The JI PBL Intereron Source
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kawachi, I.
Right arrow Articles by Gilfillan, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kawachi, I.
Right arrow Articles by Gilfillan, S.
Right arrowPubmed/NCBI databases
*Gene*GEO Profiles
*HomoloGene*Nucleotide
*Protein*UniGene
The Journal of Immunology, 2006, 176: 1618-1627.
Copyright © 2006 by The American Association of Immunologists

MR1-Restricted V{alpha}19i Mucosal-Associated Invariant T Cells Are Innate T Cells in the Gut Lamina Propria That Provide a Rapid and Diverse Cytokine Response1

Izumi Kawachi, Jorge Maldonado, Carey Strader and Susan Gilfillan2

Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Mucosal-associated invariant T (MAIT) cells reside primarily in the gut lamina propria and require commensal flora for selection/expansion. They are restricted by the highly conserved MHC class I-related molecule MR1 and, like most NK T cells, express an invariant TCR{alpha} chain. Although they probably contribute to gut immunity, MAIT cells have not been functionally characterized because they are so rare. To create a model in which they are more abundant, we generated transgenic mice expressing only the TCR{alpha} chain (V{alpha}19i) that defines MAIT cells. By directly comparing V{alpha}19i transgenic mice on MR1+/+ and MR1–/– backgrounds, we were able to distinguish and characterize a population of V{alpha}19i T cells dependent on MR1 for development. MR1-restricted V{alpha}19i transgenic T cells recapitulate what is known about MAIT cell development. Furthermore, a relatively high proportion of transgenic MAIT cells express NK1.1, and most have a cell surface phenotype similar to that of V{alpha}14i NK T cells. Finally, MR1-restricted V{alpha}19i T cells secrete IFN-{gamma}, IL-4, IL-5, and IL-10 following TCR ligation, and we provide evidence for what may be two functionally distinct MAIT cell populations. These data strongly support the idea that MAIT cells contribute to the innate immune response in the gut mucosa.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Mucosal-associated invariant T (MAIT)3 cells are a phylogenetically conserved population of T cells defined by the expression of an invariant TCR{alpha} chain that was originally identified during repertoire analysis of human CD4–CD8– double-negative (DN) peripheral blood {alpha}beta T cells (1). Lantz and colleagues (2) demonstrated that this TCRV{alpha}7.2-J{alpha}33 chain (V{alpha}7.2i) is expressed on a subset of human T cells and a homologous TCRV{alpha}19-J{alpha}33 (V{alpha}19i) chain on a similar T cell population in mice and cattle. Using quantitative PCR and limiting dilution assays, they determined that ~1 of 7.5 DN and 1 of 50 CD8{alpha}{alpha}+ T cells from human PBLs express V{alpha}7.2i, whereas 1 of 56 of DN T cells from the mesenteric lymph nodes (mLN) express the V{alpha}19i chain in mice; in cattle, the majority of V{alpha}19i T cells appear to be DN. Approximately 1 of 6000 of these cells is estimated to be CD4+ in humans, whereas a higher proportion appears to express CD4 in mice. Both human and murine MAIT cell TCR{alpha} chains preferentially associate with a limited number of TCRbeta chains. V{alpha}19i T cells are not present in nude mice, indicating that they develop in the thymus. In addition, they require B cells for selection/expansion, and the few IgA+ cells present in µMT–/– mice are sufficient for this (3). Most intriguingly, V{alpha}19i T cells are most frequent in the intestinal lamina propria (LP) and are not present in germfree mice (3), which suggests that they require commensal flora to expand and/or persist. When possible, these observations have been confirmed with human V{alpha}7.2i T cells, indicating that MAIT cells are a conserved lineage of T cells that function in the gut mucosa.

How MAIT cells recognize and respond to commensal flora is not known, but their restriction element has been identified: V{alpha}19i T cells require the MHC class I-related molecule MR1 for selection/expansion (3). The most striking feature of MR1 is the remarkable conservation of its {alpha}1 and {alpha}2 domains among mammalian species; human and mouse MR1 share 90% ({alpha}1)/89% ({alpha}2) sequence identity, whereas other functionally equivalent nonclassical and classical MHC molecules share 51–78% ({alpha}1)/41–76% ({alpha}2) (4, 5). This implies a highly conserved function for MR1, perhaps binding an invariant ligand. Supporting this, cumulative evidence from experiments with transfectants indicates that MR1 protein does not reach the cell surface in the absence of ligand (6, 7). In addition, studies with MAIT cell hybridomas and a panel of MR1 mutants strongly suggest that V{alpha}19i T cells recognize MR1 in a ligand-dependent manner (8). MAIT cells are present in TAP–/– mice, indicating that the putative ligand is not a conventional peptide. Certainly, products derived from or induced by commensal flora are highly plausible candidates for MR1 ligands, although ubiquitously expressed endogenous ligands appear to activate MAIT cell hybridomas (8).

