyzed by multiple multi-color flow cytometry panels to quantify the immune cell subsets contained therein, both as a percent of cells, and as an absolute cell count. Similar analyses were performed on colon biopsies from 17 IBD patients on a thiopurine, 17 IBD patients not on a thiopurine, and 49 healthy screening colonoscopy recipients. RESULTS Complete blood counts revealed lower lymphocyte, but not monocyte or granulocyte, counts in IBD patients who were taking thiopurines at the time of sampling. This reduction was restricted to CD3-negative lymphocytes, wherein both natural killer (NK) and B cells were significantly reduced among thiopurine recipients. Among CD19+ B cells, the transitional B cells were particularly depleted, being nearly absent in both blood and colon biopsies of thiopurine recipients. No differences were associated with thiopurine use in CD8+ T cells, mucosa-associated invariant T (MAIT) cells, invariant natural killer T (iNKT) cells, gamma/delta T cells, Th1, Th17, regulatory T cells (Tregs) or naïve CD4+ T cells. However, patients with IBD had significantly more circulating FOXP3+, Helios+ Tregs and fewer iNKT and MAIT cells than healthy controls. CONCLUSION Thiopurine use is associated with reduced B and NK cell, but not T cell, subpopulations in the blood of IBD patients

Dendritic cells tolerized with adenosine A2AR agonist attenuate acute kidney injury

PubMed Central

Li, Li; Huang, Liping; Ye, Hong; Song, Steven P.; Bajwa, Amandeep; Lee, Sang Ju; Moser, Emily K.; Jaworska, Katarzyna; Kinsey, Gilbert R.; Day, Yuan J.; Linden, Joel; Lobo, Peter I.; Rosin, Diane L.; Okusa, Mark D.

2012-01-01

DC-mediated NKT cell activation is critical in initiating the immune response following kidney ischemia/reperfusion injury (IRI), which mimics human acute kidney injury (AKI). Adenosine is an important antiinflammatory molecule in tissue inflammation, and adenosine 2A receptor (A2AR) agonists protect kidneys from IRI through their actions on leukocytes. In this study, we showed that mice with A2AR-deficient DCs are more susceptible to kidney IRI and are not protected from injury by A2AR agonists. In addition, administration of DCs treated ex vivo with an A2AR agonist protected the kidneys of WT mice from IRI by suppressing NKT production of IFN-γ and by regulating DC costimulatory molecules that are important for NKT cell activation. A2AR agonists had no effect on DC antigen presentation or on Tregs. We conclude that ex vivo A2AR–induced tolerized DCs suppress NKT cell activation in vivo and provide a unique and potent cell-based strategy to attenuate organ IRI. PMID:23093781

Activation of Invariant Natural Killer T Cells Redirects the Inflammatory Response in Neonatal Sepsis.

PubMed

Bolognese, Alexandra C; Yang, Weng-Lang; Hansen, Laura W; Sharma, Archna; Nicastro, Jeffrey M; Coppa, Gene F; Wang, Ping

2018-01-01

Sepsis is the third leading cause of death in the neonatal population, due to susceptibility to infection conferred by immaturity of both the innate and adaptive components of the immune system. Invariant natural killer T (iNKT) cells are specialized adaptive immune cells that possess important innate-like characteristics and have not yet been well-studied in septic neonates. We hypothesized that iNKT cells would play an important role in mediating the neonatal immune response to sepsis. To study this, we subjected 5- to 7-day-old neonatal C57BL/6 mice to sepsis by intraperitoneal (i.p.) cecal slurry (CS) injection. Thirty hours prior to or immediately following sepsis induction, pups received i.p. injection of the iNKT stimulator KRN7000 (KRN, 0.2 µg/g) or vehicle. Ten hours after CS injection, blood and tissues were collected for various analyses. Thirty-hour pretreatment with KRN resulted in better outcomes in inflammation, lung injury, and survival, while immediate treatment with KRN resulted in worse outcomes compared to vehicle treatment. We further analyzed the activation status of neonatal iNKT cells for 30 h after KRN administration, and showed a peak in frequency of CD69 expression on iNKT cells and serum IFN-γ levels at 5 and 10 h, respectively. We then used CD1d knockout neonatal mice to demonstrate that KRN acts through the major histocompatibility complex-like molecule CD1d to improve outcomes in neonatal sepsis. Finally, we identified that KRN pretreatment exerts its protective effect by increasing systemic levels of TGF-β1. These findings support the importance of iNKT cells for prophylactic immunomodulation in neonates susceptible to sepsis.

Co-Expansion of Cytokine-Induced Killer Cells and Vγ9Vδ2 T Cells for CAR T-Cell Therapy

PubMed Central

Chen, Can; Tan, Wee-Kiat; Chi, Zhixia; Xu, Xue-Hu; Wang, Shu

2016-01-01

Gamma delta (γδ) T cells and cytokine-induced killer (CIK) cells, which are a heterogeneous population of T lymphocytes and natural killer T (NKT) cells, have been separately expanded ex vivo and shown to be capable of targeting and mediating cytotoxicity against various tumor cells in a major histocompatibility complex-unrestricted manner. However, the co-expansion and co-administration of these immune cells have not been explored. In this study we describe an efficient method to expand simultaneously both CIK and Vγ9Vδ2 T cells, termed as CIKZ cells, from human peripheral blood mononuclear cells (PBMCs) using Zometa, interferon-gamma (IFN-γ), interleukin 2 (IL-2), anti-CD3 antibody and engineered K562 feeder cells expressing CD64, CD137L and CD86. A 21-day culture of PBMCs with this method yielded nearly 20,000-fold expansion of CIKZ cells with γδ T cells making up over 20% of the expanded population. The expanded CIKZ cells exhibited antitumor cytotoxicity and could be modified to express anti-CD19 chimeric antigen receptor (CAR), anti-CEA CAR, and anti-HER2 CAR to enhance their specificity and cytotoxicity against CD19-, CEA-, or HER2-positive tumor cells. The tumor inhibitory activity of anti-CD19 CAR-modified CIKZ cells was further demonstrated in vivo in a Raji tumor mouse model. The findings herein substantiate the feasibility of co-expanding CIK and γδ cells for adoptive cellular immunotherapy applications such as CAR T-cell therapy against cancer. PMID:27598655

Role of Natural Killer T Cells In Immunogenic Chemotherapy for Breast Cancer

DTIC Science & Technology

2012-09-01

carcinomas, hematopoietic malignancies, and their metastases. This effect is mainly due to the synthesis of IFN-g by NKT cells and to the bystander...promote IFN-g synthesis by splenic gd T cells. In con- trast, combined addition of both cytokines induced IFN-g pro- duction by gd T cells. Taken...GalCer is mainly due to the rapid synthesis of IFN-g by type I NKT cells and the by- stander activation of both NK and CD8+ CTL (15, 16). Thus, we

19. Oblique, typical cell (south cells) from rear of cell; ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

19. Oblique, typical cell (south cells) from rear of cell; view to north, 65mm lens with electronic flash illumination. - Tule Lake Project Jail, Post Mile 44.85, State Route 139, Newell, Modoc County, CA

BMP Induction of Cardiogenesis in P19 Cells Requires Prior Cell-Cell Interaction(s)

PubMed Central

ANGELLO, JOHN C.; KAESTNER, STEFANIE; WELIKSON, ROBERT E.; BUSKIN, JEAN N.; HAUSCHKA, STEPHEN D.

2008-01-01

Mouse P19 embryonal carcinoma cells undergo cardiogenesis in response to high density and DMSO. We have derived a clonal subline which undergoes cardiogenesis in response to high density, but without requiring exposure to DMSO. The new subline retains the capacity to differentiate into skeletal muscle and neuronal cells in response to DMSO and retinoic acid. However, upon aggregation, these Oct 4-positive cells, termed P19-SI because they “self-induce” cardiac muscle, exhibit increased mRNAs encoding the mesodermal factor Brachyury, cardiac transcription factors Nkx 2.5 and GATA 4, the transcriptional repressor Msx-1, and cytokines Wnt 3a, Noggin and BMP 4. Exposure of aggregated P19-SI cells to BMP 4, a known inducer of cardiogenesis, accelerates cardiogenesis, as determined by rhythmic beating and myosin staining. However, cardiogenesis is severely inhibited when P19-SI cells are aggregated in the presence of BMP 4. These results demonstrate that cell-cell interaction is required before P19-SI cells can undergo a cardiogenic response to BMP 4. A concurrent increase in the expression of Msx-1 suggests one possible process underlying the inhibition of cardiogenesis. The phenotype of P19-SI cells offers an opportunity to explore new aspects of cardiac induction. PMID:16773658

BMP induction of cardiogenesis in P19 cells requires prior cell-cell interaction(s).

PubMed

Angello, John C; Kaestner, Stefanie; Welikson, Robert E; Buskin, Jean N; Hauschka, Stephen D

2006-08-01

Mouse P19 embryonal carcinoma cells undergo cardiogenesis in response to high density and DMSO. We have derived a clonal subline that undergoes cardiogenesis in response to high density, but without requiring exposure to DMSO. The new subline retains the capacity to differentiate into skeletal muscle and neuronal cells in response to DMSO and retinoic acid. However, upon aggregation, these Oct 4-positive cells, termed P19-SI because they "self-induce" cardiac muscle, exhibit increased mRNAs encoding the mesodermal factor Brachyury, cardiac transcription factors Nkx 2.5 and GATA 4, the transcriptional repressor Msx-1, and cytokines Wnt 3a, Noggin, and BMP 4. Exposure of aggregated P19-SI cells to BMP 4, a known inducer of cardiogenesis, accelerates cardiogenesis, as determined by rhythmic beating and myosin staining. However, cardiogenesis is severely inhibited when P19-SI cells are aggregated in the presence of BMP 4. These results demonstrate that cell-cell interaction is required before P19-SI cells can undergo a cardiogenic response to BMP 4. A concurrent increase in the expression of Msx-1 suggests one possible process underlying the inhibition of cardiogenesis. The phenotype of P19-SI cells offers an opportunity to explore new aspects of cardiac induction.

Treating B-cell cancer with T cells expressing anti-CD19 chimeric antigen receptors.

PubMed

Kochenderfer, James N; Rosenberg, Steven A

2013-05-01

Most B-cell malignancies express CD19, and a majority of patients with B-cell malignancies are not cured by current standard therapies. Chimeric antigen receptors (CARs) are fusion proteins consisting of antigen recognition moieties and T-cell activation domains. T cells can be genetically modified to express CARs, and adoptive transfer of anti-CD19 CAR T cells is now being tested in clinical trials. Effective clinical treatment with anti-CD19 CAR T cells was first reported in 2010 after a patient with advanced-stage lymphoma treated at the NCI experienced a partial remission of lymphoma and long-term eradication of normal B cells. Additional patients have subsequently obtained long-term remissions of advanced-stage B-cell malignancies after infusions of anti-CD19 CAR T cells. Long-term eradication of normal CD19(+) B cells from patients receiving infusions of anti-CD19 CAR T cells demonstrates the potent antigen-specific activity of these T cells. Some patients treated with anti-CD19 CAR T cells have experienced acute adverse effects, which were associated with increased levels of serum inflammatory cytokines. Although anti-CD19 CAR T cells are at an early stage of development, the potent antigen-specific activity observed in patients suggests that infusions of anti-CD19 CAR T cells might become a standard therapy for some B-cell malignancies.

DNA damage induces down-regulation of Prp19 via impairing Prp19 stability in hepatocellular carcinoma cells.

PubMed

Yin, Jie; Zhang, Yi-An; Liu, Tao-Tao; Zhu, Ji-Min; Shen, Xi-Zhong

2014-01-01

Pre-mRNA processing factor 19 (Prp19) activates pre-mRNA spliceosome and also mediates DNA damage response. Prp19 overexpression in cells with functional p53 leads to decreased apoptosis and increases cell survival after DNA damage. Here we showed that in hepatocellular carcinoma (HCC) cells with inactive p53 or functional p53, Prp19 was down-regulated due to the impaired stability under chemotherapeutic drug treatment. Silencing Prp19 expression enhanced apoptosis of HCC cells with or without chemotherapeutic drug treatment. Furthermore high level of Prp19 may inhibit chemotherapeutic drugs induced apoptosis in hepatocellular carcinoma cells through modulating myeloid leukemia cell differentiation 1 expression. These results indicated that targeting Prp19 may potentiate pro-apoptotic effect of chemotherapeutic agents on HCC.

Norepinephrine regulates hepatic innate immune system in leptin-deficient mice with nonalcoholic steatohepatitis.

PubMed

Li, Zhiping; Oben, Jude A; Yang, Shiqi; Lin, Huizhi; Stafford, Elizabeth A; Soloski, Mark J; Thomas, Steven A; Diehl, Anna Mae

2004-08-01

It is not known why natural killer T (NKT) cells, which modulate liver injury by regulating local cytokine production, are reduced in leptin-deficient ob/ob mice. NKT cells express adrenoceptors. Thus, we hypothesize that the low norepinephrine (NE) activity of ob/ob mice promotes depletion of liver NKT cells, thereby sensitizing ob/ob livers to lipopolysaccharide (LPS) toxicity. To evaluate this hypothesis, hepatic NKT cells were quantified in wild-type mice before and after treatment with NE inhibitors, and in dopamine beta-hydroxylase knockout mice (which cannot synthesize NE) and ob/ob mice before and after 4 weeks of NE supplementation. Decreasing NE activity consistently reduces liver NKT cells, while increasing NE has the opposite effect. Analysis of hepatic and thymic NKT cells in mice of different ages demonstrate an age-related accumulation of hepatic NKT cells in normal mice, while liver NKT cells become depleted after birth in ob/ob mice, which have increased apoptosis of hepatic NKT cells. NE treatment inhibits apoptosis and restores hepatic NKT cells. In ob/ob mice with reduced hepatic NKT cells, hepatic T and NKT cells produce excessive T helper (Th)-1 proinflammatory cytokines and the liver is sensitized to LPS toxicity. NE treatment decreases Th-1 cytokines, increases production of Th-2 cytokines, and reduces hepatotoxicity. Studies of CD1d-deficient mice, which lack the receptor required for NKT cell development, demonstrate that they are also unusually sensitive to LPS hepatotoxicity. In conclusion, low NE activity increases hepatic NKT cell apoptosis and depletes liver NKT cells, promoting proinflammatory polarization of hepatic cytokine production that sensitizes the liver to LPS toxicity. Copyright 2004 American Association for the Study of Liver Diseases

Cell Aggregation-induced FGF8 Elevation Is Essential for P19 Cell Neural Differentiation

PubMed Central

Wang, Chen; Xia, Caihong; Bian, Wei; Liu, Li; Lin, Wei; Chen, Ye-Guang; Ang, Siew-Lan

2006-01-01

FGF8, a member of the fibroblast growth factor (FGF) family, has been shown to play important roles in different developing systems. Mouse embryonic carcinoma P19 cells could be induced by retinoic acid (RA) to differentiate into neuroectodermal cell lineages, and this process is cell aggregation dependent. In this report, we show that FGF8 expression is transiently up-regulated upon P19 cell aggregation, and the aggregation-dependent FGF8 elevation is pluripotent stem cell related. Overexpressing FGF8 promotes RA-induced monolayer P19 cell neural differentiation. Inhibition of FGF8 expression by RNA interference or blocking FGF signaling by the FGF receptor inhibitor, SU5402, attenuates neural differentiation of the P19 cell. Blocking the bone morphogenetic protein (BMP) pathway by overexpressing Smad6 in P19 cells, we also show that FGF signaling plays a BMP inhibition–independent role in P19 cell neural differentiation. PMID:16641368

α-Galactosylceramide-activated Vα14 natural killer T cells mediate protection against murine malaria

PubMed Central

Gonzalez-Aseguinolaza, Gloria; de Oliveira, Camila; Tomaska, Margaret; Hong, Seokmann; Bruna-Romero, Oscar; Nakayama, Toshinori; Taniguchi, Masaru; Bendelac, Albert; Van Kaer, Luc; Koezuka, Yasuhiko; Tsuji, Moriya

2000-01-01

Natural killer T (NKT) cells are a unique population of lymphocytes that coexpress a semiinvariant T cell and natural killer cell receptors, which are particularly abundant in the liver. To investigate the possible effect of these cells on the development of the liver stages of malaria parasites, a glycolipid, α-galactosylceramide (α-GalCer), known to selectively activate Vα14 NKT cells in the context of CD1d molecules, was administered to sporozoite-inoculated mice. The administration of α-GalCer resulted in rapid, strong antimalaria activity, inhibiting the development of the intrahepatocytic stages of the rodent malaria parasites Plasmodium yoelii and Plasmodium berghei. The antimalaria activity mediated by α-GalCer is stage-specific, since the course of blood-stage-induced infection was not inhibited by administration of this glycolipid. Furthermore, it was determined that IFN-γ is essential for the antimalaria activity mediated by the glycolipid. Taken together, our results provide the clear evidence that NKT cells can mediate protection against an intracellular microbial infection. PMID:10900007

Natural killer cells promote tissue injury and systemic inflammatory responses during fatal Ehrlichia-induced toxic shock-like syndrome.

PubMed

Stevenson, Heather L; Estes, Mark D; Thirumalapura, Nagaraja R; Walker, David H; Ismail, Nahed

2010-08-01

Human monocytotropic ehrlichiosis is caused by Ehrlichia chaffeensis, a Gram-negative bacterium lacking lipopolysaccharide. We have shown that fatal murine ehrlichiosis is associated with CD8(+)T cell-mediated tissue damage, tumor necrosis factor-alpha, and interleukin (IL)-10 overproduction, and CD4(+)Th1 hyporesponsiveness. In this study, we examined the relative contributions of natural killer (NK) and NKT cells in Ehrlichia-induced toxic shock. Lethal ehrlichial infection in wild-type mice induced a decline in NKT cell numbers, and late expansion and migration of activated NK cells to the liver, a main infection site that coincided with development of hepatic injury. The spatial and temporal changes in NK and NKT cells in lethally infected mice correlated with higher NK cell cytotoxic activity, higher expression of cytotoxic molecules such as granzyme B, higher production of interferon-gamma and tumor necrosis factor-alpha, increased hepatic infiltration with CD8alphaCD11c(+) dendritic cells and CD8(+)T cells, decreased splenic CD4(+)T cells, increased serum concentrations of IL-12p40, IL-18, RANTES, and monocyte chemotactic protein-1, and elevated production of IL-18 by liver mononuclear cells compared with nonlethally infected mice. Depletion of NK cells prevented development of severe liver injury, decreased serum levels of interferon-gamma, tumor necrosis factor-alpha, and IL-10, and enhanced bacterial elimination. These data indicate that NK cells promote immunopathology and defective anti-ehrlichial immunity, possibly via decreasing the protective immune response mediated by interferon-gamma producing CD4(+)Th1 and NKT cells.

