A modified α-galactosyl ceramide for staining and stimulating natural killer T cells

Liu, Yang; Goff, Randal D.; Zhou, Dapeng; Mattner, Jochen; Sullivan, Barbara A.; Khurana, Archana; Cantu, Carlos; Ravkov, Eugene V.; Ibegbu, Chris; Altman, John D.; Teyton, Luc; Bendelac, Albert; Savage, Paul B.

doi:10.1016/j.jim.2006.02.009

Cited by 167 publications

(146 citation statements)

References 13 publications

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“…In the current study, we identified CD1d-restricted NKT cells by CD1d tetramers loaded with ␣-GalCer analog PBS57 (18). At 3 h following reperfusion, we observed an increase in the percentage of NKT cells producing IFN-␥.…”

Section: Discussionmentioning

confidence: 53%

“…To identify CD1d-restricted NKT cells, we labeled kidney cells with anti-mouse CD45-FITC, anti-mouse TCR␤-PE, and CD1d tetramer-Alexa 647 loaded with PBS57 (1:500), an analog of ␣-galactosyl ceramide (␣GalCer) that yields results identical with ␣GalCer (National Institutes of Health tetramer facility; www. yerkes.emory.edu/TETRAMER/CD1d_Tetramers.html) (18). Unloaded CD1d tetramer-Alexa 647 was used as control.…”

Section: Identification Of Nkt Cells and Quantitation Of Intracellulamentioning

confidence: 99%

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NKT Cell Activation Mediates Neutrophil IFN-γ Production and Renal Ischemia-Reperfusion Injury

Li¹,

Huang²,

Sung³

et al. 2007

The Journal of Immunology

298

355

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Previous work has shown that ischemia-reperfusion (IR) injury (IRI) is dependent on CD4+ T cells from naive mice acting within 24 h. We hypothesize that NKT cells are key participants in the early innate response in IRI. Kidneys from C57BL/6 mice were subjected to IRI (0.5, 1, 3, and 24 h of reperfusion). After 30 min of reperfusion, we observed a significant increase in CD4+ cells (145% of control) from single-cell kidney suspensions as measured by flow cytometry. A significant fraction of CD4+ T cells expressed the activation marker, CD69+, and adhesion molecule, LFA-1high. Three hours after reperfusion, kidney IFN-γ-producing cells were comprised largely of GR-1+CD11b+ neutrophils, but also contained CD1d-restricted NKT cells. Kidney IRI in mice administered Abs to block CD1d, or deplete NKT cells or in mice deficient of NKT cells (Jα18−/−), was markedly attenuated. These effects were associated with a significant decrease in renal infiltration and, in activation of NKT cells, and a decrease in IFN-γ-producing neutrophils. The results support the essential role of NKT cells and neutrophils in the innate immune response of renal IRI by mediating neutrophil infiltration and production of IFN-γ.

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Section: Discussionmentioning

confidence: 53%

Section: Identification Of Nkt Cells and Quantitation Of Intracellulamentioning

confidence: 99%

NKT Cell Activation Mediates Neutrophil IFN-γ Production and Renal Ischemia-Reperfusion Injury

Li¹,

Huang²,

Sung³

et al. 2007

The Journal of Immunology

298

355

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“…Spleen cell suspensions were incubated in round-bottom plates in RPMI1640 medium supplemented with glutamine, antibiotics, and 10% FCS (Sigma Aldrich, St. Louis, MO, USA) with 50 ng/mL plate-bound anti-CD3 antibody (eBioscience, San Diego, CA, USA), PMA/Ionomycin (Cell Stimulation Cocktail, eBioscience, San Diego, CA, USA), 0.1-100 ng/mL α-GalCer [85] or an equivalent amount of DMSO (Carl Roth, Karlsruhe, Germany). CD11b-and Nur77-GFP-expression on as well as IFN-γ-and T-betexpression in iNKT cells were analyzed using FACS.…”