MAIT cells most closely resemble another phylogenetically conserved population of T cells that also express an invariant TCR{alpha} chain in association with a limited number of TCRbeta chains. A unique TCRV{alpha}24-J{alpha}18 chain and its homologous TCRV{alpha}14-J{alpha}18 murine counterpart are expressed on functionally analogous human and murine T cell subsets generally referred to as NK T cells because most coexpress NK cell receptors, including NK1.1 or NKR-P1A (CD161) (9, 10, 11). V{alpha}24/14i NK T cells recognize the MHC class Ib molecule CD1d in association with hydrophobic ligands, including the lysosomal glycosphingolipid, isoglobotrihexosylceramide, as well as glycosphingolipids from the cell walls of Gram-negative bacteria that do not contain LPS (12, 13, 14). Upon TCR ligation, V{alpha}24i/14i NK T cells rapidly secrete a number of cytokines, including large amounts of IL-4 and IFN-{gamma}, and have increased cytolytic activity. They have been implicated in antibacterial, antiviral, and antiprotozoan responses as well as tumor rejection. In addition, V{alpha}14i NK T cells provide protection against several autoimmune diseases, including diabetes and experimental autoimmune encephalomyelitis, and can induce tolerance (9, 10, 11). However, they also mediate uncontrolled Th2 responses with pathological consequences, including asthma and ulcerative colitis (15, 16, 17). In summary, V{alpha}24i/14i NK T cells are a distinct lineage of innate T cells that respond rapidly to TCR stimulation and thereby influence the adaptive response in a variety of ways (18).

Given their similarities, it is tempting to speculate that MAIT cells and V{alpha}24i/14i NK T cells play similar roles but in different environments: V{alpha}24/14i NK T cells in internal organs, including the thymus, bone marrow (BM), liver, and spleen, and MAIT cells in the gut and possibly other mucosal tissues. However, this has been difficult to test because MAIT cells are rare, comprising ~10% of DN T cells in the gut LP and ≤2% of DN {alpha}beta T cells in the mLN of wild-type (WT) mice, and analysis has been limited to relatively indirect molecular methods. Therefore, to create a model in which these cells are more abundant, we generated transgenic mice expressing the conserved TCR{alpha} chain (V{alpha}19i) characteristic of MAIT cells. To unequivocally identify MR1-restricted cells expressing the invariant V{alpha}19i chain, we crossed transgenic mice with TCR{alpha}–/– (C{alpha}–/–) mice to eliminate expression of endogenous TCR{alpha} chains and then to MR1–/– mice. By directly comparing V{alpha}19iC{alpha}–/–MR1+/+ and V{alpha}19iC{alpha}–/–MR1–/– mice, we were able to distinguish and characterize a population of V{alpha}19i T cells dependent on MR1 for development. We first demonstrated that MR1-restricted V{alpha}19i transgenic T cells recapitulate what is known about endogenous MAIT cells. Moreover, we present further characterization of this population and, for the first time, demonstrate functional capacity of MR1-restricted V{alpha}19i T cells.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Mice

To generate V{alpha}19i transgenic mice, we amplified a DNA fragment containing the canonical rearranged V{alpha}19-J{alpha}33 MAIT cell junction using a nested set of oligonucleotides located 5' of the V{alpha}19 leader sequence and 3' of J{alpha}33 and genomic DNA isolated from a sample enriched for DN {alpha}betaTCR+ cells as a template. Previous experiments demonstrated that >90% of V{alpha}19-J{alpha}33 junctions amplified from this DNA carry the canonical MAIT cell junction (3). The 1.1-kb PCR fragment was cloned and sequenced. A plasmid containing the correct rearrangement and no PCR-induced errors was cloned into the TCR{alpha} expression vector developed by D. Mathis and C. Benoist (Harvard University, Boston, MA) (19). A purified fragment free of vector sequence was injected into C57BL/6 one-cell embryos by the Department of Pathology and Immunology Transgenic/Knockout Core Facility at Washington University. We obtained two founders, one carrying 2 (V{alpha}19i2) and one carrying >10 (V{alpha}19i10) copies of the transgene. Both were bred to C{alpha}–/– C57BL/6 (B6) mice (B6.129S2-Tcratm1Mom/J; The Jackson Laboratory) and MR1–/– mice, which have been described (3). The MR1–/– mice used for these studies had been backcrossed to B6 mice for 10 generations. Mice were typed by Southern blot for the TCR{alpha} mutation and PCR for the V{alpha}19i transgenes and MR1 deletion. All animal studies were approved by the Washington University School of Medicine Animal Studies Committee.