Genomic analysis of CD8+ NK/T cell line, ‘SRIK-NKL’, with array-based CGH (aCGH), SKY/FISH and molecular mapping

PubMed Central

Rossi, Michael; LaDuca, Jeff; Cowell, John; Srivastava, Bejai I.S.; Matsui, Sei-ichi

2010-01-01

We performed aCGH, SKY /FISH, molecular mapping and expression analyses on a permanent CD8+ NK/T cell line, ‘SRIK-NKL’ established from a lymphoma (ALL) patient, in attempt to define the fundamental genetic profile of its unique NK phenotypes. aCGH revealed hemizygous deletion of 6p containing genes responsible for hematopoietic functions. The SKY demonstrated that a constitutive reciprocal translocation, rcpt(5;14)(p13.2;q11) is a stable marker. Using somatic hybrids containing der(5) derived from SRIK-NKL, we found that the breakpoint in one homologue of no. 5 is located upstream of IL7R and also that the breakpoint in no. 14 is located within TRA@. The FISH analysis using BAC which contains TRA@ and its flanking region further revealed a ~231 kb deletion within 14q11 in the der(5) but not in the normal homologue of no. 14. The RT-PCR analysis detected mRNA for TRA@ transcripts which were extending across, but not including, the deleted region. IL7R was detected at least at mRNA levels. These findings were consistent with the immunological findings that TRA@ and IL7R are both expressed at mRNA levels and TRA@ at cytoplasmic protein levels in SRIK-NKL cells. In addition to rept(5;14), aCGH identified novel copy number abnormalities suggesting that the unique phenotype of the SRIK-NKL cell line is not solely due to the TRA@ rearrangement. These findings provide supportive evidence for the notion that SRIK-NKL cells may be useful for studying not only the function of NK cells but also genetic deregulations associated with leukemiogenesis. PMID:17640729

Production of stable GFP-expressing neural cells from P19 embryonal carcinoma stem cells.

PubMed

Shirzad, Hedayatollah; Esmaeili, Fariba; Bakhshalizadeh, Shabnam; Ebrahimie, Marzieh; Ebrahimie, Esmaeil

2017-04-01

Murine P19 embryonal carcinoma (EC) cells are convenient to differentiate into all germ layer derivatives. One of the advantages of P19 cells is that the exogenous DNA can be easily inserted into them. Here, at the first part of this study, we generated stable GFP-expressing P19 cells (P19-GFP + ). FACS and western-blot analysis confirmed stable expression of GFP in the cells. We previously demonstrated the efficient induction of neuronal differentiation from mouse ES and EC cells by application of a neuroprotective drug, selegiline In the second part of this study selegiline was used to induce differentiation of P19-GFP + into stable GFP-expressing neuron-like cells. Cresyl violet staining confirmed neuronal morphology of the differentiated cells. Furthermore, real-time PCR and immunoflourescence approved the expression of neuron specific markers. P19-GFP + cells were able to survive, migrate and integrated into host tissues when transplanted to developing chick embryo CNS. The obtained live GFP-expressing cells can be used as an abundant source of developmentally pluripotent material for transplantation studies, investigating the cellular and molecular aspects of early differentiation. Copyright © 2016 Elsevier Ltd. All rights reserved.

A Single Subset of Dendritic Cells Controls the Cytokine Bias of Natural Killer T Cell Responses to Diverse Glycolipid Antigens

PubMed Central

Arora, Pooja; Baena, Andres; Yu, Karl O.A.; Saini, Neeraj K.; Kharkwal, Shalu S.; Goldberg, Michael F.; Kunnath-Velayudhan, Shajo; Carreño, Leandro J.; Venkataswamy, Manjunatha M.; Kim, John; Lazar-Molnar, Eszter; Lauvau, Gregoire; Chang, Young-tae; Liu, Zheng; Bittman, Robert; Al-Shamkhani, Aymen; Cox, Liam R.; Jervis, Peter J.; Veerapen, Natacha; Besra, Gurdyal S.; Porcelli, Steven A.

2014-01-01

Summary Many hematopoietic cell types express CD1d and are capable of presenting glycolipid antigens to invariant natural killer T cells (iNKT cells). However, the question of which cells are the principal presenters of glycolipid antigens in vivo remains controversial, and it has been suggested that this might vary depending on the structure of a particular glycolipid antigen. Here we have shown that a single type of cell, the CD8α+ DEC-205+ dendritic cell, was mainly responsible for capturing and presenting a variety of different glycolipid antigens, including multiple forms of α-galactosylceramide that stimulate widely divergent cytokine responses. After glycolipid presentation, these dendritic cells rapidly altered their expression of various costimulatory and coinhibitory molecules in a manner that was dependent on the structure of the antigen. These findings show flexibility in the outcome of two-way communication between CD8α+ dendritic cells and iNKT cells, providing a mechanism for biasing toward either proinflammatory or anti-inflammatory responses. PMID:24412610

Infiltration of invariant natural killer T cells occur and accelerate brain infarction in permanent ischemic stroke in mice.

PubMed

Wang, Zhen-Kui; Xue, Li; Wang, Tao; Wang, Xiu-Jie; Su, Zhi-Qiang

2016-10-28

Invariant natural killer T (iNKT) cells are a unique subset of T cells that have been implicated in inflammation, atopy, autoimmunity, infections, and cancer. Although iNKT cells have been extensively studied over the past decade, its role in the pathogenesis of ischemic brain injury is still largely unknown. In our study, we determined whether iNKT cells infiltration occur in a mouse model of permanent cerebral ischemia. C57BL6/J male mice were treated with either alpha-galactosylceramide (α-GalCer) or vehicle control before undergoing permanent middle cerebral artery occlusion (pMCAO). α-GalCer, a glycolipid antigen, specifically activates iNKT cells by a CD1d-restricted mechanism. Using flow cytometry, 10,000 leukocytes (CD45 high cells) from the ischemic hemisphere and peripheral blood respectively were analyzed to determine the number of NK1.1 + CD3 + cells at 3, 12, 24 and 48h post-pMCAO. Cerebral infarct size, brain edema and morphological characteristics were measured at the stipulated time points by 2,3,5-triphenyltetrazolium chloride (TTC) staining, weighing, and H&E staining. The levels of IFN-γ and TNF-α in brain tissue and serum were assessed by immunohistochemistry and ELISA respectively. We found that the number of iNKT cells started increasing from 12h (PB sample) and 24h (ischemic hemisphere sample) respectively in the vehicle treated group. iNKT cells infiltration occurred at an earlier time-point compared in the α-GalCer treated group (T=3H vs T=12H in PB sample; T=12H vs T=24H in ischemic hemisphere sample). Brain water content at 12h and 24h was significantly higher in pMCAO+α-GalCer mice compared to pMCAO+vehicle mice which was in turn higher than mice that underwent sham surgery. Aggravated morphological abnormalities in HE-stained neurons and significantly increased neurons with pyknotic nuclei and cavitation in the ischemic region were observed at 24h in the pMCAO+α-GalCer and pMCAO+vehicle groups. Cerebral infarct volume

Fibroblast growth factor receptor 4 (FGFR4) and fibroblast growth factor 19 (FGF19) autocrine enhance breast cancer cells survival.

PubMed

Tiong, Kai Hung; Tan, Boon Shing; Choo, Heng Lungh; Chung, Felicia Fei-Lei; Hii, Ling-Wei; Tan, Si Hoey; Khor, Nelson Tze Woei; Wong, Shew Fung; See, Sze-Jia; Tan, Yuen-Fen; Rosli, Rozita; Cheong, Soon-Keng; Leong, Chee-Onn

2016-09-06

Basal-like breast cancer is an aggressive tumor subtype with poor prognosis. The discovery of underlying mechanisms mediating tumor cell survival, and the development of novel agents to target these pathways, is a priority for patients with basal-like breast cancer. From a functional screen to identify key drivers of basal-like breast cancer cell growth, we identified fibroblast growth factor receptor 4 (FGFR4) as a potential mediator of cell survival. We found that FGFR4 mediates cancer cell survival predominantly via activation of PI3K/AKT. Importantly, a subset of basal-like breast cancer cells also secrete fibroblast growth factor 19 (FGF19), a canonical ligand specific for FGFR4. siRNA-mediated silencing of FGF19 or neutralization of extracellular FGF19 by anti-FGF19 antibody (1A6) decreases AKT phosphorylation, suppresses cancer cell growth and enhances doxorubicin sensitivity only in the FGFR4+/FGF19+ breast cancer cells. Consistently, FGFR4/FGF19 co-expression was also observed in 82 out of 287 (28.6%) primary breast tumors, and their expression is strongly associated with AKT phosphorylation, Ki-67 staining, higher tumor stage and basal-like phenotype. In summary, our results demonstrated the presence of an FGFR4/FGF19 autocrine signaling that mediates the survival of a subset of basal-like breast cancer cells and suggest that inactivation of this autocrine loop may potentially serve as a novel therapeutic intervention for future treatment of breast cancers.

Adoptive immunotherapy utilizing anti-CD19 chimeric antigen receptor T-cells for B-cell malignancies.

PubMed

Oh, Iekuni; Oh, Yukiko; Ohmine, Ken

2016-01-01

Genetically modified T-cells with forced expression of anti-CD19 chimeric antigen receptor (CD19 CAR) have demonstrated promising clinical results for relapsed and refractory B cell malignancies in early clinical trial settings. The first beneficial tumor regressions were identified among approximately half of CLL patients in 2011. Similarly, CD19 CAR T-cells achieved remissions in about 80% of aggressive B-cell lymphomas in 2012. Furthermore, in 2013 this cellular therapy showed an extremely high rate of efficacy against refractory CD19 positive acute lymphoid leukemia, which had been regarded as the most difficult to treat hematologic disease. Recently, despite the absence of CD19 expression by neoplastic plasma cells, patients with refractory multiple myeloma achieved stringent complete remission after this therapy coupled with high dose chemotherapy and autologous stem cell transplantation. However, there are significant toxicities. Cytokine releasing syndrome and neurotoxicity are recognized as life-threatening adverse events. Although phase I/II clinical trials have just started in Japan, given the exciting results obtained to date, this cellular therapy is expected to be a novel breakthrough immunotherapy for treating refractory B-cell malignancies.

Saposins modulate human invariant Natural Killer T cells self-reactivity and facilitate lipid exchange with CD1d molecules during antigen presentation

PubMed Central

Salio, Mariolina; Ghadbane, Hemza; Dushek, Omer; Shepherd, Dawn; Cypen, Jeremy; Gileadi, Uzi; Aichinger, Michael C.; Napolitani, Giorgio; Qi, Xiaoyang; van der Merwe, P. Anton; Wojno, Justyna; Veerapen, Natacha; Cox, Liam R.; Besra, Gurdyal S.; Yuan, Weiming; Cresswell, Peter; Cerundolo, Vincenzo

2013-01-01

Lipid transfer proteins, such as molecules of the saposin family, facilitate extraction of lipids from biological membranes for their loading onto CD1d molecules. Although it has been shown that prosaposin-deficient mice fail to positively select invariant natural killer T (iNKT) cells, it remains unclear whether saposins can facilitate loading of endogenous iNKT cell agonists in the periphery during inflammatory responses. In addition, it is unclear whether saposins, in addition to loading, also promote dissociation of lipids bound to CD1d molecules. To address these questions, we used a combination of cellular assays and demonstrated that saposins influence CD1d-restricted presentation to human iNKT cells not only of exogenous lipids but also of endogenous ligands, such as the self-glycosphingolipid β-glucopyranosylceramide, up-regulated by antigen-presenting cells following bacterial infection. Furthermore, we demonstrated that in human myeloid cells CD1d-loading of endogenous lipids after bacterial infection, but not at steady state, requires trafficking of CD1d molecules through an endo-lysosomal compartment. Finally, using BIAcore assays we demonstrated that lipid-loaded saposin B increases the off-rate of lipids bound to CD1d molecules, providing important insights into the mechanisms by which it acts as a “lipid editor,” capable of fine-tuning loading and unloading of CD1d molecules. These results have important implications in understanding how to optimize lipid-loading onto antigen-presenting cells, to better harness iNKT cells central role at the interface between innate and adaptive immunity. PMID:24248359

In Vivo Cytometry of Antigen-Specific T Cells Using 19F MRI

PubMed Central

Srinivas, Mangala; Turner, Michael S.; Janjic, Jelena M.; Morel, Penelope A.; Laidlaw, David H.; Ahrens, Eric T.

2009-01-01

Noninvasive methods to image the trafficking of phenotypically defined immune cells are paramount as we attempt to understand adaptive immunity. A 19F MRI-based methodology for tracking and quantifying cells of a defined phenotype is presented. These methods were applied to a murine inflammation model using antigen-specific T cells. The T cells that were intracellularly labeled ex vivo with a perfluoropolyether (PFPE) nanoemulsion and cells were transferred to a host receiving a localized inoculation of antigen. Longitudinal 19F MRI over 21 days revealed a dynamic accumulation and clearance of T cells in the lymph node (LN) draining the antigen. The apparent T-cell numbers were calculated in the LN from the time-lapse 19F MRI data. The effect of in vivo T-cell division on the 19F MRI cell quantification accuracy was investigated using fluorescence assays. Overall, in vivo cytometry using PFPE labeling and 19F MRI is broadly applicable to studies of whole-body cell biodistribution. PMID:19585593

Frequency of γδ T Cells and Invariant Natural Killer T Cells in Helicobacter Pylori-infected Patients with Peptic Ulcer and Gastric Cancer.

PubMed

Shadman, Mojtaba; Rajabian, Zeinab; Ajami, Abolghasem; Hussein-Nattaj, Hadi; Rafiei, Alireza; Hosseini, Vahid; Taghvaei, Tarang; Abbasi, Ali; Tehrani, Mohsen

2015-10-01

To clarify the effect of γδ T cells and invariant Natural Killer T (iNKT) cells in pathophysiology of dyspeptic disorders, number of these two cells in patients with non-ulcer dyspepsia (NUD), peptic ulcer disease (PUD), and gastric cancer (GC) were compared.Patients with dyspepsia were divided into three groups of NUD, PUD, and GC according to their endoscopic and histopathological examinations. Helicobacter pylori infection was diagnosed by rapid urease test and histopathology. The number of peripheral blood CD3+TCRγδ(+) T cells and CD3+Va24Ja18+ iNKT cells were determined by flow cytometry. Immunohistochemistry (IHC) was also used for identifying the TCRγδ+ cells.Forty two patients with NUD (31.6%), 44 with PUD (33.1%), and 47 with GC (35.3%) were included in the study. The frequency of CD3+TCRγδ(+) T cells in peripheral blood of patients with GC (2.71±0.25) was significantly lower than that in NUD (3.97±0.32, p<0.05) and PUD groups (3.87±0.32, p<0.05). However, there was no significant difference in CD3+TCRγδ(+) T cell percentage between the NUD and PUD groups. The frequency of TCRγδ(+) lymphocytes was significantly lower in tissue samples from patients with GC (4.81±0.53) than in NUD (11.09±1.09, p<0.0001) and PUD groups (11.11±1.01, p<0.0001). Also, we could not find any significant difference in the percentage of mucosal TCRγδ+ cells between the NUD and PUD groups. The results showed no significant difference in iNKT cells percentage among the three groups of patients.The results suggest that decreasing number of γδ T cells may be related to development and progression of gastric cancer.

Extranodal NK/T Cell Lymphoma, Nasal Type (ENKTL-NT): An Update on Epidemiology, Clinical Presentation, and Natural History in North American and European Cases.

PubMed

Haverkos, Bradley M; Pan, Zenggang; Gru, Alejandro A; Freud, Aharon G; Rabinovitch, Rachel; Xu-Welliver, Meng; Otto, Brad; Barrionuevo, Carlos; Baiocchi, Robert A; Rochford, Rosemary; Porcu, Pierluigi

2016-12-01

Extranodal NK/T cell lymphoma, nasal type (ENKTL-NT) is an aggressive extranodal non-Hodgkin lymphoma most commonly occurring in East Asia and Latin America but with increasing incidence in the United States. Data on epidemiology, disease presentation, and outcome for European and North American ("Western") cases are very limited. We review published landmark clinical studies on ENKTL-NT in the West and report in detail recent data, including our institutional experience. We highlight key observations in its epidemiology, natural history, and trends in clinical management. In the USA, ENKTL-NT is more common among Asian Pacific Islanders (API) and Hispanics compared to non-Hispanic whites. Published studies indicate less heterogeneity in clinical presentation in Western ENKTL-NT compared to Asian patients. While there is variation in age at diagnosis, presence of antecedent lymphoproliferative disorders, and outcomes among racial/ethnic groups, the universal association of ENKTL-NT with EBV and the poor response of this neoplasm to anthracycline-based therapy is consistent across all geographic areas. Data on epidemiology, disease presentation, and clinical outcomes in mature T cell and NK cell (T/NK cell) neoplasms, including ENKTL-NT, in Europe and North America are very limited. As the classification and diagnostic characterization of the currently recognized T/NK cell lymphoma disease entities continue to evolve, gaps and inconsistencies in data reporting across different studies are being recognized. Despite these limitations, several studies from the USA suggest that the incidence of ENKTL-NT is higher in Asian Pacific Islanders (API) and non-white Hispanics and that outcomes may be worse in non-whites. However, the universal association of ENKTL-NT with Epstein-Barr virus (EBV) across all ethnic groups suggests a common pathogenesis. Given the overlap between the entities included in the category of T/NK cell neoplasms, there is a need to further define

Targeted antibody-mediated depletion of murine CD19 CAR T cells permanently reverses B cell aplasia

PubMed Central

Paszkiewicz, Paulina J.; Fräßle, Simon P.; Srivastava, Shivani; Sommermeyer, Daniel; Hudecek, Michael; Sadelain, Michel; Liu, Lingfeng; Jensen, Michael C.; Riddell, Stanley R.; Busch, Dirk H.

2016-01-01

The adoptive transfer of T cells that have been genetically modified to express a CD19-specific chimeric antigen receptor (CAR) is effective for treating human B cell malignancies. However, the persistence of functional CD19 CAR T cells causes sustained depletion of endogenous CD19+ B cells and hypogammaglobulinemia. Thus, there is a need for a mechanism to ablate transferred T cells after tumor eradication is complete to allow recovery of normal B cells. Previously, we developed a truncated version of the epidermal growth factor receptor (EGFRt) that is coexpressed with the CAR on the T cell surface. Here, we show that targeting EGFRt with the IgG1 monoclonal antibody cetuximab eliminates CD19 CAR T cells both early and late after adoptive transfer in mice, resulting in complete and permanent recovery of normal functional B cells, without tumor relapse. EGFRt can be incorporated into many clinical applications to regulate the survival of gene-engineered cells. These results support the concept that EGFRt represents a promising approach to improve safety of cell-based therapies. PMID:27760047

Targeted antibody-mediated depletion of murine CD19 CAR T cells permanently reverses B cell aplasia.