Section: In Vitro Stimulation Of Inkt Cellsmentioning

confidence: 99%

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

et al. 2016

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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.Keywords: ADAP r Cytokine r iNKT cell r Integrin r slp-76Additional supporting information may be found in the online version of this article at the publisher's web-site Eur. J. Immunol. 2016Immunol. . 46: 2121Immunol. -2136 Introduction iNKT cells express a panoply of NK-cell receptors [1] and a canonical TCR through which they recognize (glyco-)lipid antigens [2]. iNKT cells activate similar signaling cascades after TCR ligation like other T lymphocytes [3], but utilize unique transcription factors for their development such as the promyelocytic leukemia zinc finger (PLZF) [4][5][6]. According to two different developmental models PLZF characterizes distinct maturation stages and polarized subsets. The sequential lineage model suggests a gradual decrease of PLZFexpression following selection of iNKT cells which show a Th2-dominated cytokine profile during earlier and a Th1-dominated cytokine profile during later stages of intrathymic maturation [1,7]. The second model describes lineage diversification and simultaneous differentiation into Th1-, Th2-, or Th17-polarized subsets that are defined by the level of PLZF-expression [8]. Although many iNKT cells release both Th1 and Th2 cytokines on a single cell level [9], the production of IL-17 and IFN-γ is mutually exclusive within NK1.1 − cells [10][11][12]. Several transcription factors, such as Egr2, T-bet, ThPOK, Id2, Id3, and the Tec kinases Itk and Rlk have been implicated in the differentiation of iNKT cell subsets [6,8,[13][14][15][16][17] which home to distinct tissues. Specifically, liver and spleen constitute the main source for the Th1-polarized sublineage which is PLZF low . Th2-or ...

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“…It can also be applied to humans, opening a new avenue for iNKT cell-based immunotherapy that has the potential to provide patients with therapeutic levels of iNKT cells throughout life. TCRs (16). We included TCR Vβ8 staining to focus on the dominant Vβ8 + population of mouse iNKT cells (1,2).…”

Section: Cloning Of Inkt Tcr Genes and Construction Of Retroviral Delmentioning

confidence: 99%

Genetic engineering of hematopoietic stem cells to generate invariant natural killer T cells

Smith

Liu

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Invariant natural killer T (iNKT) cells comprise a small population of αβ T lymphocytes. They bridge the innate and adaptive immune systems and mediate strong and rapid responses to many diseases, including cancer, infections, allergies, and autoimmunity. However, the study of iNKT cell biology and the therapeutic applications of these cells are greatly limited by their small numbers in vivo (∼0.01-1% in mouse and human blood). Here, we report a new method to generate large numbers of iNKT cells in mice through T-cell receptor (TCR) gene engineering of hematopoietic stem cells (HSCs). We showed that iNKT TCR-engineered HSCs could generate a clonal population of iNKT cells. These HSC-engineered iNKT cells displayed the typical iNKT cell phenotype and functionality. They followed a two-stage developmental path, first in thymus and then in the periphery, resembling that of endogenous iNKT cells. When tested in a mouse melanoma lung metastasis model, the HSC-engineered iNKT cells effectively protected mice from tumor metastasis. This method provides a powerful and high-throughput tool to investigate the in vivo development and functionality of clonal iNKT cells in mice. More importantly, this method takes advantage of the self-renewal and longevity of HSCs to generate a long-term supply of engineered iNKT cells, thus opening up a new avenue for iNKT cell-based immunotherapy. αβ T lymphocytes highly conserved from mice to humans. Like conventional αβ T cells, iNKT cells are derived from hematopoietic stem cells (HSCs) and develop in the thymus. However, they differ from conventional T cells in several important aspects, including their display of NK cell markers, their recognition of glycolipid antigens presented by the nonclassical monomorphic major histocompatibility complex (MHC) molecule CD1d, and their expression of semi-invariant T-cell receptors (identical α chains paired with a limited selection of β chains) (1, 2). Despite their small numbers in vivo (∼0.1-1% in mouse blood and ∼0.01-1% in human blood), iNKT cells have been suggested to play important roles in regulating many diseases, including cancer, infections, allergies, and autoimmunity (3). When stimulated, iNKT cells rapidly release a large amount of effector cytokines like IFN-γ and IL-4, both as a cell population and at the single-cell level. These cytokines then activate various immune effector cells, such as natural killer (NK) cells and dendritic cells (DCs) of the innate immune system, as well as CD4 helper and CD8 cytotoxic conventional αβ T cells of the adaptive immune system via activated DCs (3, 4). Because of their unique activation mechanism, iNKT cells can attack multiple diseases independent of antigen and MHC restrictions, making them attractive universal therapeutic agents (3, 4). Notably, because of the capacity of effector NK cells and conventional αβ T cells to specifically recognize diseased tissue cells, iNKT cell-induced immune reactions result in limited off-target side effects (3, 4).Restricted by their extremely low numbers, ...

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A modified α-galactosyl ceramide for staining and stimulating natural killer T cells

Cited by 167 publications

References 13 publications

NKT Cell Activation Mediates Neutrophil IFN-γ Production and Renal Ischemia-Reperfusion Injury

NKT Cell Activation Mediates Neutrophil IFN-γ Production and Renal Ischemia-Reperfusion Injury

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

Genetic engineering of hematopoietic stem cells to generate invariant natural killer T cells

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