Isolation of cells

Thymus, spleen, and mLN cell suspensions were prepared by mechanical disruption of tissues. Intraepithelial lymphocytes (IEL) and LP lymphocytes (LPL) were isolated from the small intestine. Briefly, intestines were excised at the stomach and the cecum and flushed with PBS; after Peyer’s patches were removed, they were opened longitudinally and washed in HBSS. The intestines were then cut into small fragments and shaken vigorously in 100 ml of HBSS with 2 mM DTT (Sigma-Aldrich) for 30 min at 37°C. The cell suspensions were passed through a 70-µm cell strainer and centrifuged, and IEL were isolated on 30–70% discontinuous Percoll gradients. The tissue fragments were incubated for an additional 30 min, shaking at 37°C in HBSS with 1 mM DTT and 5 mM EDTA, and the remaining IEL/enterocytes were discarded. The remaining tissue was incubated in 20 ml of DMEM/5% BCS (HyClone) with 0.1 mg/ml DNase I (Roche) and 35 µg/ml Liberase Blenzymes 3 (Roche) for 20–30 min at 37°C and filtered through a 70-µm cell strainer, and LPL were isolated on 30–70% Percoll gradients.

Flow cytometry and cell sorting

The following mAbs were purchased from BD Pharmingen: anti-TCR{alpha}beta (H57-597), anti-TCR{gamma}{delta} (GL3), anti-Vbeta2TCR (B20.6), anti-Vbeta4TCR (KT4), anti-Vbeta5.1–5.2TCR (MR9-4), anti-Vbeta6TCR (RR4-7), anti-Vbeta7TCR (TR310), anti-Vbeta8.1–2TCR (MR5-2), anti-Vbeta8.3TCR (1B3.3), anti-Vbeta9 (MR10-2), anti-Vbeta10bTCR (B21.5), anti-Vbeta11TCR (RR3-15), anti-Vbeta12TCR (MR11-1), anti-Vbeta13TCR (MR12-3), anti-Vbeta14TCR (14-2), anti-CD4 (GK1.5 or RM4-5), anti-CD8{alpha} (53-6.7), anti-CD45RB (16A), anti-CD62L (Mel-14), anti-NK1.1 (PK136), anti-CD69 (H1.2F3), anti-CD44 (IM7), and anti-CD25 (7D4). Anti-ICOS (15F9) mAb was purchased from eBioscience. Cell suspensions were stained using a combination of Abs conjugated to TriColor, FITC, PE, and allophycocyanin (or biotin followed by allophycocyanin-conjugated streptavidin) (Molecular Probes), and analyzed on a FACScan with CellQuest software (BD Biosciences). Dead cells were excluded by propidium iodide gating. T cell subsets were sorted as indicated in the figures from spleen and mLN using a MoFlo cytometer (DakoCytomation). Analysis of sorted cells confirmed ≥95% purity.

Cytokine secretion

Cells were cultured at the indicated density for 24–72 h at 37°C with 5% CO2 in 100 µl of RPMI 1640 medium containing 10% BCS (HyClone) in 96-well plates that had been coated overnight at 4°C with the indicated concentrations of anti-CD3{epsilon} (145-2C11 or KT3) and anti-CD28 (37.51) mAbs (BD Pharmingen). Supernatants were assayed for IFN-{gamma}, TNF-{alpha}, IL-5, IL-4, IL-2, IL-10, IL-6, MCP-1, and IL-12p70 using cytometric bead arrays included in the mouse inflammation and mouse Th1/Th2 cytokine kits (BD Pharmingen) according to the manufacturer’s instructions.