PubMed

Paszkiewicz, Paulina J; Fräßle, Simon P; Srivastava, Shivani; Sommermeyer, Daniel; Hudecek, Michael; Drexler, Ingo; Sadelain, Michel; Liu, Lingfeng; Jensen, Michael C; Riddell, Stanley R; Busch, Dirk H

2016-11-01

The adoptive transfer of T cells that have been genetically modified to express a CD19-specific chimeric antigen receptor (CAR) is effective for treating human B cell malignancies. However, the persistence of functional CD19 CAR T cells causes sustained depletion of endogenous CD19+ B cells and hypogammaglobulinemia. Thus, there is a need for a mechanism to ablate transferred T cells after tumor eradication is complete to allow recovery of normal B cells. Previously, we developed a truncated version of the epidermal growth factor receptor (EGFRt) that is coexpressed with the CAR on the T cell surface. Here, we show that targeting EGFRt with the IgG1 monoclonal antibody cetuximab eliminates CD19 CAR T cells both early and late after adoptive transfer in mice, resulting in complete and permanent recovery of normal functional B cells, without tumor relapse. EGFRt can be incorporated into many clinical applications to regulate the survival of gene-engineered cells. These results support the concept that EGFRt represents a promising approach to improve safety of cell-based therapies.

Invariant natural killer T cells from children with versus without food allergy exhibit differential responsiveness to milk-derived sphingomyelin.

PubMed

Jyonouchi, Soma; Abraham, Valsamma; Orange, Jordan S; Spergel, Jonathan M; Gober, Laura; Dudek, Emily; Saltzman, Rushani; Nichols, Kim E; Cianferoni, Antonella

2011-07-01

A key immunologic feature of food allergy (FA) is the presence of a T(h)2-type cytokine bias. Ligation of the invariant natural killer T cell (iNKT) T-cell receptor (TCR) by sphingolipids presented via the CD1d molecule leads to copious secretion of T(h)2-type cytokines. Major food allergens (eg, milk, egg) are the richest dietary source of sphingolipids (food-derived sphingolipids [food-SLs]). Nonetheless, the role of iNKTs in FA is unknown. To investigate the role of iNKTs in FA and to assess whether food-SL-CD1d complexes can engage the iNKT-TCR and induce iNKT functions. PBMCs from 15 children with cow's milk allergy (MA), 12 children tolerant to cow's milk but with allergy to egg, and 13 healthy controls were incubated with α-galactosylceramide (αGal), cow's milk-sphingomyelin, or hen's egg-ceramide. iNKTs were quantified, and their cytokine production and proliferation were assessed. Human CD1d tetramers loaded with milk-sphingomyelin or egg-ceramide were used to determine food-SL binding to the iNKT-TCR. Milk-sphingomyelin, but not egg-ceramide, can engage the iNKT-TCR and induce iNKT proliferation and T(h)2-type cytokine secretion. Children with FA, especially those with MA, had significantly fewer peripheral blood iNKTs and their iNKTs exhibited a greater T(h)2 response to αGal and milk-sphingomyelin than iNKTs of healthy controls. iNKTs from children with FA, especially those with MA, are reduced in number and exhibit a T(h)2 bias in response to αGal and milk-sphingomyelin. These data suggest a potential role for iNKTs in FA. Copyright © 2011 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

In vivo MRI cell tracking using perfluorocarbon probes and fluorine-19 detection

PubMed Central

Ahrens, Eric T.; Zhong, Jia

2013-01-01

This article is a brief survey of preclinical in vivo cell tracking methods and applications using perfluorocarbon (PFC) probes and fluorine-19 (19F) MRI detection. Detection of the 19F signal offers high cell specificity and quantification abilities in spin-density weighted MR images. We discuss the compositions of matter, methods, and applications of PFC-based cell tracking using ex vivo and in situ PFC labeling in preclinical studies of inflammation and cellular therapeutics. We will also address potential applicability of 19F cell tracking to clinical trials. PMID:23606473

CD19 chimeric antigen receptor (CD19 CAR)-redirected adoptive T-cell immunotherapy for the treatment of relapsed or refractory B-cell Non-Hodgkin's Lymphomas.

PubMed

Onea, Alexandra S; Jazirehi, Ali R

2016-01-01

Recovery rates for B-cell Non-Hodgkin's Lymphoma (NHL) are up to 70% with current standard-of-care treatments including rituximab (chimeric anti-CD20 monoclonal antibody) in combination with chemotherapy (R-CHOP). However, patients who do not respond to first-line treatment or develop resistance have a very poor prognosis. This signifies the need for the development of an optimal treatment approach for relapsed/refractory B-NHL. Novel CD19- chimeric antigen receptor (CAR) T-cell redirected immunotherapy is an attractive option for this subset of patients. Anti-CD19 CAR T-cell therapy has already had remarkable efficacy in various leukemias as well as encouraging outcomes in phase I clinical trials of relapsed/refractory NHL. In going forward with additional clinical trials, complementary treatments that may circumvent potential resistance mechanisms should be used alongside anti-CD19 T-cells in order to prevent relapse with resistant strains of disease. Some such supplementary tactics include conditioning with lymphodepletion agents, sensitizing with kinase inhibitors and Bcl-2 inhibitors, enhancing function with multispecific CAR T-cells and CD40 ligand-expressing CAR T-cells, and safeguarding with lymphoma stem cell-targeted treatments. A therapy regimen involving anti-CD19 CAR T-cells and one or more auxiliary treatments could dramatically improve prognoses for patients with relapsed/refractory B-cell NHL. This approach has the potential to revolutionize B-NHL salvage therapy in much the same way rituximab did for first-line treatments.

Adhesion of Epstein–Barr virus-positive natural killer cell lines to cultured endothelial cells stimulated with inflammatory cytokines

PubMed Central

Kanno, H; Watabe, D; Shimizu, N; Sawai, T

2008-01-01

Chronic active Epstein–Barr virus (EBV) infection (CAEBV) is characterized by chronic recurrent infectious mononucleosis-like symptoms. Approximately one-fourth of CAEBV patients develop vascular lesions with infiltration of EBV-positive lymphoid cells. Furthermore, EBV-positive natural killer (NK)/T cell lymphomas often exhibit angiocentric or angiodestructive lesions. These suggest an affinity of EBV-positive NK/T cells to vascular components. In this study, we evaluated the expression of adhesion molecules and cytokines in EBV-positive NK lymphoma cell lines, SNK1 and SNK6, and examined the role of cytokines in the interaction between NK cell lines and endothelial cells. SNKs expressed intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) at much higher levels than those in EBV-negative T cell lines. SNKs produced the larger amount of tumour necrosis factor (TNF)-α, which caused increased expression of ICAM-1 and VCAM-1 in cultured human endothelial cells, than that from EBV-negative T cell lines. Furthermore, SNKs exhibited increased adhesion to cultured endothelial cells stimulated with TNF-α or interleukin (IL)-1β, and the pretreatment of cytokine-stimulated endothelial cells with anti-VCAM-1-antibodies reduced cell adhesion. These indicate that the up-regulated expression of VCAM-1 on cytokine-stimulated endothelial cells would be important for the adhesion of EBV-positive NK cells and might initiate the vascular lesions. PMID:18190605

Granzyme B mediated function of Parvovirus B19-specific CD4+ T cells

PubMed Central

Kumar, Arun; Perdomo, Maria F; Kantele, Anu; Hedman, Lea; Hedman, Klaus; Franssila, Rauli

2015-01-01

A novel conception of CD4+ T cells with cytolytic potential (CD4+ CTL) is emerging. These cells appear to have a part in controlling malignancies and chronic infections. Human parvovirus B19 can cause a persistent infection, yet no data exist on the presence of B19-specific CD4+ CTLs. Such cells could have a role in the pathogenesis of some autoimmune disorders reported to be associated with B19. We explored the cytolytic potential of human parvovirus B19-specific T cells by stimulating peripheral blood mononuclear cell (PBMC) with recombinant B19-VP2 virus-like particles. The cytolytic potential was determined by enzyme immunoassay-based quantitation of granzyme B (GrB) and perforin from the tissue culture supernatants, by intracellular cytokine staining (ICS) and by detecting direct cytotoxicity. GrB and perforin responses with the B19 antigen were readily detectable in B19-seropositive individuals. T-cell depletion, HLA blocking and ICS experiments showed GrB and perforin to be secreted by CD4+ T cells. CD4+ T cells with strong GrB responses were found to exhibit direct cytotoxicity. As anticipated, ICS of B19-specific CD4+ T cells showed expected co-expression of GrB, perforin and interferon gamma (IFN-γ). Unexpectedly, also a strong co-expression of GrB and interleukin 17 (IL-17) was detected. These cells expressed natural killer (NK) cell surface marker CD56, together with the CD4 surface marker. To our knowledge, this is the first report on virus-specific CD4+ CTLs co-expressing CD56 antigen. Our results suggest a role for CD4+ CTL in B19 immunity. Such cells could function within both immune regulation and triggering of autoimmune phenomena such as systemic lupus erythematosus (SLE) or rheumatoid arthritis. PMID:26246896

Rapid and preferential distribution of blood-borne αCD3εAb to the liver is followed by local stimulation of T cells and natural killer T cells

PubMed Central

Wingender, Gerhard; Schumak, Beatrix; Schurich, Anna; Gessner, J Engelbert; Endl, Elmar; Limmer, Andreas; Knolle, Percy A

2006-01-01

Dissemination of soluble molecules or antigens via the blood stream is considered to lead to a uniform distribution in the various organs of the body, but organ-specific microarchitecture and vascularization may influence this. Following intravenous injection of αCD3ε antibody (αCD3εAb) we observed clear differences in antibody binding to Fcγ receptor (FcγR)+ antigen-presenting cells (APCs) or T lymphocytes in different organs. Significant binding of blood-borne αCD3εAb was only detected in the spleen and liver and not in the thymus or lymph node. In the spleen, only 10% of dendritic cells/macrophages and 40% of T-cell receptor (TCR)-β+ cells were positive for αCD3εAb, and, dependent on FcγR-mediated cross-linking of αCD3εAb, a similar percentage of splenic TCR-β+ cells were stimulated and became CD69+. Stimulation of TCR-β+ cells in the liver was at least as efficient as in the spleen, but almost all T cells and all scavenger liver sinusoidal endothelial cells bound αCD3εAb. In contrast to CD69 up-regulation, only CD4+ natural killer T (NKT) cells and CD11ahigh CD8+ T cells were activated by αCD3εAb and expressed interferon (IFN)-γ. Again, IFN-γ release from NKT/T cells was at least as efficient in the liver as in the spleen. Taken together, our results support the notion that the combination of extensive hepatic vascularization and very high scavenger activity allows the liver to fulfill its metabolic tasks and to promote stimulation of the large but widely distributed hepatic population of NKT/T cells. PMID:16423047

Paving the road ahead for CD19 CAR T-cell therapy.

PubMed

Nellan, Anandani; Lee, Daniel W

2015-11-01

Modern immunotherapies, most notably in the form of anti-CD19 chimeric antigen receptor (CAR) T cells, have produced significant clinical responses in otherwise refractory pre-B-cell acute lymphoblastic leukemia patients. Several groups have simultaneously reported robust response rates in children and adults alike. These early studies indicate an impending shift in paradigm for the treatment of acute lymphoblastic leukemia. Incorporating CD19 CAR T-cell therapy into upfront or salvage regimens has its challenges and opportunities. Most CD19 CAR T-cell products in trial today are excellent at inducing minimal residual disease negative remissions, and most responding patients experience cytokine release syndrome and/or neurotoxicity. The challenges facing the CAR community involve how best to minimize the severity of cytokine release syndrome and neurotoxicity while maximizing antitumor efficacy, determining what role this therapy will play for the prophylaxis and treatment of central nervous system leukemia, and its implications on subsequent hematopoietic stem cell transplant given the emergence of CD19-negative relapses. CD19 CAR T-cell therapy is a powerful new tool in the oncologist's arsenal. How it is incorporated into standard practice and how it will shift survival curves are the exciting questions that are waiting to be answered.

Adipose derived mesenchymal stem cells partially rescue mitomycin C treated ARPE19 cells from death in co-culture condition.

PubMed

Singh, Amar K; Srivastava, Girish K; García-Gutiérrez, María T; Pastor, J Carlos

2013-12-01

Age-related macular degeneration is a retinal disease with important damage at the RPE layer. This layer is considered a target for therapeutical approaches. Stem cell transplantation is a promising option for retinal diseases. Adipose derived mesenchymal stem cells secret growth factors which might play a significant role in RPE maintenance. This study aimed to evaluate human AD-MSCs ability to rescue mitomycin C treated dying ARPE19 cells in co-culture condition. ARPE19 cells were treated with MMC (50 μg/ml, 100 μg/ml and 200 μg/ml) for 2 hours to induce cell death. These treated cells were co-cultured with hAD-MSCs in indirect co-culture system for 3 days and 3 weeks. Then the viability, growth and proliferation of these ARPE19 cells were evaluated by a cell viability/cytotoxicity assay kit and Alamar Blue (AB) assay. Untreated ARPE19 cells and human skin fibroblasts (HSF) were used as controls. MMC blocked ARPE19 cell proliferation significantly in 3 days and cells were almost completely dead after 3 weeks. Cell toxicity of MMC increased significantly with concentration. When these cells were co-cultured with hAD-MSCs, a significant growth difference was observed in treated cells compared to untreated cells. hAD-MSCs rescue capacity was also significantly higher than HSF for treated ARPE19 cells. This study showed that hAD-MSCs rescued MMC treated ARPE19 cells from death. It probably occurred due to undefined growth factors secreted by hAD-MSCs in the medium, shared by treated ARPE19 cells in co-culture conditions. This study supports further evaluation of the effect of hAD-MSCs subretinal transplantation over the RPE degeneration process in AMD patients.

Activation of CD11b+ Kupffer Cells/Macrophages as a Common Cause for Exacerbation of TNF/Fas-Ligand-Dependent Hepatitis in Hypercholesterolemic Mice

PubMed Central

Nakashima, Hiroyuki; Ogawa, Yoshiko; Shono, Satoshi; Kinoshita, Manabu; Nakashima, Masahiro; Sato, Atsushi; Ikarashi, Masami; Seki, Shuhji

2013-01-01

We have reported that the mouse hepatic injury induced by either α-galactosylceramide (α-GalCer) or bacterial DNA motifs (CpG-ODN) is mediated by the TNF/NKT cell/Fas-ligand (FasL) pathway. In addition, F4/80+ Kupffer cells can be subclassified into CD68+ subset with a phagocytosing capacity and CD11b+ subset with a TNF-producing capacity. CD11b+ subset increase if mice are fed high-fat and cholesterol diet (HFCD). The present study examined how a HFCD affects the function of NKT cells and F4/80+ CD11b+ subset and these hepatitis models. After the C57BL/6 mice received a HFCD, high-cholesterol diet (HCD), high-fat diet (HFD) and control diet (CD) for four weeks, the HFCD mice increased surface CD1d and intracellular TLR-9 expression by the CD11b+ population compared to CD mice. Hepatic injury induced either by α-GalCer or CpG-ODN was more severe in HCD and HFCD mice compared to CD mice, which was in proportion to the serum TNF levels. In addition, liver cholesterol levels but not serum cholesterol levels nor liver triglyceride levels were involved in the aggravation of hepatitis. The FasL expression of NKT cells induced by both reagents was upregulated in HFCD mice. Furthermore, the liver mononuclear cells and purified F4/80+ CD11b+ subset from HFCD mice stimulated with either reagent in vitro produced a larger amount of TNF than did those from CD mice. Intracellular TNF production in F4/80+ CD11b+ cells was confirmed. The increased number of F4/80+ CD11b+ Kupffer cells/macrophages by HFCD and their enhanced TNF production thus play a pivotal role in TNF/NKT cell/FasL dependent hepatic injury. PMID:23372642

Activation of CD11b+ Kupffer cells/macrophages as a common cause for exacerbation of TNF/Fas-ligand-dependent hepatitis in hypercholesterolemic mice.

PubMed

Nakashima, Hiroyuki; Ogawa, Yoshiko; Shono, Satoshi; Kinoshita, Manabu; Nakashima, Masahiro; Sato, Atsushi; Ikarashi, Masami; Seki, Shuhji

2013-01-01

We have reported that the mouse hepatic injury induced by either α-galactosylceramide (α-GalCer) or bacterial DNA motifs (CpG-ODN) is mediated by the TNF/NKT cell/Fas-ligand (FasL) pathway. In addition, F4/80(+) Kupffer cells can be subclassified into CD68(+) subset with a phagocytosing capacity and CD11b(+) subset with a TNF-producing capacity. CD11b(+) subset increase if mice are fed high-fat and cholesterol diet (HFCD). The present study examined how a HFCD affects the function of NKT cells and F4/80(+) CD11b(+) subset and these hepatitis models. After the C57BL/6 mice received a HFCD, high-cholesterol diet (HCD), high-fat diet (HFD) and control diet (CD) for four weeks, the HFCD mice increased surface CD1d and intracellular TLR-9 expression by the CD11b(+) population compared to CD mice. Hepatic injury induced either by α-GalCer or CpG-ODN was more severe in HCD and HFCD mice compared to CD mice, which was in proportion to the serum TNF levels. In addition, liver cholesterol levels but not serum cholesterol levels nor liver triglyceride levels were involved in the aggravation of hepatitis. The FasL expression of NKT cells induced by both reagents was upregulated in HFCD mice. Furthermore, the liver mononuclear cells and purified F4/80(+) CD11b(+) subset from HFCD mice stimulated with either reagent in vitro produced a larger amount of TNF than did those from CD mice. Intracellular TNF production in F4/80(+) CD11b(+) cells was confirmed. The increased number of F4/80(+) CD11b(+) Kupffer cells/macrophages by HFCD and their enhanced TNF production thus play a pivotal role in TNF/NKT cell/FasL dependent hepatic injury.

CD19 chimeric antigen receptor (CD19 CAR)-redirected adoptive T-cell immunotherapy for the treatment of relapsed or refractory B-cell Non-Hodgkin’s Lymphomas

PubMed Central

Onea, Alexandra S; Jazirehi, Ali R

2016-01-01

Recovery rates for B-cell Non-Hodgkin’s Lymphoma (NHL) are up to 70% with current standard-of-care treatments including rituximab (chimeric anti-CD20 monoclonal antibody) in combination with chemotherapy (R-CHOP). However, patients who do not respond to first-line treatment or develop resistance have a very poor prognosis. This signifies the need for the development of an optimal treatment approach for relapsed/refractory B-NHL. Novel CD19- chimeric antigen receptor (CAR) T-cell redirected immunotherapy is an attractive option for this subset of patients. Anti-CD19 CAR T-cell therapy has already had remarkable efficacy in various leukemias as well as encouraging outcomes in phase I clinical trials of relapsed/refractory NHL. In going forward with additional clinical trials, complementary treatments that may circumvent potential resistance mechanisms should be used alongside anti-CD19 T-cells in order to prevent relapse with resistant strains of disease. Some such supplementary tactics include conditioning with lymphodepletion agents, sensitizing with kinase inhibitors and Bcl-2 inhibitors, enhancing function with multispecific CAR T-cells and CD40 ligand-expressing CAR T-cells, and safeguarding with lymphoma stem cell-targeted treatments. A therapy regimen involving anti-CD19 CAR T-cells and one or more auxiliary treatments could dramatically improve prognoses for patients with relapsed/refractory B-cell NHL. This approach has the potential to revolutionize B-NHL salvage therapy in much the same way rituximab did for first-line treatments. PMID:27186412

Mechanisms of Invariant Natural Killer T Cell-Mediated Immunoregulation in Cancer

DTIC Science & Technology

2012-05-01

by mesenchymal stem cells . Intriguingly, the increased metastatic ability was dependent on the production of CCL5 by mesenchymal stem cells , which...Tubo, R., &Weinberg, R.A.(2007) Mesenchymal stem cells within tumour stroma promote breast cancer metastasis. Nature. Vol. 449:pp557-563. Breast...can induce preferential secretion of immunosuppressive cytokines ; 2) iNKT cells inhibit effector T cell priming by killing dendritic cells that

Fatal natural killer cell lymphoma arising in a patient with a crop of Epstein-Barr virus-associated disorders.

PubMed

Nitta, Yukiko; Iwatsuki, Keiji; Kimura, Hiroshi; Kojima, Seiji; Morishima, Tsuneo; Tsuji, Kazuhide; Oono, Takashi

2005-01-01

Natural killer (NK) lymphoma in Asia is frequently associated with latent Epstein-Barr (EBV) infection. Unlike the adult cases, EBV-associated NK/T cell lymphomas in children are often preceded by various EBV-related disorders, including chronic active EBV infection (CAEBV), hypersensitivity to mosquito bites (HMB), virus-associated haemophagocytic syndrome (VAHS), and hydroa vacciniforme (HV)-like eruptions. Here, we report a 14-year-old Japanese girl who sequentially developed all the symptoms related to EBV-associated NK/T cell lymphoproliferative disorders in a 12-year clinical course. Our observations confirm the spectrum of EBV-associated cutaneous disorders and indicate the importance of long-term follow-up.

Fluorine-19 Labeling of Stromal Vascular Fraction Cells for Clinical Imaging Applications

PubMed Central

Rose, Laura C.; Kadayakkara, Deepak K.; Wang, Guan; Bar-Shir, Amnon; Helfer, Brooke M.; O’Hanlon, Charles F.; Kraitchman, Dara L.; Rodriguez, Ricardo L.

2015-01-01

Stromal vascular fraction (SVF) cells are used clinically for various therapeutic targets. The location and persistence of engrafted SVF cells are important parameters for determining treatment failure versus success. We used the GID SVF-1 platform and a clinical protocol to harvest and label SVF cells with the fluorinated (19F) agent CS-1000 as part of a first-in-human phase I trial (clinicaltrials.gov identifier NCT02035085) to track SVF cells with magnetic resonance imaging during treatment of radiation-induced fibrosis in breast cancer patients. Flow cytometry revealed that SVF cells consisted of 25.0% ± 15.8% CD45+, 24.6% ± 12.5% CD34+, and 7.5% ± 3.3% CD31+ cells, with 2.1 ± 0.7 × 105 cells per cubic centimeter of adipose tissue obtained. Fluorescent CS-1000 (CS-ATM DM Green) labeled 87.0% ± 13.5% of CD34+ progenitor cells compared with 47.8% ± 18.5% of hematopoietic CD45+ cells, with an average of 2.8 ± 2.0 × 1012 19F atoms per cell, determined using nuclear magnetic resonance spectroscopy. The vast majority (92.7% ± 5.0%) of CD31+ cells were also labeled, although most coexpressed CD34. Only 16% ± 22.3% of CD45−/CD31−/CD34− (triple-negative) cells were labeled with CS-ATM DM Green. After induction of cell death by either apoptosis or necrosis, >95% of 19F was released from the cells, indicating that fluorine retention can be used as a surrogate marker for cell survival. Labeled-SVF cells engrafted in a silicone breast phantom could be visualized with a clinical 3-Tesla magnetic resonance imaging scanner at a sensitivity of approximately 2 × 106 cells at a depth of 5 mm. The current protocol can be used to image transplanted SVF cells at clinically relevant cell concentrations in patients. Significance Stromal vascular fraction (SVF) cells harvested from adipose tissue offer great promise in regenerative medicine, but methods to track such cell therapies are needed to ensure correct administration and monitor survival. A clinical protocol

Larger number of invariant natural killer T cells in PBSC allografts correlates with improved GVHD-free and progression-free survival.

PubMed

Malard, Florent; Labopin, Myriam; Chevallier, Patrice; Guillaume, Thierry; Duquesne, Alix; Rialland, Fanny; Derenne, Sophie; Peterlin, Pierre; Leauté, Anne-Gaelle; Brissot, Eolia; Gregoire, Marc; Moreau, Philippe; Saas, Philippe; Gaugler, Béatrice; Mohty, Mohamad

2016-04-07

We studied the impact of a set of immune cells contained within granulocyte colony-stimulating factor-mobilized peripheral blood stem cell grafts (naïve and memory T-cell subsets, B cells, regulatory T cells, invariant natural killer T cells [iNKTs], NK cells, and dendritic cell subsets) in patients (n = 80) undergoing allogeneic stem cell transplantation (SCT), using the composite end point of graft-versus-host disease (GVHD)-free and progression-free survival (GPFS) as the primary end point. We observed that GPFS incidences in patients receiving iNKT doses above and below the median were 49% vs 22%, respectively (P= .007). In multivariate analysis, the iNKT dose was the only parameter with a significant impact on GPFS (hazard ratio = 0.48; 95% confidence interval, 0.27-0.85;P= .01). The incidences of severe grade III to IV acute GVHD and National Institutes of Health grade 2 to 3 chronic GVHD (12% and 16%, respectively) were low and associated with the use of antithymocyte globulin in 91% of patients. No difference in GVHD incidence was reported according to the iNKT dose. In conclusion, a higher dose of iNKTs within the graft is associated with an improved GPFS. These data may pave the way for prospective and active interventions aiming to manipulate the graft content to improve allo-SCT outcome. © 2016 by The American Society of Hematology.

Donor-derived CD19-targeted T cells cause regression of malignancy persisting after allogeneic hematopoietic stem cell transplantation.

PubMed

Kochenderfer, James N; Dudley, Mark E; Carpenter, Robert O; Kassim, Sadik H; Rose, Jeremy J; Telford, William G; Hakim, Frances T; Halverson, David C; Fowler, Daniel H; Hardy, Nancy M; Mato, Anthony R; Hickstein, Dennis D; Gea-Banacloche, Juan C; Pavletic, Steven Z; Sportes, Claude; Maric, Irina; Feldman, Steven A; Hansen, Brenna G; Wilder, Jennifer S; Blacklock-Schuver, Bazetta; Jena, Bipulendu; Bishop, Michael R; Gress, Ronald E; Rosenberg, Steven A

2013-12-12

New treatments are needed for B-cell malignancies persisting after allogeneic hematopoietic stem cell transplantation (alloHSCT). We conducted a clinical trial of allogeneic T cells genetically modified to express a chimeric antigen receptor (CAR) targeting the B-cell antigen CD19. T cells for genetic modification were obtained from each patient's alloHSCT donor. All patients had malignancy that persisted after alloHSCT and standard donor lymphocyte infusions (DLIs). Patients did not receive chemotherapy prior to the CAR T-cell infusions and were not lymphocyte depleted at the time of the infusions. The 10 treated patients received a single infusion of allogeneic anti-CD19-CAR T cells. Three patients had regressions of their malignancies. One patient with chronic lymphocytic leukemia (CLL) obtained an ongoing complete remission after treatment with allogeneic anti-CD19-CAR T cells, another CLL patient had tumor lysis syndrome as his leukemia dramatically regressed, and a patient with mantle cell lymphoma obtained an ongoing partial remission. None of the 10 patients developed graft-versus-host disease (GVHD). Toxicities included transient hypotension and fever. We detected cells containing the anti-CD19-CAR gene in the blood of 8 of 10 patients. These results show for the first time that donor-derived allogeneic anti-CD19-CAR T cells can cause regression of B-cell malignancies resistant to standard DLIs without causing GVHD.

Pure Red Cell Aplasia due to B19 Parvovirus Infection after Autologous Stem Cell Transplantation

PubMed Central

Tsirigotis, Panagiotis; Girkas, Konstantinos; Economopoulou, Christina; Bouchla, Anthoula; Papanicolaou, Nikolaos; Economopoulou, Panagiota; Papageorgiou, Sotirios; Pappa, Vassiliki; Dervenoulas, John

2011-01-01

Parvovirus B19 is recognized as a rare cause of pure red cell aplasia (PRCA) in allogeneic stem cell (SCT) and solid organ transplant patients. We report a patient with Hodgkin's disease who developed PRCA due to parvovirus B19 after autologous SCT and who had an excellent response after treatment with gamma-globulin. PMID:23198254

Hydroxyurea inhibits parvovirus B19 replication in erythroid progenitor cells.

PubMed

Bonvicini, Francesca; Bua, Gloria; Conti, Ilaria; Manaresi, Elisabetta; Gallinella, Giorgio

2017-07-15

Parvovirus B19 (B19V) infection is restricted to erythroid progenitor cells (EPCs) of the human bone marrow, leading to transient arrest of erythropoiesis and severe complications mainly in subjects with underlying hematological disorders or with immune system deficits. Currently, there are no specific antiviral drugs for B19V treatment, but identification of compounds inhibiting B19V replication can be pursued by a drug repositioning strategy. In this frame, the present study investigates the activity of hydroxyurea (HU), the only disease-modifying therapy approved for sickle cell disease (SCD), towards B19V replication in the two relevant cellular systems, the UT7/EpoS1 cell line and EPCs. Results demonstrate that HU inhibits B19V replication with EC 50 values of 96.2µM and 147.1µM in UT7/EpoS1 and EPCs, respectively, providing experimental evidence of the antiviral activity of HU towards B19V replication, and confirming the efficacy of a drug discovery process by drug repositioning strategy. The antiviral activity occurs in vitro at concentrations lower than those affecting cellular DNA replication and viability, and at levels measured in plasma samples of SCD patients undergoing HU therapy. HU might determine a dual beneficial effect on SCD patients, not only for the treatment of the disease but also towards a virus responsible for severe complications. Copyright © 2017 Elsevier Inc. All rights reserved.

The extended leukocyte differential count using the Cytodiff flow cytometric system reveals that higher CD16+ cytotoxic NK+T lymphocyte levels predict superior survival outcomes in patients with metastatic carcinoma.

PubMed

Park, Borae G; Park, Chan-Jeoung; Yoon, Chan-Hee; Jang, Seongsoo; Chi, Hyun-Sook; Ryu, Min-Hee; Kim, Sang-We

2013-05-01

The recently developed Cytodiff flow cytometric system (Beckman Coulter, Miami, FL) enables leukocyte analysis using a single immunophenotyping panel tube composed of six markers and five colors and that can detect 16 leukocyte subpopulations. We performed a preliminary investigation of whether changes in any of 16 leukocyte differentials were associated with survival and treatment outcomes in patients with metastatic carcinoma or not. We measured 16 leukocyte differential counts using the Cytodiff flow cytometric system in peripheral blood samples from 40 patients with metastatic malignancy (27 stomach cancer and 13 lung cancer) before chemotherapy and at 15 day intervals after chemotherapy for 2 months. A higher percentage of CD16+ cytotoxic NK+T lymphocytes was found to be the only significant prognostic factor among by Cox regression analysis and a higher percentage of CD16+ cytotoxic NK+T lymphocytes (>5.0%) showed significantly longer survival outcomes by Kaplan-Meier analysis (P = 0.003). The Cytodiff system enables 16 leukocyte subpopulations in a one tube assay and also can operate with only small amounts of sample, although it cannot differentiate NK cells from T lymphocytes. Hence, the monitoring of all leukocyte subpopulations using Cytodiff flow cytometry may be a helpful prognostic tool for patients with metastatic carcinoma. Copyright © 2012 International Clinical Cytometry Society.

Knockdown of Mediator Complex Subunit 19 Suppresses the Growth and Invasion of Prostate Cancer Cells

PubMed Central

Zhao, Hongwei; Lv, Wei; Chen, Jian; Wan, Fengchun; Liu, Dongfu; Gao, Zhenli; Wu, Jitao

2017-01-01

Prostate cancer (PCa) is one of the most common cancers in elderly men. Mediator Complex Subunit 19 (Med19) is overexpressed and plays promotional roles in many cancers. However, the roles of Med19 in PCa are still obscure. In this study, by using immunohistochemical staining, we found higher expression level of Med19 in PCa tissues than in adjacent benign prostate tissues. We then knocked down the Med19 expression in PCa cell lines LNCaP and PC3 by using lentivirus siRNA. Cell proliferation, anchor-independent growth, migration, and invasion were suppressed in Med19 knockdown PCa cells. In nude mice xenograft model, we found that Med19 knockdown PCa cells formed smaller tumors with lower proliferation index than did control cells. In the mechanism study, we found that Med19 could regulate genes involved in cell proliferation, cell cycle, and epithelial-mesenchymal transition, including P27, pAKT, pPI3K, IGF1R, E-Cadherin, N-Cadherin, Vimentin, ZEB2, Snail-1 and Snail-2. Targeting Med19 in PCa cells could inhibit the PCa growth and metastasis, and might be a therapeutic option for PCa in the future. PMID:28125713

Low susceptibility of NC/Nga mice to the lipopolysaccharide-mediated lethality with D-galactosamine sensitization and the involvement of fewer natural killer T cells.

PubMed

Koide, Naoki; Morikawa, Akiko; Odkhuu, Erdenezaya; Haque, Abedul; Badamtseren, Battuvshin; Naiki, Yoshikazu; Komatsu, Takayuki; Yoshida, Tomoaki; Yokochi, Takashi

2012-02-01

The LPS-mediated lethality of NC/Nga mice, having fewer NKT cells, was examined by using d-galactosamine (d-GalN)-sensitization. The NC/Nga mice were not killed by a simultaneous administration of d-GalN and LPS whereas all C57BL/6 (B6) control mice were killed. The injection of d-GalN and LPS failed to elevate the levels of serum alanine aminotransferase and caspase 3 in the liver tissues of NC/Nga mice. Further, the nitric oxide (NO) level of the d-GalN- and LPS-injected NC/Nga mice was much lower than those of the B6 mice. The expression of an inducible NO synthase (iNOS) was significantly reduced in the livers of NC/Nga mice. However, there was no significant difference in LPS-induced TNF-α production between B6 mice and NC/Nga mice. The NC/Nga mice had an impaired expression of IFN-γ protein and mRNA in response to d-GalN and LPS. The pretreatment with α-galactosylceramide (α-GalCer), which activates Vα14(+) NKT cells and induces the production of IFN-γ, rendered NC/Nga mice more susceptible to the LPS-mediated lethality. The livers of NC/Nga mice had fewer NKT cells compared to B6 mice. Taken together, it is suggested that the resistance of NC/Nga mice to the LPS-mediated lethality with d-GalN sensitization depended on the impaired IFN-γ production caused by fewer NKT cells and reduced NO production that followed.

Tailoring CD19xCD3-DART exposure enhances T-cells to eradication of B-cell neoplasms.

PubMed

Circosta, Paola; Elia, Angela Rita; Landra, Indira; Machiorlatti, Rodolfo; Todaro, Maria; Aliberti, Sabrina; Brusa, Davide; Deaglio, Silvia; Chiaretti, Sabina; Bruna, Riccardo; Gottardi, Daniela; Massaia, Massimo; Giacomo, Filomena Di; Guarini, Anna Rita; Foà, Robin; Kyriakides, Peter W; Bareja, Rohan; Elemento, Olivier; Chichili, Gurunadh R; Monteleone, Emanuele; Moore, Paul A; Johnson, Syd; Bonvini, Ezio; Cignetti, Alessandro; Inghirami, Giorgio

2018-01-01

Many patients with B-cell malignancies can be successfully treated, although tumor eradication is rarely achieved. T-cell-directed killing of tumor cells using engineered T-cells or bispecific antibodies is a promising approach for the treatment of hematologic malignancies. We investigated the efficacy of CD19xCD3 DART bispecific antibody in a broad panel of human primary B-cell malignancies. The CD19xCD3 DART identified 2 distinct subsets of patients, in which the neoplastic lymphocytes were eliminated with rapid or slow kinetics. Delayed responses were always overcome by a prolonged or repeated DART exposure. Both CD4 and CD8 effector cytotoxic cells were generated, and DART-mediated killing of CD4 + cells into cytotoxic effectors required the presence of CD8 + cells. Serial exposures to DART led to the exponential expansion of CD4 + and CD8 + cells and to the sequential ablation of neoplastic cells in absence of a PD-L1-mediated exhaustion. Lastly, patient-derived neoplastic B-cells (B-Acute Lymphoblast Leukemia and Diffuse Large B Cell Lymphoma) could be proficiently eradicated in a xenograft mouse model by DART-armed cytokine induced killer (CIK) cells. Collectively, patient tailored DART exposures can result in the effective elimination of CD19 positive leukemia and B-cell lymphoma and the association of bispecific antibodies with unmatched CIK cells represents an effective modality for the treatment of CD19 positive leukemia/lymphoma.

Donor-derived CD19-targeted T cells cause regression of malignancy persisting after allogeneic hematopoietic stem cell transplantation

PubMed Central

Dudley, Mark E.; Carpenter, Robert O.; Kassim, Sadik H.; Rose, Jeremy J.; Telford, William G.; Hakim, Frances T.; Halverson, David C.; Fowler, Daniel H.; Hardy, Nancy M.; Mato, Anthony R.; Hickstein, Dennis D.; Gea-Banacloche, Juan C.; Pavletic, Steven Z.; Sportes, Claude; Maric, Irina; Feldman, Steven A.; Hansen, Brenna G.; Wilder, Jennifer S.; Blacklock-Schuver, Bazetta; Jena, Bipulendu; Bishop, Michael R.; Gress, Ronald E.; Rosenberg, Steven A.