Intracellular detection of cytokines

Cells were stimulated with plate-bound anti-CD3{epsilon} (145-2C11) and anti-CD28 (37.51) mAbs (BD Pharmingen) for 48 h. PMA (10–7 M; Sigma-Aldrich) and ionomycin (0.8 µg/ml; Sigma-Aldrich) were added for the last 6 h and monensin (2 µM; Sigma-Aldrich) for the last 4 h of stimulation. Cells were stained for cell surface markers before fixation with 2% formaldehyde for 15 min. Cells were washed and then permeabilized with saponin (0.5%; Sigma-Aldrich) for intracellular staining with the following FITC- and allophycocyanin-labeled mAb as indicated: anti-IFN-{gamma}-FITC (XMG1.2), anti-IL10-allophycocyanin (JES5-16E3), and anti-IL4-allophycocyanin (11B11) (BD Pharmingen).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
MR1-restricted V{alpha}19i transgenic T cells recapitulate what is known about endogenous MAIT cells

We obtained two V{alpha}19i transgenic founders, one carrying 2 (V{alpha}19i2) and one carrying >10 (V{alpha}19i10) copies of the transgene. Both founders were bred to C{alpha}–/– B6 and MR1–/– B6 mice to generate V{alpha}19iC{alpha}–/–MR1+/+ and V{alpha}19iC{alpha}–/–MR1–/– mice; all data shown are from mice on these backgrounds. The low- and high-copy mice had very similar phenotypes, and the subtle differences noted are probably due to higher expression of the transgene in the V{alpha}19i10 mice. Total thymocyte numbers were reduced ~40% in the low-copy-number and 60% in the high-copy-number V{alpha}19i transgenic mice, compared with B6 controls (Table I). Total spleen cell numbers also were reduced, and the peripheral lymph nodes were <25% of WT size in both lines of V{alpha}19i transgenic mice, whereas the mLN were of normal size and cellularity. T cell frequency was reduced ~2-fold in the mLN and spleens of both transgenic lines, compared with WT mice. However, T cell frequency and numbers were similar in the thymus, mLN, spleen, and IEL of V{alpha}19iC{alpha}–/–MR1–/– and V{alpha}19iC{alpha}–/–MR1+/+ animals, and a slight increase in T cell frequency was observed in the intestinal LP of V{alpha}19iC{alpha}–/–MR1+/+ mice (Table I).


View this table:
[in this window]
[in a new window]
  		Table I. T cell subsets in V{alpha}19i transgenic micea

 
To identify T cells dependent on MR1 for development, we compared T cell subsets in the thymus, mLN, spleen, IEL, and LP of V{alpha}19iC{alpha}–/–MR1–/– and V{alpha}19iC{alpha}–/–MR1+/+ mice. Thymii from all transgenic mice had an increased proportion of DN and very few CD4+ or CD8+ single-positive T cells (Fig. 1A); the majority of TCRbetahigh thymocytes were DN in these mice (Fig. 1B, Table I). Because an aberrant population of mature DN T cells develops in most TCR{alpha} transgenic mice due to premature expression of the transgene, this was not unexpected (20, 21). In fact, a high proportion of mature T cells were DN in the periphery of both lines of transgenic mice on C{alpha}–/–MR1–/– and C{alpha}–/–MR1+/+ backgrounds. However, the DN and CD4low population of T cells was augmented in all lymphoid organs and highly enriched in the LP of V{alpha}19iC{alpha}–/–MR1+/+ mice, compared with V{alpha}19iC{alpha}–/–MR1–/– mice (Fig. 1B and Table I). This was evident in both lines of transgenic mice, indicating that at least a proportion of DN and CD4low transgenic T cells are dependent on MR1 for development in the V{alpha}19iC{alpha}–/–MR1+/+ mice.


Figure 1
View larger version (58K):
[in this window]
[in a new window]
  		FIGURE 1. A, CD4 and CD8 expression on total thymocytes isolated from transgenic and B6 mice. The percentage of cells in each quadrant is shown. B, CD4 and CD8 expression on TCRbeta+ cells from V{alpha}19i2 transgenic mice. The percentage of cells in each quadrant is shown. Compiled data are presented in Table I.