2013-01-01

New treatments are needed for B-cell malignancies persisting after allogeneic hematopoietic stem cell transplantation (alloHSCT). We conducted a clinical trial of allogeneic T cells genetically modified to express a chimeric antigen receptor (CAR) targeting the B-cell antigen CD19. T cells for genetic modification were obtained from each patient’s alloHSCT donor. All patients had malignancy that persisted after alloHSCT and standard donor lymphocyte infusions (DLIs). Patients did not receive chemotherapy prior to the CAR T-cell infusions and were not lymphocyte depleted at the time of the infusions. The 10 treated patients received a single infusion of allogeneic anti-CD19-CAR T cells. Three patients had regressions of their malignancies. One patient with chronic lymphocytic leukemia (CLL) obtained an ongoing complete remission after treatment with allogeneic anti-CD19-CAR T cells, another CLL patient had tumor lysis syndrome as his leukemia dramatically regressed, and a patient with mantle cell lymphoma obtained an ongoing partial remission. None of the 10 patients developed graft-versus-host disease (GVHD). Toxicities included transient hypotension and fever. We detected cells containing the anti-CD19-CAR gene in the blood of 8 of 10 patients. These results show for the first time that donor-derived allogeneic anti-CD19-CAR T cells can cause regression of B-cell malignancies resistant to standard DLIs without causing GVHD. This trial was registered at www.clinicaltrials.gov as #NCT01087294. PMID:24055823

Selective resistance of CD44hi T cells to p53-dependent cell death results in persistence of immunologic memory after total body irradiation.

PubMed

Yao, Zhenyu; Jones, Jennifer; Kohrt, Holbrook; Strober, Samuel

2011-10-15

Our previous studies showed that treatment of mice with total body irradiation (TBI) or total lymphoid tissue irradiation markedly changes the balance of residual T cell subsets to favor CD4(+)CD44(hi) NKT cells because of the differential resistance of the latter subset to cell death. The object of the current study was to further elucidate the changed balance and mechanisms of differential radioresistance of T cell subsets after graded doses of TBI. The experimental results showed that CD4(+) T cells were markedly more resistant than CD8(+) T cells, and CD44(hi) T cells, including NKT cells and memory T cells, were markedly more resistant than CD44(lo) (naive) T cells. The memory T cells immunized to alloantigens persisted even after myeloablative (1000 cGy) TBI and were able to prevent engraftment of bone marrow transplants. Although T cell death after 1000 cGy was prevented in p53(-/-) mice, there was progressive T cell death in p53(-/-) mice at higher doses. Although p53-dependent T cell death changed the balance of subsets, p53-independent T cell death did not. In conclusion, resistance of CD44(hi) T cells to p53-dependent cell death results in the persistence of immunological memory after TBI and can explain the immune-mediated rejection of marrow transplants in sensitized recipients.

Adoptive transfer of syngeneic T cells transduced with a chimeric antigen receptor that recognizes murine CD19 can eradicate lymphoma and normal B cells.

PubMed

Kochenderfer, James N; Yu, Zhiya; Frasheri, Dorina; Restifo, Nicholas P; Rosenberg, Steven A

2010-11-11

Adoptive T-cell therapy with anti-CD19 chimeric antigen receptor (CAR)-expressing T cells is a new approach for treating advanced B-cell malignancies. To evaluate anti-CD19-CAR-transduced T cells in a murine model of adoptive T-cell therapy, we developed a CAR that specifically recognized murine CD19. We used T cells that were retrovirally transduced with this CAR to treat mice bearing a syngeneic lymphoma that naturally expressed the self-antigen murine CD19. One infusion of anti-CD19-CAR-transduced T cells completely eliminated normal B cells from mice for at least 143 days. Anti-CD19-CAR-transduced T cells eradicated intraperitoneally injected lymphoma cells and large subcutaneous lymphoma masses. The antilymphoma efficacy of anti-CD19-CAR-transduced T cells was critically dependent on irradiation of mice before anti-CD19-CAR-transduced T-cell infusion. Anti-CD19-CAR-transduced T cells had superior antilymphoma efficacy compared with the anti-CD19 monoclonal antibody from which the anti-CD19 CAR was derived. Our results demonstrated impressive antilymphoma activity and profound destruction of normal B cells caused by anti-CD19-CAR-transduced T cells in a clinically relevant murine model.

Parvovirus B19 Replication and Expression in Differentiating Erythroid Progenitor Cells

PubMed Central

Bua, Gloria; Manaresi, Elisabetta; Bonvicini, Francesca; Gallinella, Giorgio

2016-01-01

The pathogenic Parvovirus B19 (B19V) is characterized by a strict adaptation to erythroid progenitor cells (EPCs), a heterogeneous population of differentiating cells with diverse phenotypic and functional properties. In our work, we studied the dynamics of B19V infection in EPCs in dependence on the cell differentiation stage, in terms of distribution of infected cells, synthesis of viral nucleic acids and production of infectious virus. EPCs at early differentiation stage led to an abortive infection, without viral genome replication and a very low transcriptional activity. EPCs at later stages were permissive, with highest levels of viral replicative activity at day 9 (+3.0 Log from 2 to 48 hpi) and lower levels at day 18 (+1.5 Log from 2 to 48 hpi). B19V DNA increment was in accordance with the percentage of cells positive to flow-FISH assay (41.4% at day 9, 1.1% at day 18). Quantitation of total RNA indicated a close association of genome replication and transcription with viral RNA accumulation within infected cells related to viral DNA increase during the course of infection. Analysis of the different classes of mRNAs revealed two distinct pattern of genome expression profile with a fine regulation in the frequency utilization of RNA processing signals: an early phase, when cleavage at the proximal site leading to a higher relative production of mRNA for NS protein, and a late phase, when cleavage at the distal site was more frequent leading to higher relative abundance of mRNA for VP and 11 kDA proteins. Infectious virus was released from cells at day 6–15, but not at day 18. Our results, providing a detailed description of B19V replication and expression profile in differentiating EPCs, highlight the very tight adaptation of B19V to a specific cellular target defined both by its erythroid lineage and its differentiation stage. PMID:26845771

Elevated expression of pleiotrophin in lymphocytic leukemia CD19+ B cells.

PubMed

Du, Chun-Xian; Wang, Lan; Li, Yan; Xiao, Wei; Guo, Qin-Lian; Chen, Fei; Tan, Xin-Ti

2014-10-01

Pleiotrophin (PTN) has been demonstrated to be strongly expressed in many fetal tissues, but seldom in healthy adult tissues. While PTN has been reported to be expressed in many types of tumors as well as at high serum concentrations in patients with many types of cancer, to date, there has been no report that PTN is expressed in leukemia, especially in lymphocytic leukemia. We isolated the CD19(+) subset of B cells from peripheral blood from healthy adults, B-cell acute lymphocytic leukemia (B-ALL) patients, and B-cell chronic lymphocytic leukemia (B-CLL) patients and examined these cells for PTN mRNA and protein expression. We used immunocytochemistry, western blotting, and enzyme-linked immunosorbent assay to show that PTN protein is highly expressed in CD19(+) B cells from B-ALL and B-CLL patients, but barely expressed in B cells from healthy adults. We also examined PTN expression at the nucleic acid level using reverse transcription polymerase chain reaction (RT-PCR) and northern blotting and detected a high levels of PTN transcripts in the CD19(+) B cells from both groups of leukemia patients, but very few in the CD19(+) B cells from the healthy controls. Interestingly, the quantity of the PTN transcripts correlated with the severity of disease. Moreover, suppression of PTN activity with an anti-PTN antibody promoted apoptosis of cells from leukemia patients and cell lines SMS-SB and JVM-2. This effect of the anti-PTN antibody suggests that PTN may be a new target for the treatment of lymphocytic leukemia. © 2014 APMIS. Published by John Wiley & Sons Ltd.

High-dose therapy and autologous stem cell transplantation for extra-nodal NK/T lymphoma in patients from the Western hemisphere: a study from the European Society for Blood and Marrow Transplantation.

PubMed

Fox, Christopher P; Boumendil, Ariane; Schmitz, Norbert; Finel, Herve; Luan, Jian J; Sucak, Gülsan; Blaise, Didier; Finke, Jürgen; Pflüger, Karl-Heinz; Veelken, Hendrik; Gorin, Norbert-Claude; Poiré, Xavier; Ganser, Arnold; Dreger, Peter; Sureda, Anna

2015-01-01

Extra-nodal NK/T lymphoma (ENKTL) is rare and more frequently encountered in East Asia. The role of high-dose therapy and autologous stem cell transplantation (HDT-ASCT) for ENKTL is unclear. Twenty-eight evaluable patients who had undergone HDT-ASCT in Europe from 2000-2009 were studied. The median age was 47 years and patients had received a median of two lines of prior therapy. Some 57% of patients were not in complete remission or beyond first complete remission at HDT-ASCT. The 1-year non-relapse mortality (NRM) was 11%; 2-year progression-free survival (PFS) and overall survival (OS) rates were 41% and 52%, respectively. Notably, the 2-year PFS and OS for those with stage III/IV disease were 33% and 40%, respectively, with no relapses beyond 1-year post-HDT-ASCT. This is the largest analysis of HDT-ASCT for patients with ENKTL reported from the Western hemisphere. Survival is comparable to East Asian cohorts and outcomes are encouraging for patients with advanced disease.

CD4+ T cell-mediated cytotoxicity is associated with MHC class II expression on malignant CD19+ B cells in diffuse large B cell lymphoma.

PubMed

Zhou, Yong; Zha, Jie; Lin, Zhijuan; Fang, Zhihong; Zeng, Hanyan; Zhao, Jintao; Luo, Yiming; Li, Zhifeng; Xu, Bing

2018-01-15

Diffuse large B cell lymphoma (DLBCL) is a common B cell malignancy with approximately 30% of patients present relapsed or refractory disease after first-line therapy. Research of further treatment options is needed. Cytotoxic CD4 + T cells express cytolytic molecules and have potential antitumor function. Here, we showed that the CD19 + cells from DLBCL patients presented significantly reduced expression of MHC II molecules than those from healthy controls. Three years after the first-line treatment, patients that presented relapsed disease had significantly lower MHC II expression on their CD19 + cells than patients who did not show recurrence. Examining cytotoxic CD4 + T cells show that DLBCL patients presented significantly elevated frequencies of granzyme A-, granzyme B-, and/or perforin-expressing cytotoxic CD4 + T cells. Also, frequency of cytotoxic CD4 + T cells in DLBCL patients was positively correlated with the MHC II expression level. Subsequently, the cytotoxic potential of CD4 + T cells against autologous CD19 + cells was investigated. We found that the cytotoxic potential of CD4 + T cells was highest in MHC II-high, intermediate in MHC II-mid, and lowest in MHC II-low patients. The percentage of MHC II-expressing viable CD19 + cells presented a significant reduction after longer incubation with cytotoxic CD4 + T cells, suggesting that cytotoxic CD4 + T cells preferentially eliminated MHC II-expressing CD19 + cells. Blocking MHC II on CD19 + cells significantly reduced the cytolytic capacity of CD4 + T cells. Despite these discoveries, the frequency of cytotoxic CD4 + T cells did not predict the clinical outcome of DLBCL patients. Together, these results demonstrated that cytotoxic CD4 + T cells presented an MHC II-dependent cytotoxic potential against autologous CD19 + cells and could potentially represent a future treatment option for DLBCL. Copyright © 2017 Elsevier Inc. All rights reserved.

Innate NKTγδ and NKTαβ cells exert similar functions and compete for a thymic niche.

PubMed

Pereira, Pablo; Boucontet, Laurent

2012-05-01

The transcriptional regulator promyelocytic leukemia zinc finger (PLZF) is highly expressed during the differentiation of natural killer T (NKT) cells and is essential for the acquisition of their effector/memory innate-like phenotype. Staining with anti-PLZF and anti-NK1.1 Abs allows the definition of two subsets of NKTαβ and NKTγδ thymocytes that differ phenotypically and functionally: a PLZF(+) NK1.1(-) subset composed of mostly quiescent cells that secrete more IL-4 than IFN-γ upon activation and a PLZF(+/-) NK1.1(+) subset that expresses CD127, NK1.1, and other NK-cell markers, secrete more IFN-γ than IL-4 upon activation and contains a sizable fraction of dividing cells. The size of the NK1.1(+) population is very tightly regulated and NK1.1(+) αβ and γδ thymocytes compete for a thymic niche. Furthermore, the relative representation of the PLZF(+) and NK1.1(+) subsets varies in a strain-specific manner with C57BL/6 (B6) mice containing more NK1.1(+) cells and (B6 × DBA/2)F1 (B6D2F1) mice more PLZF(+) cells. Consequently, activation of NKT cells in vivo is expected to result in higher levels of IL-4 secreted in B6D2F1 mice than in B6 mice. Consistent with this possibility, B6D2F1 mice, when compared with B6 mice, contain more "innate" CD8(+) thymocytes, the generation of which depends on IL-4 secreted by NKT cells. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Selection of a novel CD19 aptamer for targeted delivery of doxorubicin to lymphoma cells.

PubMed

Hu, Yan; Li, Xiaoou; An, Yacong; Duan, Jinhong; Yang, Xian-Da

2018-06-01

CD19 is overexpressed in most human B cell malignancies and considered an important tumor marker for diagnosis and treatment. Aptamers are oligonucleotides that may potentially serve as tumor-homing ligand for targeted cancer therapy with excellent affinity and specificity. In this study, we selected a novel CD19 aptamer (LC1) that was a 59-nucleotide single strand DNA. The aptamer could bind to recombinant CD19 protein with a K d of 85.4 nM, and had minimal cross reactivity to bovine serum albumin (BSA) or ovalbumin (OVA). Moreover, the aptamer was found capable of binding with the CD19-positive lymphoma cells (Ramos and Raji), but not the CD19-negative cell lines (Jurkat and NB4). An aptamer-doxorubicin complex (Apt-Dox) was also formulated, and selectively delivered doxorubicin to CD19-positive lymphoma cells in vitro . The results indicate that aptamer LC1 can recognize CD19-positive tumor cells and may potentially function as a CD19-targeting ligand.

Ligation of TLR7 on CD19(+) CD1d(hi) B cells suppresses allergic lung inflammation via regulatory T cells.

PubMed

Khan, Adnan R; Amu, Sylvie; Saunders, Sean P; Hams, Emily; Blackshields, Gordon; Leonard, Martin O; Weaver, Casey T; Sparwasser, Tim; Sheils, Orla; Fallon, Padraic G

2015-06-01

B cells have been described as having the capacity to regulate cellular immune responses and suppress inflammatory processes. One such regulatory B-cell population is defined as IL-10-producing CD19(+) CD1d(hi) cells. Previous work has identified an expansion of these cells in mice infected with the helminth, Schistosoma mansoni. Here, microarray analysis of CD19(+) CD1d(hi) B cells from mice infected with S. mansoni demonstrated significantly increased Tlr7 expression, while CD19(+) CD1d(hi) B cells from uninfected mice also demonstrated elevated Tlr7 expression. Using IL-10 reporter, Il10(-/-) and Tlr7(-/-) mice, we formally demonstrate that TLR7 ligation of CD19(+) CD1d(hi) B cells increases their capacity to produce IL-10. In a mouse model of allergic lung inflammation, the adoptive transfer of TLR7-elicited CD19(+) CD1d(hi) B cells reduced airway inflammation and associated airway hyperresponsiveness. Using DEREG mice to deplete FoxP3(+) T regulatory cells in allergen-sensitized mice, we show that that TLR7-elicited CD19(+) CD1d(hi) B cells suppress airway hyperresponsiveness via a T regulatory cell dependent mechanism. These studies identify that TLR7 stimulation leads to the expansion of IL-10-producing CD19(+) CD1d(hi) B cells, which can suppress allergic lung inflammation via T regulatory cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Long Noncoding RNA H19 Inhibits Cell Viability, Migration, and Invasion Via Downregulation of IRS-1 in Thyroid Cancer Cells

PubMed Central

Wang, Peng; Xu, Weimin; Liu, Haixia; Bu, Qingao; Sun, Diwen

2017-01-01

Thyroid cancer is a common endocrine gland malignancy which exhibited rapid increased incidence worldwide in recent decades. This study was aimed to investigate the role of long noncoding RNA H19 in thyroid cancer. Long noncoding RNA H19 was overexpressed or knockdown in thyroid cancer cells SW579 and TPC-1, and the expression of long noncoding RNA H19 was detected by real-time polymerase chain reaction. The cell viability, migration, and invasion were determined by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide assay, Transwell assay, and wound healing assay, respectively. Furthermore, cell apoptosis was analyzed by flow cytometry, and expressions of some factors that were related to phosphatidyl inositide 3-kinases/protein kinase B and nuclear factor κB signal pathway were measured by Western blotting. This study revealed that cell viability and migration/invasion of SW579 and TPC-1 were significantly decreased by long noncoding RNA H19 overexpression compared with the control group (P < .05), whereas cell apoptosis was statistically increased (P < .001). Meanwhile, cell viability and migration/invasion were significantly increased after long noncoding RNA H19 knockdown (P < .05). Furthermore, long noncoding RNA H19 negatively regulated the expression of insulin receptor substrate 1 and thus effect on cell proliferation and apoptosis. Insulin receptor substrate 1 regulated the activation of phosphatidyl inositide 3-kinases/AKT and nuclear factor κB signal pathways. In conclusion, long noncoding RNA H19 could suppress cell viability, migration, and invasion via downregulation of insulin receptor substrate 1 in SW579 and TPC-1 cells. These results suggested the important role of long noncoding RNA H19 in thyroid cancer, and long noncoding RNA H19 might be a potential target of thyroid cancer treatment. PMID:29332545

IL-21 Promotes Late Activator APC-Mediated T Follicular Helper Cell Differentiation in Experimental Pulmonary Virus Infection

PubMed Central

Yoo, Jae-Kwang; Braciale, Thomas J.

2014-01-01

IL-21 is a type-I cytokine that has pleiotropic immuno-modulatory effects. Primarily produced by activated T cells including NKT and TFH cells, IL-21 plays a pivotal role in promoting TFH differentiation through poorly understood cellular and molecular mechanisms. Here, employing a mouse model of influenza A virus (IAV) infection, we demonstrate that IL-21, initially produced by NKT cells, promotes TFH differentiation by promoting the migration of late activator antigen presenting cell (LAPC), a recently identified TFH inducer, from the infected lungs into the draining lymph nodes (dLN). LAPC migration from IAV-infected lung into the dLN is CXCR3-CXCL9 dependent. IL-21-induced TNF-α production by conventional T cells is critical to stimulate CXCL9 expression by DCs in the dLN, which supports LAPC migration into the dLN and ultimately facilitates TFH differentiation. Our results reveal a previously unappreciated mechanism for IL-21 modulation of TFH responses during respiratory virus infection. PMID:25251568

Bisphenol A downregulates CYP19 transcription in JEG-3 cells.

PubMed

Huang, Hui; Leung, Lai K

2009-09-28

Bisphenol A is an industrial contaminant and is considered to be an endocrine disruptor; its estrogenic property has been reported in many studies. Because of its ubiquitous existence in our environment, bisphenol A has drawn much discussion on its safety issues. Estrogen is important in the maintenance of human pregnancy, and the placenta is the major site of synthesis during this period of time. Aromatase or CYP19 catalyses the conversion of estrogen from its precursor, and is highly expressed in placental cells. In the present study, we examined the ability of the toxicant in suppressing the transcription of CYP19 in JEG-3 cells. Cells treated with bisphenol A displayed a reduced aromatase activity. Real-time PCR analysis indicated that 5muM of the compound significantly reduced the mRNA expression in these cells. As the transcriptional activity of CYP19 gene is controlled by the proximal promoter region of exon I.1 in placental cells, the promoter activity of this gene fragment and exon-I.1-spliced mRNA abundance were also evaluated. Both results indicated that bisphenol A repressed the transcriptional control of promoter I.1. The present study showed that bisphenol A potentially reduced estrogen synthesis by downregulating CYP of placental cells. This information could be useful for evaluating the exposure limit of bisphenol A.

Fluorine-19 MRI Contrast Agents for Cell Tracking and Lung Imaging

PubMed Central

Fox, Matthew S.; Gaudet, Jeffrey M.; Foster, Paula J.

2015-01-01

Fluorine-19 (19F)-based contrast agents for magnetic resonance imaging stand to revolutionize imaging-based research and clinical trials in several fields of medical intervention. First, their use in characterizing in vivo cell behavior may help bring cellular therapy closer to clinical acceptance. Second, their use in lung imaging provides novel noninvasive interrogation of the ventilated airspaces without the need for complicated, hard-to-distribute hardware. This article reviews the current state of 19F-based cell tracking and lung imaging using magnetic resonance imaging and describes the link between the methods across these fields and how they may mutually benefit from solutions to mutual problems encountered when imaging 19F-containing compounds, as well as hardware and software advancements. PMID:27042089

Detection of invariant natural killer T cells in ejaculates from infertile patients with chronic inflammation of genital tract.

PubMed

Duan, Yong-Gang; Chen, Shujian; Haidl, Gerhard; Allam, Jean-Pierre

2017-08-01

Chronic inflammation of genital tract is thought to play a major role in male fertility disorder. Natural killer (NK) T cells are a heterogeneous group of T cells that share properties of both T cells and NK cells which display immunoregulatory properties. However, little is known regarding the presence and function of NK T cells in ejaculates from patients with chronic inflammation of genital tract. Invariant NK T (iNK T) cells were detected by invariant (Vα24-JαQ) TCR chain in ejaculates from patients suffering from chronic inflammation of genital tract (CIGT) using flow cytometry and immunofluorescence of double staining (n=40). Inflammatory cytokines interleukin (IL)-6, IL-17, and IFN-γ were detected in cell-free seminal plasma using an enzyme-linked immunosorbent assay (ELISA). The correlation between the percentage of iNK T cells and spermatozoa count, motility, vitality, seminal IL-6, IL-17, and IFN-γ was investigated. Significant percentages of iNK T cells above 10% were detected in 50% (CIGT-NKT + group). A negative correlation was detected between the percentage of iNK T cells and spermatozoa count (r=-.5957, P=.0056), motility (r=-.6163, P=.0038), and vitality (r=-.8032, P=.0019) in CIGT-NKT + group (n=20). Interestingly, a significant correlation of iNK T cells to seminal IL-6 (r=.7083, P=.0005), IFN-γ (r=.9578, P<.0001) was detected whereas lack of correlation between iNK T cells and IL-17 (r=-.1557, P=.5122) in CIGT-NKT + group. The proliferative response of iNK T cells could accompany an inflammatory response to spermatozoa and consequently influence sperm quality through secretion of IFN-γ but not IL-17 under chronic inflammatory condition. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Peripheral killer cells do not differentiate between asthma patients with or without fixed airway obstruction.

PubMed

Tubby, Carolyn; Negm, Ola H; Harrison, Timothy; Tighe, Patrick J; Todd, Ian; Fairclough, Lucy C

2017-06-01

The three main types of killer cells - CD8 + T cells, NK cells and NKT cells - have been linked to asthma and chronic obstructive pulmonary disease (COPD). However, their role in a small subset of asthma patients displaying fixed airway obstruction (FAO), similar to that seen in COPD, has not been explored. The objective of the present study was to investigate killer cell numbers, phenotype and function in peripheral blood from asthma patients with FAO, asthma patients without FAO, and healthy individuals. Peripheral CD8 + T cells (CD8 + CD3 + CD56 - ), NK cells (CD56 + CD3 - ) and NKT-like cells (CD56 + CD3 + ) of 14 asthma patients with FAO (post-bronchodilator FEV/FVC <0.7, despite clinician-optimised treatment), 7 asthma patients without FAO (post-bronchodilator FEV/FVC ≥ 0.7), and 9 healthy individuals were studied. No significant differences were seen between the number, receptor expression, MAPK signalling molecule expression, cytotoxic mediator expression, and functional cytotoxicity of peripheral killer cells from asthma patients with FAO, asthma patients without FAO and healthy individuals. Peripheral killer cell numbers or functions do not differentiate between asthma patients with or without fixed airway obstruction.

Ordering human CD34+CD10−CD19+ pre/pro-B-cell and CD19− common lymphoid progenitor stages in two pro-B-cell development pathways

PubMed Central

Sanz, Eva; Muñoz-A., Norman; Monserrat, Jorge; Van-Den-Rym, Ana; Escoll, Pedro; Ranz, Ismael; Álvarez-Mon, Melchor; de-la-Hera, Antonio

2010-01-01

Studies here respond to two long-standing questions: Are human “pre/pro-B” CD34+CD10−CD19+ and “common lymphoid progenitor (CLP)/early-B” CD34+CD10+CD19− alternate precursors to “pro-B” CD34+CD19+CD10+ cells, and do the pro-B cells that arise from these progenitors belong to the same or distinct B-cell development pathways? Using flow cytometry, gene expression profiling, and Ig VH-D-JH sequencing, we monitor the initial 10 generations of development of sorted cord blood CD34highLineage− pluripotential progenitors growing in bone marrow S17 stroma cocultures. We show that (i) multipotent progenitors (CD34+CD45RA+CD10−CD19−) directly generate an initial wave of Pax5+TdT− “unilineage” pre/pro-B cells and a later wave of “multilineage” CLP/early-B cells and (ii) the cells generated in these successive stages act as precursors for distinct pro-B cells through two independent layered pathways. Studies by others have tracked the origin of B-lineage leukemias in elderly mice to the mouse B-1a pre/pro-B lineage, which lacks the TdT activity that diversifies the VH-D-JH Ig heavy chain joints found in the early-B or B-2 lineage. Here, we show a similar divergence in human B-cell development pathways between the Pax5+TdT− pre/pro-B differentiation pathway that gives rise to infant B-lineage leukemias and the early-B pathway. PMID:20231472

The Addition of the BTK Inhibitor Ibrutinib to Anti-CD19 Chimeric Antigen Receptor T Cells (CART19) Improves Responses against Mantle Cell Lymphoma.

PubMed

Ruella, Marco; Kenderian, Saad S; Shestova, Olga; Fraietta, Joseph A; Qayyum, Sohail; Zhang, Qian; Maus, Marcela V; Liu, Xiaobin; Nunez-Cruz, Selene; Klichinsky, Michael; Kawalekar, Omkar U; Milone, Michael; Lacey, Simon F; Mato, Anthony; Schuster, Stephen J; Kalos, Michael; June, Carl H; Gill, Saar; Wasik, Mariusz A

2016-06-01

Responses to therapy with chimeric antigen receptor T cells recognizing CD19 (CART19, CTL019) may vary by histology. Mantle cell lymphoma (MCL) represents a B-cell malignancy that remains incurable despite novel therapies such as the BTK inhibitor ibrutinib, and where data from CTL019 therapy are scant. Using MCL as a model, we sought to build upon the outcomes from CTL019 and from ibrutinib therapy by combining these in a rational manner. MCL cell lines and primary MCL samples were combined with autologous or normal donor-derived anti-CD19 CAR T cells along with ibrutinib. The effect of the combination was studied in vitro and in mouse xenograft models. MCL cells strongly activated multiple CTL019 effector functions, and MCL killing by CTL019 was further enhanced in the presence of ibrutinib. In a xenograft MCL model, we showed superior disease control in the CTL019- as compared with ibrutinib-treated mice (median survival not reached vs. 95 days, P < 0.005) but most mice receiving CTL019 monotherapy eventually relapsed. Therefore, we added ibrutinib to CTL019 and showed that 80% to 100% of mice in the CTL019 + ibrutinib arm and 0% to 20% of mice in the CTL019 arm, respectively, remained in long-term remission (P < 0.05). Combining CTL019 with ibrutinib represents a rational way to incorporate two of the most recent therapies in MCL. Our findings pave the way to a two-pronged therapeutic strategy in patients with MCL and other types of B-cell lymphoma. Clin Cancer Res; 22(11); 2684-96. ©2016 AACR. ©2016 American Association for Cancer Research.

Arsenic inhibits hedgehog signaling during P19 cell differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Jui Tung; Bain, Lisa J., E-mail: lbain@clemson.edu; Department of Biological Sciences, Clemson University, 132 Long Hall, Clemson, SC 29634

Arsenic is a toxicant found in ground water around the world, and human exposure mainly comes from drinking water or from crops grown in areas containing arsenic in soils or water. Epidemiological studies have shown that arsenic exposure during development decreased intellectual function, reduced birth weight, and altered locomotor activity, while in vitro studies have shown that arsenite decreased muscle and neuronal cell differentiation. The sonic hedgehog (Shh) signaling pathway plays an important role during the differentiation of both neurons and skeletal muscle. The purpose of this study was to investigate whether arsenic can disrupt Shh signaling in P19 mousemore » embryonic stem cells, leading to changes muscle and neuronal cell differentiation. P19 embryonic stem cells were exposed to 0, 0.25, or 0.5 μM of sodium arsenite for up to 9 days during cell differentiation. We found that arsenite exposure significantly reduced transcript levels of genes in the Shh pathway in both a time and dose-dependent manner. This included the Shh ligand, which was decreased 2- to 3-fold, the Gli2 transcription factor, which was decreased 2- to 3-fold, and its downstream target gene Ascl1, which was decreased 5-fold. GLI2 protein levels and transcriptional activity were also reduced. However, arsenic did not alter GLI2 primary cilium accumulation or nuclear translocation. Moreover, additional extracellular SHH rescued the inhibitory effects of arsenic on cellular differentiation due to an increase in GLI binding activity. Taken together, we conclude that arsenic exposure affected Shh signaling, ultimately decreasing the expression of the Gli2 transcription factor. These results suggest a mechanism by which arsenic disrupts cell differentiation. - Highlights: • Arsenic exposure decreases sonic hedgehog pathway-related gene expression. • Arsenic decreases GLI2 protein levels and transcriptional activity in P19 cells. • Arsenic exposure does not alter the levels of SHH

The Tol2 transposon system mediates the genetic engineering of T-cells with CD19-specific chimeric antigen receptors for B-cell malignancies.

PubMed

Tsukahara, T; Iwase, N; Kawakami, K; Iwasaki, M; Yamamoto, C; Ohmine, K; Uchibori, R; Teruya, T; Ido, H; Saga, Y; Urabe, M; Mizukami, H; Kume, A; Nakamura, M; Brentjens, R; Ozawa, K

2015-02-01

Engineered T-cell therapy using a CD19-specific chimeric antigen receptor (CD19-CAR) is a promising strategy for the treatment of advanced B-cell malignancies. Gene transfer of CARs to T-cells has widely relied on retroviral vectors, but transposon-based gene transfer has recently emerged as a suitable nonviral method to mediate stable transgene expression. The advantages of transposon vectors compared with viral vectors include their simplicity and cost-effectiveness. We used the Tol2 transposon system to stably transfer CD19-CAR into human T-cells. Normal human peripheral blood lymphocytes were co-nucleofected with the Tol2 transposon donor plasmid carrying CD19-CAR and the transposase expression plasmid and were selectively propagated on NIH3T3 cells expressing human CD19. Expanded CD3(+) T-cells with stable and high-level transgene expression (~95%) produced interferon-γ upon stimulation with CD19 and specifically lysed Raji cells, a CD19(+) human B-cell lymphoma cell line. Adoptive transfer of these T-cells suppressed tumor progression in Raji tumor-bearing Rag2(-/-)γc(-/-) immunodeficient mice compared with control mice. These results demonstrate that the Tol2 transposon system could be used to express CD19-CAR in genetically engineered T-cells for the treatment of refractory B-cell malignancies.

Ex vivo Akt inhibition promotes the generation of potent CD19CAR T cells for adoptive immunotherapy.

PubMed

Urak, Ryan; Walter, Miriam; Lim, Laura; Wong, ChingLam W; Budde, Lihua E; Thomas, Sandra; Forman, Stephen J; Wang, Xiuli

2017-01-01

Insufficient persistence and effector function of chimeric antigen receptor (CAR)-redirected T cells have been challenging issues for adoptive T cell therapy. Generating potent CAR T cells is of increasing importance in the field. Studies have demonstrated the importance of the Akt pathway in the regulation of T cell differentiation and memory formation. We now investigate whether inhibition of Akt signaling during ex vivo expansion of CAR T cells can promote the generation of CAR T cells with enhanced antitumor activity following adoptive therapy in a murine leukemia xenograft model. Various T cell subsets including CD8+ T cells, bulk T cells, central memory T cells and naïve/memory T cells were isolated from PBMC of healthy donors, activated with CD3/CD28 beads, and transduced with a lentiviral vector encoding a second-generation CD19CAR containing a CD28 co-stimulatory domain. The transduced CD19CAR T cells were expanded in the presence of IL-2 (50U/mL) and Akt inhibitor (Akti) (1 μM) that were supplemented every other day. Proliferative/expansion potential, phenotypical characteristics and functionality of the propagated CD19CAR T cells were analyzed in vitro and in vivo after 17-21 day ex vivo expansion. Anti-tumor activity was evaluated after adoptive transfer of the CD19CAR T cells into CD19+ tumor-bearing immunodeficient mice. Tumor signals were monitored with biophotonic imaging, and survival rates were analyzed by the end of the experiments. We found that Akt inhibition did not compromise CD19CAR T cell proliferation and expansion in vitro, independent of the T cell subsets, as comparable CD19CAR T cell expansion was observed after culturing in the presence or absence of Akt inhibitor. Functionally, Akt inhibition did not dampen cell-mediated effector function, while Th1 cytokine production increased. With respect to phenotype, Akti-treated CD19CAR T cells expressed higher levels of CD62L and CD28 as compared to untreated CD19CAR T cells. Once

Infusion of donor-derived CD19-redirected virus-specific T cells for B-cell malignancies relapsed after allogeneic stem cell transplant: a phase 1 study.

PubMed

Cruz, Conrad Russell Y; Micklethwaite, Kenneth P; Savoldo, Barbara; Ramos, Carlos A; Lam, Sharon; Ku, Stephanie; Diouf, Oumar; Liu, Enli; Barrett, A John; Ito, Sawa; Shpall, Elizabeth J; Krance, Robert A; Kamble, Rammurti T; Carrum, George; Hosing, Chitra M; Gee, Adrian P; Mei, Zhuyong; Grilley, Bambi J; Heslop, Helen E; Rooney, Cliona M; Brenner, Malcolm K; Bollard, Catherine M; Dotti, Gianpietro

2013-10-24

Autologous T cells expressing a CD19-specific chimeric antigen receptor (CD19.CAR) are active against B-cell malignancies, but it is unknown whether allogeneic CD19.CAR T cells are safe or effective. After allogeneic hematopoietic stem cell transplantation (HSCT), infused donor-derived virus-specific T cells (VSTs) expand in vivo, persist long term, and display antiviral activity without inducing graft-vs-host disease; therefore, we determined whether donor VSTs, engineered to express CD19.CAR, retained the characteristics of nonmanipulated allogeneic VSTs while gaining antitumor activity. We treated 8 patients with allogeneic (donor-derived) CD19.CAR-VSTs 3 months to 13 years after HSCT. There were no infusion-related toxicities. VSTs persisted for a median of 8 weeks in blood and up to 9 weeks at disease sites. Objective antitumor activity was evident in 2 of 6 patients with relapsed disease during the period of CD19.CAR-VST persistence, whereas 2 patients who received cells while in remission remain disease free. In 2 of 3 patients with viral reactivation, donor CD19.CAR-VSTs expanded concomitantly with VSTs. Hence CD19.CAR-VSTs display antitumor activity and, because their number may be increased in the presence of viral stimuli, earlier treatment post-HSCT (when lymphodepletion is greater and the incidence of viral infection is higher) or planned vaccination with viral antigens may enhance disease control.

Skeletal and cardiac myogenesis accompany adipogenesis in P19 embryonal stem cells.

PubMed

Bouchard, Frédéric; Paquin, Joanne

2009-09-01

P19 embryonic carcinoma cells resemble normal embryonic stem (ES) cells. They generate cardiac and skeletal myocytes in response to retinoic acid (RA) or oxytocin (OT). RA treatment followed by exposure to triiodothyronine (T3) and insulin induces ES cells differentiation into adipocytes and skeletomyocytes. On the other hand, OT (10(-7) M) was reported to inhibit 3T3 preadipocyte maturation. The present work was undertaken to determine whether P19 cells have an adipogenic potential that could be affected by OT. Cells were treated with RA (10(-6) M)/T3+insulin (adipogenic protocol) or 10(-7) M OT (cardiomyogenic protocol), and analyzed by polymerase chain reaction, immunotechniques, and cytochemistry. Oil-Red-O staining and expression of peroxisome proliferator-activated receptor-gamma (PPARgamma) and aP2 indicated the generation of adipocytes in cultures submitted to the adipogenic protocol. Contracting cells were also generated. Cells positive for sarcomeric actinin and negative for cardiac troponin inhibitor (cTpnI) indicated generation of skeletomyocytes, and cTpnI positive cells revealed generation of cardiomyocytes. Levels of cTpnI and of the skeletal marker MyoD were almost similar in both protocols, whereas no Oil-Red-O staining was associated with the cardiomyogenic protocol. Addition of 10(-7) M OT to the adipogenic protocol did not affect Oil-Red-O staining and PPARgamma expression. Interestingly, Oct3/4 pluripotency marker disappeared in the adipogenic protocol but remained expressed in the cardiomyogenic one. P19 cells thus have an adipogenic potential non affected by 10(-7) M OT. RA/T3+insulin combination generates a larger spectrum of mesodermal cell derivatives and is a more potent morphogenic treatment than OT. P19 cells could help investigating mechanisms of cell fate decision during development.

19F-perfluorocarbon-labeled human peripheral blood mononuclear cells can be detected in vivo using clinical MRI parameters in a therapeutic cell setting.