 
Because the V{alpha}19i TCR{alpha} chain preferentially associates with Vbeta8.1, Vbeta8.2, and Vbeta6 on endogenous MAIT cells (2), we evaluated TCRbeta expression on CD4high, CD8+, and DN/CD4low mLN T cells from transgenic mice on C{alpha}–/–MR1–/– and C{alpha}–/–MR1+/+ backgrounds as well as WT controls (Fig. 2). A slight increase in the frequency of CD4high T cells expressing Vbeta6 and Vbeta8.1/2 was seen in V{alpha}19i transgenic mice on an MR1+ background, and differences between MR1–/– and MR1+ mice were more evident in the V{alpha}19i10 line (Fig. 2B). An increase in the frequency of CD8+ T cells expressing Vbeta8.1/2 was evident only in the high-copy-number V{alpha}19iC{alpha}–/–MR1+/+ mice, and the frequency of Vbeta6+ CD8+ T cells was slightly higher in both V{alpha}19i transgenic lines on MR1+/+ vs MR1–/– backgrounds (Fig. 2C). Much more striking, the frequency of DN/CD4low T cells expressing either Vbeta6 or Vbeta8.1/2 in both high- and low-copy V{alpha}19iC{alpha}–/–MR1+/+ mice was double that observed in V{alpha}19iC{alpha}–/–MR1–/– mice (Fig. 2D). In MR1-deficient transgenic mice, 23–25% of mLN DN/CD4low T cells expressed Vbeta6 or Vbeta8.1/2, whereas 47–51% of this population expressed one of these three TCRbeta chains in MR1+/+ mice (Fig. 2D and Table II). This skewing also was evident in DN/CD4low T cells from the thymus and spleen and was more pronounced in those isolated from the LP of V{alpha}19iC{alpha}–/–MR1+/+ mice (Table II). In summary, MR1-restricted V{alpha}19i transgenic T cells recapitulate much of what is known about endogenous MAIT cells: they are primarily DN or CD4low; preferentially express Vbeta8.1, Vbeta8.2, or Vbeta6; and are more frequent in the gut LP. Therefore, these V{alpha}19i transgenic mice provide a valid model for further characterization of MAIT cell development and function.


Figure 2
View larger version (24K):
[in this window]
[in a new window]
  		FIGURE 2. TCRVbeta repertoire of T cells from V{alpha}19i transgenic mice. Cells were gated as shown in A, and the percentage of TCRbeta+ cells expressing the Vbeta chains indicated are shown for CD4high (B), CD8+ (C), and DN/CD4low (D) T cells. T cells from V{alpha}19i2 and V{alpha}19i10 transgenic mice on a C{alpha}–/–MR1–/– background had similar repertoires, so these data were pooled. The data are derived from five B6, two V{alpha}19iC{alpha}–/–MR1–/–, two V{alpha}192C{alpha}–/–MR1+/+, and three V{alpha}19i10C{alpha}–/–MR1+/+ mice. Compiled data for Vbeta8.1, Vbeta8.2, and Vbeta6 expression are presented in Table II.

 

View this table:
[in this window]
[in a new window]
  		Table II. Frequency of {alpha}betaT cells expressing Vbeta8.1, Vbeta8.2, and Vbeta6a

 
MR1-restricted V{alpha}19i T cells have an NK T cell phenotype

Given the parallels between MAIT and V{alpha}24/14i NK T cells, we assessed NK1.1 expression on T cells from V{alpha}19i transgenic mice. Indeed, a high proportion of TCRbeta+ cells expressed NK1.1 in V{alpha}19iC{alpha}–/–MR1+/+ but not V{alpha}19iC{alpha}–/–MR1–/– mice (Fig. 3A). Six to 9% of TCRbeta+ cells from the mLN and 11–15% from the spleen of V{alpha}19iC{alpha}–/–MR1+/+ mice expressed NK1.1; 20–30% of V{alpha}19i transgenic T cells in the LP were NK1.1+ (Fig. 3A and Table I). In the spleen and mLN, the majority of V{alpha}19i NK1.1+ T cells were DN (60–80%), and the remaining were either CD4low (10–30%) or CD8low (10%). A slightly higher proportion expressed CD8 in the LP (data not shown). As expected, the majority of MR1-restricted NK1.1+ V{alpha}19i T cells expressed either Vbeta6 or Vbeta8.1/2 (Table II).