PubMed

Fink, Corby; Gaudet, Jeffrey M; Fox, Matthew S; Bhatt, Shashank; Viswanathan, Sowmya; Smith, Michael; Chin, Joseph; Foster, Paula J; Dekaban, Gregory A

2018-01-12

A 19 Fluorine ( 19 F) perfluorocarbon cell labeling agent, when employed with an appropriate cellular MRI protocol, allows for in vivo cell tracking. 19 F cellular MRI can be used to non-invasively assess the location and persistence of cell-based cancer vaccines and other cell-based therapies. This study was designed to determine the feasibility of labeling and tracking peripheral blood mononuclear cells (PBMC), a heterogeneous cell population. Under GMP-compliant conditions human PBMC were labeled with a 19 F-based MRI cell-labeling agent in a manner safe for autologous re-injection. Greater than 99% of PBMC labeled with the 19 F cell-labeling agent without affecting functionality or affecting viability. The 19 F-labeled PBMC were detected in vivo in a mouse model at the injection site and in a draining lymph node. A clinical cellular MR protocol was optimized for the detection of PBMC injected both at the surface of a porcine shank and at a depth of 1.2 cm, equivalent to depth of a human lymph node, using a dual 1 H/ 19 F dual switchable surface radio frequency coil. This study demonstrates it is feasible to label and track 19 F-labeled PBMC using clinical MRI protocols. Thus, 19 F cellular MRI represents a non-invasive imaging technique suitable to assess the effectiveness of cell-based cancer vaccines.

The novel cyclophilin D inhibitor compound 19 protects retinal pigment epithelium cells and retinal ganglion cells from UV radiation.

PubMed

Xie, Laiqing; Cheng, Long; Xu, Guoxu; Zhang, Ji; Ji, Xiaoyan; Song, E

2017-06-10

Excessive Ultra violet (UV) radiation induces injuries to retinal pigment epithelium (RPE) cells (RPEs) and retinal ganglion cells (RGCs), causing retinal degeneration. Cyclophilin D (Cyp-D)-dependent mitochondrial permeability transition pore (mPTP) opening mediates UV-induced cell death. In this study, we show that a novel Cyp-D inhibitor compound 19 efficiently protected RPEs and RGCs from UV radiation. Compound 19-mediated cytoprotection requires Cyp-D, as it failed to further protect RPEs/RGCs from UV when Cyp-D was silenced by targeted shRNAs. Compound 19 almost blocked UV-induced p53-Cyp-D mitochondrial association, mPTP opening and subsequent cytochrome C release. Further studies showed that compound 19 inhibited UV-induced reactive oxygen species (ROS) production, lipid peroxidation and DNA damage. Together, compound 19 protects RPEs and RGCs from UV radiation, possibly via silencing Cyp-D-regulated intrinsic mitochondrial death pathway. Compound 19 could a lead compound for treating UV-associated retinal degeneration diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Simplified process for the production of anti-CD19-CAR engineered T cells

PubMed Central

Tumaini, Barbara; Lee, Daniel W.; Lin, Tasha; Castiello, Luciano; Stroncek, David F.; Mackall, Crystal; Wayne, Alan; Sabatino, Marianna

2014-01-01

Background Adoptive Immunotherapy using chimeric antigen receptor (CAR) engineered T cells specific for CD19 has shown promising results for the treatment of B cell lymphomas and leukemia. This therapy involves the transduction of autologous T cells with a viral vector and the subsequent cell expansion. Here, we describe a new, simplified method to produce anti-CD19-CAR T cells. Methods T cells were isolated from peripheral blood mononuclear cell (PBMC) with anti-CD3/anti-CD28 paramagnetic beads. After 2 days, the T cells were added to culture bags pre-treated with RetroNectin and loaded with the retroviral anti-CD19 CAR vector. The cells, beads and vector were incubated for 24 hours and then a second transduction was performed. No spinoculation was used. Cells were then expanded for an additional 9 days. Results The method was validated using 2 PBMC products from a patient with B-CLL and one PBMC product from a healthy subject. The 2 PBMC products from the B-CLL patient contained 11.4% and 12.9% T cells. The manufacture process led to final products highly enriched in T cells with a mean CD3+ cell content of 98%, a mean expansion of 10.6 fold and a mean transduction efficiency of 68%. Similar results were obtained from the PBMCs of the first 4 ALL patients treated at our institution. Discussion We developed a simplified semi-closed system for the initial selection, activation, transduction and expansion of T cells using anti-CD3/anti-CD28 beads and bags, to produce autologous anti-CD19 CAR transduced T cells to support an ongoing clinical trial. PMID:23992830

MicroRNA-19a and CD22 Comprise a Feedback Loop for B Cell Response in Sepsis.

PubMed

Jiang, Yinan; Zhou, Hongmin; Ma, Dandan; Chen, Zhonghua Klaus; Cai, Xun

2015-05-28

MicroRNA-19a (miR-19a), an oncogenic microRNA, has been recently reported to target CD22 in B cell lymphoma cell lines, but its role in inflammatory response is unclear. CD22 is a negative regulator for BCR signaling, and we hypothesize that miR-19a regulates B cell response by targeting CD22 in sepsis. In order to determine whether miR-19a-CD22 pathway was involved in sepsis, and what role it played in the regulatory mechanisms, we detected the levels of miR-19a in B cells obtained from patients with sepsis, and measured the levels of miR-19a and CD22 expression in B cells activated by LPS in vitro. Additionally, we investigated the correlation between miR-19a and CD22, as well as the influence of this pathway on BCR signaling, in transfected B cells. We found that septic patients displayed up-regulated miR-19a in B cells. In vitro, miR-19a was increased in activated B cells, with CD22 expression initially enhanced but subsequently decreased. Moreover, overexpression of miR-19a resulted in an amplified BCR signaling, while overexpression of CD22 attenuated the effect of miR-19a and increased its expression. Our study demonstrated that miR-19a and CD22 comprised a feedback loop for B cell response in sepsis, providing a potential therapeutic target to recover the immune homeostasis.

MicroRNA-19a and CD22 Comprise a Feedback Loop for B Cell Response in Sepsis

PubMed Central

Jiang, Yinan; Zhou, Hongmin; Ma, Dandan; Chen, Zhonghua Klaus; Cai, Xun

2015-01-01

Background MicroRNA-19a (miR-19a), an oncogenic microRNA, has been recently reported to target CD22 in B cell lymphoma cell lines, but its role in inflammatory response is unclear. CD22 is a negative regulator for BCR signaling, and we hypothesize that miR-19a regulates B cell response by targeting CD22 in sepsis. Material/Methods In order to determine whether miR-19a-CD22 pathway was involved in sepsis, and what role it played in the regulatory mechanisms, we detected the levels of miR-19a in B cells obtained from patients with sepsis, and measured the levels of miR-19a and CD22 expression in B cells activated by LPS in vitro. Additionally, we investigated the correlation between miR-19a and CD22, as well as the influence of this pathway on BCR signaling, in transfected B cells. Results We found that septic patients displayed up-regulated miR-19a in B cells. In vitro, miR-19a was increased in activated B cells, with CD22 expression initially enhanced but subsequently decreased. Moreover, overexpression of miR-19a resulted in an amplified BCR signaling, while overexpression of CD22 attenuated the effect of miR-19a and increased its expression. Conclusions Our study demonstrated that miR-19a and CD22 comprised a feedback loop for B cell response in sepsis, providing a potential therapeutic target to recover the immune homeostasis. PMID:26017478

Infusion of donor-derived CD19-redirected virus-specific T cells for B-cell malignancies relapsed after allogeneic stem cell transplant: a phase 1 study

PubMed Central

Cruz, Conrad Russell Y.; Micklethwaite, Kenneth P.; Savoldo, Barbara; Ramos, Carlos A.; Lam, Sharon; Ku, Stephanie; Diouf, Oumar; Liu, Enli; Barrett, A. John; Ito, Sawa; Shpall, Elizabeth J.; Krance, Robert A.; Kamble, Rammurti T.; Carrum, George; Hosing, Chitra M.; Gee, Adrian P.; Mei, Zhuyong; Grilley, Bambi J.; Heslop, Helen E.; Rooney, Cliona M.; Brenner, Malcolm K.; Bollard, Catherine M.

2013-01-01

Autologous T cells expressing a CD19-specific chimeric antigen receptor (CD19.CAR) are active against B-cell malignancies, but it is unknown whether allogeneic CD19.CAR T cells are safe or effective. After allogeneic hematopoietic stem cell transplantation (HSCT), infused donor-derived virus-specific T cells (VSTs) expand in vivo, persist long term, and display antiviral activity without inducing graft-vs-host disease; therefore, we determined whether donor VSTs, engineered to express CD19.CAR, retained the characteristics of nonmanipulated allogeneic VSTs while gaining antitumor activity. We treated 8 patients with allogeneic (donor-derived) CD19.CAR-VSTs 3 months to 13 years after HSCT. There were no infusion-related toxicities. VSTs persisted for a median of 8 weeks in blood and up to 9 weeks at disease sites. Objective antitumor activity was evident in 2 of 6 patients with relapsed disease during the period of CD19.CAR-VST persistence, whereas 2 patients who received cells while in remission remain disease free. In 2 of 3 patients with viral reactivation, donor CD19.CAR-VSTs expanded concomitantly with VSTs. Hence CD19.CAR-VSTs display antitumor activity and, because their number may be increased in the presence of viral stimuli, earlier treatment post-HSCT (when lymphodepletion is greater and the incidence of viral infection is higher) or planned vaccination with viral antigens may enhance disease control. This study is registered at clinicaltrials.gov as #NCT00840853. PMID:24030379

miR-19a correlates with poor prognosis of clear cell renal cell carcinoma patients via promoting cell proliferation and suppressing PTEN/SMAD4 expression.

PubMed

Ma, Qiang; Peng, Zhiqiang; Wang, Lei; Li, Yanming; Wang, Kaizhen; Zheng, Junfang; Liang, Zhiyong; Liu, Tonghua

2016-12-01

MicroRNAs (miRNAs) were reported to be involved in the development of clear cell renal cell carcinoma (ccRCC). However, the study on miRNAs in ccRCC is far from complete. The present study identified miRNAs which could act as potential novel prognostic markers for ccRCC, and analyzed its possible mechanism. We found that miR-19a correlated with poor prognosis of ccRCC patients via promoting cell proliferation and suppressing PTEN/SMAD4 expression. Both the microarray screening result and TCGA KIRC dataset analysis showed that miR-19a was significantly upregulated in ccRCC tissues, and further analysis of TCGA data revealed that the upregulated level of miR-19a was strongly associated with advanced T stage and poor prognosis of ccRCC patients. Consistent with clinical observations, miR-19a overexpression significantly promoted ccRCC cell proliferation in vitro. To further explore the mechanism by which miR-19a correlated with cell proliferation and poor prognosis of ccRCC, we performed gene set enrichment analysis (GSEA) for target genes of miR-19a in ccRCC patients. Result indicated that the key target genes of miR-19a included SMAD4 and PTEN. In ccRCC tissues, expression levels of SMAD4 and PTEN were negatively correlated with expression level of miR-19a, revealing that miR-19a suppressed the expression of SMAD4 and PTEN in ccRCC patients. miR-19a overexpression significantly suppressed the expression of SMAD4 and PTEN in vitro, further verifying that SMAD4 and PTEN were the target genes of miR-19a in ccRCC cells. Our results elucidated the tumor promoting role of miR-19a and established miR-19a as a potential novel prognostic marker for ccRCC.

Potent neutralizing anti-CD1d antibody reduces lung cytokine release in primate asthma model.

PubMed

Nambiar, Jonathan; Clarke, Adam W; Shim, Doris; Mabon, David; Tian, Chen; Windloch, Karolina; Buhmann, Chris; Corazon, Beau; Lindgren, Matilda; Pollard, Matthew; Domagala, Teresa; Poulton, Lynn; Doyle, Anthony G

2015-01-01

CD1d is a receptor on antigen-presenting cells involved in triggering cell populations, particularly natural killer T (NKT) cells, to release high levels of cytokines. NKT cells are implicated in asthma pathology and blockade of the CD1d/NKT cell pathway may have therapeutic potential. We developed a potent anti-human CD1d antibody (NIB.2) that possesses high affinity for human and cynomolgus macaque CD1d (KD ∼100 pM) and strong neutralizing activity in human primary cell-based assays (IC50 typically <100 pM). By epitope mapping experiments, we showed that NIB.2 binds to CD1d in close proximity to the interface of CD1d and the Type 1 NKT cell receptor β-chain. Together with data showing that NIB.2 inhibited stimulation via CD1d loaded with different glycolipids, this supports a mechanism whereby NIB.2 inhibits NKT cell activation by inhibiting Type 1 NKT cell receptor β-chain interactions with CD1d, independent of the lipid antigen in the CD1d antigen-binding cleft. The strong in vitro potency of NIB.2 was reflected in vivo in an Ascaris suum cynomolgus macaque asthma model. Compared with vehicle control, NIB.2 treatment significantly reduced bronchoalveolar lavage (BAL) levels of Ascaris-induced cytokines IL-5, IL-8 and IL-1 receptor antagonist, and significantly reduced baseline levels of GM-CSF, IL-6, IL-15, IL-12/23p40, MIP-1α, MIP-1β, and VEGF. At a cellular population level NIB.2 also reduced numbers of BAL lymphocytes and macrophages, and blood eosinophils and basophils. We demonstrate that anti-CD1d antibody blockade of the CD1d/NKT pathway modulates inflammatory parameters in vivo in a primate inflammation model, with therapeutic potential for diseases where the local cytokine milieu is critical.

Selective Resistance of CD44hi T Cells to p53 Dependent Cell Death Results in Persistence of Immunologic Memory after Total Body Irradiation1,2,3

PubMed Central

Yao, Zhenyu; Jones, Jennifer; Kohrt, Holbrook; Strober, Samuel

2011-01-01

Our previous studies showed that treatment of mice with total body irradiation (TBI) or total lymphoid tissue irradiation (TLI) markedly changes the balance of residual T cell subsets to favor CD4+CD44hi natural killer T (NKT) cells due to differential resistance of the latter subset to cell death. The object of the current study was to further elucidate the changed balance and mechanisms of differential radioresistance of T cell subsets after graded doses of TBI. The experimental results show that CD4+ T cells were markedly more resistant than CD8+ T cells, and CD44hi T cells including NKT cells and memory T cells were markedly more resistant than CD44lo (naïve) T cells. The memory T cells immunized to alloantigens persisted even after myeloabloative (1,000cGy) TBI, and were able to prevent engraftment of bone marrow transplants. Although T cell death after 1,000cGy was prevented in p53−/− mice, there was progressive T cell death in p53−/− mice at higher doses. Whereas, p53 dependent T cell death changed the balance of subsets, the p53 independent T cell death did not. In conclusion, resistance of CD44hi T cells to p53 dependent cell death results in the persistence of immunological memory after TBI, and can explain the immune mediated rejection of marrow transplants in sensitized recipients. PMID:21930972

Restricting nonclassical MHC genes coevolve with TRAV genes used by innate-like T cells in mammals

PubMed Central

Boudinot, Pierre; Mondot, Stanislas; Jouneau, Luc; Teyton, Luc; Lefranc, Marie-Paule; Lantz, Olivier

2016-01-01

Whereas major histocompatibility class-1 (MH1) proteins present peptides to T cells displaying a large T-cell receptor (TR) repertoire, MH1Like proteins, such as CD1D and MR1, present glycolipids and microbial riboflavin precursor derivatives, respectively, to T cells expressing invariant TR-α (iTRA) chains. The groove of such MH1Like, as well as iTRA chains used by mucosal-associated invariant T (MAIT) and natural killer T (NKT) cells, respectively, may result from a coevolution under particular selection pressures. Herein, we investigated the evolutionary patterns of the iTRA of MAIT and NKT cells and restricting MH1Like proteins: MR1 appeared 170 Mya and is highly conserved across mammals, evolving more slowly than other MH1Like. It has been pseudogenized or independently lost three times in carnivores, the armadillo, and lagomorphs. The corresponding TRAV1 gene also evolved slowly and harbors highly conserved complementarity determining regions 1 and 2. TRAV1 is absent exclusively from species in which MR1 is lacking, suggesting that its loss released the purifying selection on MR1. In the rabbit, which has very few NKT and no MAIT cells, a previously unrecognized iTRA was identified by sequencing leukocyte RNA. This iTRA uses TRAV41, which is highly conserved across several groups of mammals. A rabbit MH1Like gene was found that appeared with mammals and is highly conserved. It was independently lost in a few groups in which MR1 is present, like primates and Muridae, illustrating compensatory emergences of new MH1Like/Invariant T-cell combinations during evolution. Deciphering their role is warranted to search similar effector functions in humans. PMID:27170188

Cell cycle inhibitor, p19INK4d, promotes cell survival and decreases chromosomal aberrations after genotoxic insult due to enhanced DNA repair.

PubMed

Scassa, María E; Marazita, Mariela C; Ceruti, Julieta M; Carcagno, Abel L; Sirkin, Pablo F; González-Cid, Marcela; Pignataro, Omar P; Cánepa, Eduardo T

2007-05-01

Genome integrity and cell proliferation and survival are regulated by an intricate network of pathways that includes cell cycle checkpoints, DNA repair and recombination, and programmed cell death. It makes sense that there should be a coordinated regulation of these different processes, but the components of such mechanisms remain unknown. In this report, we demonstrate that p19INK4d expression enhances cell survival under genotoxic conditions. By using p19INK4d-overexpressing clones, we demonstrated that p19INK4d expression correlates with the cellular resistance to UV treatment with increased DNA repair activity against UV-induced lesions. On the contrary, cells transfected with p19INK4d antisense cDNA show reduced ability to repair DNA damage and increased sensitivity to genotoxic insult when compared with their p19INK4d-overexpressing counterparts. Consistent with these findings, our studies also show that p19INK4d-overexpressing cells present not only a minor accumulation of UV-induced chromosomal aberrations but a lower frequency of spontaneous chromosome abnormalities than p19INK4d-antisense cells. Lastly, we suggest that p19INK4d effects are dissociated from its role as CDK4/6 inhibitor. The results presented herein support a crucial role for p19INK4d in regulating genomic stability and overall cell viability under conditions of genotoxic stress. We propose that p19INK4d would belong to a protein network that would integrate DNA repair, apoptotic and checkpoint mechanisms in order to maintain the genomic integrity.

Thio-isoglobotrihexosylceramide, an agonist for activating invariant natural killer T cells.

PubMed

Xia, Chengfeng; Zhou, Dapeng; Liu, Chengwen; Lou, Yanyan; Yao, Qingjia; Zhang, Wenpeng; Wang, Peng George

2006-11-23

Thio-isoglobotrihexosylceramide (S-iGb3) might be resistant to alpha-galactosidases in antigen-presenting cells and have a longer retaining time in the lysosome before being loaded to CD1d. The biological assay showed that S-iGb3 demonstrates a much higher increase as a stimulatory ligand toward invariant natural killer T (iNKT) cells as compared to iGb3. [structure: see text].

A unique 19F MRI agent for the tracking of non phagocytic cells in vivo.

PubMed

Moonshi, Shehzahdi S; Zhang, Cheng; Peng, Hui; Puttick, Simon; Rose, Stephen; Fisk, Nicholas M; Bhakoo, Kishore; Stringer, Brett W; Qiao, Greg G; Gurr, Paul A; Whittaker, Andrew K

2018-05-03

There is currently intense interest in new methods for understanding the fate of therapeutically-relevant cells, such as mesenchymal stem cells (MSCs). The absence of a confounding background signal and consequent unequivocal assignment makes 19F MRI one of the most attractive modalities for the tracking of injected cells in vivo. We describe here the synthesis of novel partly-fluorinated polymeric nanoparticles with small size and high fluorine content as MRI agents. The polymers, constructed from perfluoropolyether methacrylate (PFPEMA) and oligo(ethylene glycol) methacrylate (OEGMA) have favourable cell uptake profiles and excellent MRI performance. To facilitate cell studies the polymer was further conjugated with a fluorescent dye creating a dual-modal imaging agent. The efficacy of labelling of MSCs was assessed using 19F NMR, flow cytometry and confocal microscopy. The labelling efficiency of 2.6 ± 0.1 × 1012 19F atoms per cell, and viability of >90% demonstrates high uptake and good tolerance by the cells. This loading translates to a minimum 19F MRI detection sensitivity of ∼7.4 × 103 cells per voxel. Importantly, preliminary in vivo data demonstrate that labelled cells can be readily detected within a short acquisition scan period (12 minutes). Hence, these copolymers show outstanding potential for 19F MRI cellular tracking and for quantification of non-phagocytic and therapeutically-relevant cells in vivo.

Ribosomal protein L19 overexpression activates the unfolded protein response and sensitizes MCF7 breast cancer cells to endoplasmic reticulum stress-induced cell death.

PubMed

Hong, Mina; Kim, HyungRyong; Kim, Inki

2014-07-18

Although first identified for their roles in protein synthesis, certain ribosomal proteins exert pleiotropic physiological functions in the cell. Ribosomal protein L19 is overexpressed in breast cancer cells by amplification and copy number variation. In this study, we examined the novel pro-apoptotic role of ribosomal protein L19 in the breast cancer cell line MCF7. Overexpression of RPL19 sensitized MCF7 cells to endoplasmic reticulum stress-induced cell death. RPL19 overexpression itself was not cytotoxic; however, cell death induction was enhanced when RPL19 overexpressing cells were incubated with endoplasmic reticulum stress-inducing agents, and this sensitizing effect was specific to MCF7 cells. Examination of the cell signaling pathways that mediate the unfolded protein response (UPR) revealed that overexpression of RPL19 induced pre-activation of the UPR, including phosphorylation of pERK-like ER kinase (PERK), phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α), and activation of p38 MAPK-associated stress signaling. Our findings suggest that upregulation of RPL19 induces ER stress, resulting in increased sensitivity to ER stress and enhanced cell death in MCF7 breast cancer cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Chemotherapy-Refractory Diffuse Large B-Cell Lymphoma and Indolent B-Cell Malignancies Can Be Effectively Treated With Autologous T Cells Expressing an Anti-CD19 Chimeric Antigen Receptor

PubMed Central

Kochenderfer, James N.; Dudley, Mark E.; Kassim, Sadik H.; Somerville, Robert P.T.; Carpenter, Robert O.; Stetler-Stevenson, Maryalice; Yang, James C.; Phan, Giao Q.; Hughes, Marybeth S.; Sherry, Richard M.; Raffeld, Mark; Feldman, Steven; Lu, Lily; Li, Yong F.; Ngo, Lien T.; Goy, Andre; Feldman, Tatyana; Spaner, David E.; Wang, Michael L.; Chen, Clara C.; Kranick, Sarah M.; Nath, Avindra; Nathan, Debbie-Ann N.; Morton, Kathleen E.; Toomey, Mary Ann; Rosenberg, Steven A.

2015-01-01

Purpose T cells can be genetically modified to express an anti-CD19 chimeric antigen receptor (CAR). We assessed the safety and efficacy of administering autologous anti-CD19 CAR T cells to patients with advanced CD19+ B-cell malignancies. Patients and Methods We treated 15 patients with advanced B-cell malignancies. Nine patients had diffuse large B-cell lymphoma (DLBCL), two had indolent lymphomas, and four had chronic lymphocytic leukemia. Patients received a conditioning chemotherapy regimen of cyclophosphamide and fludarabine followed by a single infusion of anti-CD19 CAR T cells. Results Of 15 patients, eight achieved complete remissions (CRs), four achieved partial remissions, one had stable lymphoma, and two were not evaluable for response. CRs were obtained by four of seven evaluable patients with chemotherapy-refractory DLBCL; three of these four CRs are ongoing, with durations ranging from 9 to 22 months. Acute toxicities including fever, hypotension, delirium, and other neurologic toxicities occurred in some patients after infusion of anti-CD19 CAR T cells; these toxicities resolved within 3 weeks after cell infusion. One patient died suddenly as a result of an unknown cause 16 days after cell infusion. CAR T cells were detected in the blood of patients at peak levels, ranging from nine to 777 CAR-positive T cells/μL. Conclusion This is the first report to our knowledge of successful treatment of DLBCL with anti-CD19 CAR T cells. These results demonstrate the feasibility and effectiveness of treating chemotherapy-refractory B-cell malignancies with anti-CD19 CAR T cells. The numerous remissions obtained provide strong support for further development of this approach. PMID:25154820

Recombinant Human Parvovirus B19 Vectors: Erythroid Cell-Specific Delivery and Expression of Transduced Genes

PubMed Central

Ponnazhagan, Selvarangan; Weigel, Kirsten A.; Raikwar, Sudhanshu P.; Mukherjee, Pinku; Yoder, Mervin C.; Srivastava, Arun

1998-01-01

A novel packaging strategy combining the salient features of two human parvoviruses, namely the pathogenic parvovirus B19 and the nonpathogenic adeno-associated virus type 2 (AAV), was developed to achieve erythroid cell-specific delivery as well as expression of the transduced gene. The development of such a chimeric vector system was accomplished by packaging heterologous DNA sequences cloned within the inverted terminal repeats of AAV and subsequently packaging the DNA inside the capsid structure of B19 virus. Recombinant B19 virus particles were assembled, as evidenced by electron microscopy as well as DNA slot blot analyses. The hybrid vector failed to transduce nonerythroid human cells, such as 293 cells, as expected. However, MB-02 cells, a human megakaryocytic leukemia cell line which can be infected by B19 virus following erythroid differentiation with erythropoietin (N. C. Munshi, S. Z. Zhou, M. J. Woody, D. A. Morgan, and A. Srivastava, J. Virol. 67:562–566, 1993) but lacks the putative receptor for AAV (S. Ponnazhagan, X.-S. Wang, M. J. Woody, F. Luo, L. Y. Kang, M. L. Nallari, N. C. Munshi, S. Z. Zhou, and A. Srivastava, J. Gen. Virol. 77:1111–1122, 1996), were readily transduced by this vector. The hybrid vector was also found to specifically target the erythroid population in primary human bone marrow cells as well as more immature hematopoietic progenitor cells following erythroid differentiation, as evidenced by selective expression of the transduced gene in these target cells. Preincubation with anticapsid antibodies against B19 virus, but not anticapsid antibodies against AAV, inhibited transduction of primary human erythroid cells. The efficiency of transduction of primary human erythroid cells by the recombinant B19 virus vector was significantly higher than that by the recombinant AAV vector. Further development of the AAV-B19 virus hybrid vector system should prove beneficial in gene therapy protocols aimed at the correction of inherited

Phase 1 studies of central memory-derived CD19 CAR T-cell therapy following autologous HSCT in patients with B-cell NHL.

PubMed

Wang, Xiuli; Popplewell, Leslie L; Wagner, Jamie R; Naranjo, Araceli; Blanchard, M Suzette; Mott, Michelle R; Norris, Adam P; Wong, ChingLam W; Urak, Ryan Z; Chang, Wen-Chung; Khaled, Samer K; Siddiqi, Tanya; Budde, Lihua E; Xu, Jingying; Chang, Brenda; Gidwaney, Nikita; Thomas, Sandra H; Cooper, Laurence J N; Riddell, Stanley R; Brown, Christine E; Jensen, Michael C; Forman, Stephen J

2016-06-16

Myeloablative autologous hematopoietic stem cell transplantation (HSCT) is a mainstay of therapy for relapsed intermediate-grade B-cell non-Hodgkin lymphoma (NHL); however, relapse rates are high. In phase 1 studies designed to improve long-term remission rates, we administered adoptive T-cell immunotherapy after HSCT, using ex vivo-expanded autologous central memory-enriched T cells (TCM) transduced with lentivirus expressing CD19-specific chimeric antigen receptors (CARs). We present results from 2 safety/feasibility studies, NHL1 and NHL2, investigating different T-cell populations and CAR constructs. Engineered TCM-derived CD19 CAR T cells were infused 2 days after HSCT at doses of 25 to 200 × 10(6) in a single infusion. In NHL1, 8 patients safely received T-cell products engineered from enriched CD8(+) TCM subsets, expressing a first-generation CD19 CAR containing only the CD3ζ endodomain (CD19R:ζ). Four of 8 patients (50%; 95% confidence interval [CI]: 16-84%) were progression free at both 1 and 2 years. In NHL2, 8 patients safely received T-cell products engineered from enriched CD4(+) and CD8(+) TCM subsets and expressing a second-generation CD19 CAR containing the CD28 and CD3ζ endodomains (CD19R:28ζ). Six of 8 patients (75%; 95% CI: 35-97%) were progression free at 1 year. The CD4(+)/CD8(+) TCM-derived CD19 CAR T cells (NHL2) exhibited improvement in expansion; however, persistence was ≤28 days, similar to that seen by others using CD28 CARs. Neither cytokine release syndrome nor delayed hematopoietic engraftment was observed in either trial. These data demonstrate the safety and feasibility of CD19 CAR TCM therapy after HSCT. Trials were registered at www.clinicaltrials.gov as #NCT01318317 and #NCT01815749. © 2016 by The American Society of Hematology.

Orbital involvement in extranodal natural killer T cell lymphoma: an atypical case presentation and review of the literature.

PubMed

Ely, A; Evans, J; Sundstrom, J M; Malysz, J; Specht, C S; Wilkinson, M

2012-08-01

To report a rare case of extranodal NK/T cell lymphoma (NKTL) and to compare its features with those cases previously reported. Case report, observational and literature review. Complete ophthalmologic examinations followed by excisional biopsy, histopathologic examination and therapy with radiation and chemotherapy. Evaluation of clinical presenting features and histopathologic diagnosis along with patient outcome. A 22 year old female presented as a referral with right orbital swelling, decreased vision and eye pain for 5 weeks. Subsequent orbital CT and multiple biopsies resulted in a diagnosis of extranodal natural killer (NK)/T cell lymphoma (NKTL). Despite continued chemotherapy and orbital radiation the patient expired within 3 months of diagnosis. To our knowledge, only 8 cases of orbital involvement without nasal mucosal involvement are reported in the literature, the majority in patients of male gender around the fifth decade. Here we present an atypical and aggressive case of extranodal NK/T cell lymphoma presenting in a 22 year old Caucasian female as orbital swelling without evidence of nasal mucosal involvement. It is important to distinguish NKTL from the more common benign lymphoproliferative lesions of the orbital adnexa as prognosis of these two clinical entities varies and timely diagnosis is key. The present case demonstrates that extranodal NKTL can occur in the orbit without evidence of the more common nasal mucosal presentations and should be included in the differential diagnosis of ocular adnexal lesions suspicious for a lymphoproliferative disorder and/or an inflammatory process.

Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells

PubMed Central

Kebriaei, Partow; Singh, Harjeet; Huls, M. Helen; Figliola, Matthew J.; Bassett, Roland; Olivares, Simon; Jena, Bipulendu; Dawson, Margaret J.; Kumaresan, Pappanaicken R.; Su, Shihuang; Maiti, Sourindra; Dai, Jianliang; Moriarity, Branden; Forget, Marie-Andrée; Senyukov, Vladimir; Orozco, Aaron; Liu, Tingting; McCarty, Jessica; Jackson, Rineka N.; Moyes, Judy S.; Rondon, Gabriela; Qazilbash, Muzaffar; Ciurea, Stefan; Alousi, Amin; Nieto, Yago; Rezvani, Katy; Marin, David; Popat, Uday; Hosing, Chitra; Shpall, Elizabeth J.; Kantarjian, Hagop; Keating, Michael; Wierda, William; Do, Kim Anh; Largaespada, David A.; Lee, Dean A.; Hackett, Perry B.; Champlin, Richard E.; Cooper, Laurence J.N.

2016-01-01

BACKGROUND. T cells expressing antigen-specific chimeric antigen receptors (CARs) improve outcomes for CD19-expressing B cell malignancies. We evaluated a human application of T cells that were genetically modified using the Sleeping Beauty (SB) transposon/transposase system to express a CD19-specific CAR. METHODS. T cells were genetically modified using DNA plasmids from the SB platform to stably express a second-generation CD19-specific CAR and selectively propagated ex vivo with activating and propagating cells (AaPCs) and cytokines. Twenty-six patients with advanced non-Hodgkin lymphoma and acute lymphoblastic leukemia safely underwent hematopoietic stem cell transplantation (HSCT) and infusion of CAR T cells as adjuvant therapy in the autologous (n = 7) or allogeneic settings (n = 19). RESULTS. SB-mediated genetic transposition and stimulation resulted in 2,200- to 2,500-fold ex vivo expansion of genetically modified T cells, with 84% CAR expression, and without integration hotspots. Following autologous HSCT, the 30-month progression-free and overall survivals were 83% and 100%, respectively. After allogeneic HSCT, the respective 12-month rates were 53% and 63%. No acute or late toxicities and no exacerbation of graft-versus-host disease were observed. Despite a low antigen burden and unsupportive recipient cytokine environment, CAR T cells persisted for an average of 201 days for autologous recipients and 51 days for allogeneic recipients. CONCLUSIONS. CD19-specific CAR T cells generated with SB and AaPC platforms were safe, and may provide additional cancer control as planned infusions after HSCT. These results support further clinical development of this nonviral gene therapy approach. TRIAL REGISTRATION. Autologous, NCT00968760; allogeneic, NCT01497184; long-term follow-up, NCT01492036. FUNDING. National Cancer Institute, private foundations, and institutional funds. Please see Acknowledgments for details. PMID:27482888

Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells.

PubMed

Kebriaei, Partow; Singh, Harjeet; Huls, M Helen; Figliola, Matthew J; Bassett, Roland; Olivares, Simon; Jena, Bipulendu; Dawson, Margaret J; Kumaresan, Pappanaicken R; Su, Shihuang; Maiti, Sourindra; Dai, Jianliang; Moriarity, Branden; Forget, Marie-Andrée; Senyukov, Vladimir; Orozco, Aaron; Liu, Tingting; McCarty, Jessica; Jackson, Rineka N; Moyes, Judy S; Rondon, Gabriela; Qazilbash, Muzaffar; Ciurea, Stefan; Alousi, Amin; Nieto, Yago; Rezvani, Katy; Marin, David; Popat, Uday; Hosing, Chitra; Shpall, Elizabeth J; Kantarjian, Hagop; Keating, Michael; Wierda, William; Do, Kim Anh; Largaespada, David A; Lee, Dean A; Hackett, Perry B; Champlin, Richard E; Cooper, Laurence J N

2016-09-01

T cells expressing antigen-specific chimeric antigen receptors (CARs) improve outcomes for CD19-expressing B cell malignancies. We evaluated a human application of T cells that were genetically modified using the Sleeping Beauty (SB) transposon/transposase system to express a CD19-specific CAR. T cells were genetically modified using DNA plasmids from the SB platform to stably express a second-generation CD19-specific CAR and selectively propagated ex vivo with activating and propagating cells (AaPCs) and cytokines. Twenty-six patients with advanced non-Hodgkin lymphoma and acute lymphoblastic leukemia safely underwent hematopoietic stem cell transplantation (HSCT) and infusion of CAR T cells as adjuvant therapy in the autologous (n = 7) or allogeneic settings (n = 19). SB-mediated genetic transposition and stimulation resulted in 2,200- to 2,500-fold ex vivo expansion of genetically modified T cells, with 84% CAR expression, and without integration hotspots. Following autologous HSCT, the 30-month progression-free and overall survivals were 83% and 100%, respectively. After allogeneic HSCT, the respective 12-month rates were 53% and 63%. No acute or late toxicities and no exacerbation of graft-versus-host disease were observed. Despite a low antigen burden and unsupportive recipient cytokine environment, CAR T cells persisted for an average of 201 days for autologous recipients and 51 days for allogeneic recipients. CD19-specific CAR T cells generated with SB and AaPC platforms were safe, and may provide additional cancer control as planned infusions after HSCT. These results support further clinical development of this nonviral gene therapy approach. Autologous, NCT00968760; allogeneic, NCT01497184; long-term follow-up, NCT01492036. National Cancer Institute, private foundations, and institutional funds. Please see Acknowledgments for details.

New data for iimoriite.

USGS Publications Warehouse

Foord, E.E.; Staatz, M.H.; Conklin, N.M.

1984-01-01

In its first reported occurrence outside of Japan, iimoriite, Y2(SiO4)(CO3), has been found in a thorite- and uraninite-bearing quartz and albite vein from Bokan Mountain on Prince of Wales Island, Alaska. It occurs as buff-tan subhedral and anhedral grains, 0.01-0.5 mm in diameter, with vitreous lustre, white streak, and H. 51/2-6. Colourless in thin section, with no pleochroism, iimoriite is biaxially negative with alpha 1.753, beta 1.824, gamma 1.830, 2Valpha (obs.) 31o, 2Valpha (calc.) 31.5o. The mineral is triclinic, space group P1 or P1, a 6.573(1), b 6.651(1), c 6.454(1) A, alpha 116.44, beta 92.34, gamma 95.63o, Z = 2, Dobs. 4.47, Dcalc. 4.56 (from Gladstone-Dale relationship), Dcalc. 4.91 (from calculated formula weight and cell volume) g/cm3. The strongest XRD lines are 2.88(100), 2.95(80), 2.78(40), 3.02(36), 2.84(35) A. Chemical analyses are tabulated. -J.A.Z.

19. CONSTRUCTION PROGRESS PHOTO SHOWING (TYPICALLY COMPLEX) WASTE HOLDING CELL ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

19. CONSTRUCTION PROGRESS PHOTO SHOWING (TYPICALLY COMPLEX) WASTE HOLDING CELL PIPING. INEEL PHOTO NUMBER NRTS-59-3212. - Idaho National Engineering Laboratory, Old Waste Calcining Facility, Scoville, Butte County, ID

Induction of the Hajdu-Cheney Syndrome Mutation in CD19 B Cells in Mice Alters B-Cell Allocation but Not Skeletal Homeostasis.