Figure 3
View larger version (39K):
[in this window]
[in a new window]
  		FIGURE 3. A, NK1.1 expression on T cells from V{alpha}19i2 transgenic and control mice. Cells were stained for TCRbeta and NK1.1; the percentage of NK1.1+ cells in each sample is shown. Compiled data for NK1.1 expression are presented in Table I. B, Cell surface phenotype of MR1-restricted transgenic T cells. Left, mLN cells from V{alpha}192C{alpha}–/–MR1–/– and V{alpha}192C{alpha}–/–MR1+/+ mice were stained for CD4/CD8 and Vbeta8.1/8.2/6 and the panel of cell surface markers indicated; expression of these markers on the CD4lowCD8lowVbeta8.1/8.2/6+ population is shown below. Right, Spleen cells from V{alpha}192C{alpha}–/–MR1+/+ and B6 mice were stained for TCRbeta and NK1.1, and expression of the panel of cell surface markers on NK1.1+ TCRbeta+ T cells is shown below. Bottom, The percentage of NK1.1+ cells in the CD4lowCD8lowVbeta8.1/8.2/6+ mLN population and ICOS expression on NK1.1+TCRbeta+ T cells.

 
To further characterize transgenic MAIT cells, we stained V{alpha}19i transgenic T cells with a panel of cell surface markers, initially focusing on Vbeta8.1/2+Vbeta6+CD4lowCD8lowDN cells. In mLN cells from V{alpha}19iC{alpha}–/–MR1+/+ mice, 50–60% of this population was CD44high, CD45RBlow, and CD62Lneg and expressed low levels of CD25; ~40% expressed CD69; and most expressed intermediate-high levels of ICOS (Fig. 3B). Relatively few Vbeta8.1/2+Vbeta6+CD4lowCD8lowDN cells from V{alpha}19iC{alpha}–/–MR1–/– mice expressed similar levels of these cell surface markers, indicating that this phenotype results from interaction of V{alpha}19i with MR1. In similar analyses, we found that almost 90% of NK1.1+ T cells from V{alpha}19iC{alpha}–/–MR1+/+ mice share this phenotype as do most NK1.1+ T cells from B6 mice. High levels of CD25 are characteristic of T regulatory cells (Treg) dependent on the transcription factor FoxP3 (22). Although MR1-restricted V{alpha}19i T cells express low levels of CD25, they do not express FoxP3 as determined by RT-PCR of RNA from sorted cells (data not shown). ICOS is associated with IL-10 production (23) and may reflect encounter with self ligand; for instance, ICOS is expressed on naive DO11.1 transgenic T cells specific for OVA323–339 from mice expressing high levels of OVA under control of the rat insulin promoter (24). Overall, these data are consistent with the observation that human MAIT cells have an activated/memory phenotype (2) and, furthermore, demonstrate that a T cell population phenotypically similar to V{alpha}14i NK T cells develops in V{alpha}19i transgenic mice on an MR1+/+ (but not MR1–/–) background and that 25–50% of this population expresses NK1.1.

Transgenic MAIT cells secrete IFN-{gamma}, IL-4, IL-5, and IL-10 following TCR ligation

We initially assessed T cell proliferation in response to plate-bound anti-CD3 or anti-CD3 and anti-CD28 mAbs in bulk cultures from spleen and/or mLN and found that T cells from V{alpha}19iC{alpha}–/–MR1+/+ mice proliferated poorly in comparison to those from V{alpha}19iC{alpha}–/–MR1–/– and WT mice (data not shown). We then measured cytokine secretion from V{alpha}19iC{alpha}–/–MR1–/–, V{alpha}19iC{alpha}–/–MR1+/+, and control mLN and splenic T cells after 24–72 h of culture with plate-bound anti-CD3 with or without anti-CD28. Although all T cells secreted IFN-{gamma}, and TNF-{alpha}, we detected significant amounts of IL-4, IL-5, and IL-10 in supernatants from V{alpha}19iC{alpha}–/–MR1+/+ but not V{alpha}19iC{alpha}–/–MR1–/– or B6 mice (Fig. 4A). Conversely, more IL-2 was detected in supernatants from V{alpha}19iC{alpha}–/–MR1–/– and B6 T cells than from V{alpha}19iC{alpha}–/–MR1+/+ T cells. Very similar results were obtained with LPL in these assays; again, only those isolated from V{alpha}19iC{alpha}–/–MR1+/+ mice secreted significant amounts of IL-4, IL-5, and IL-10; in contrast, IELs from V{alpha}19iC{alpha}–/–MR1+/+ secreted IL-4 and IL-10 but very little IL-5. (Fig. 4B). We repeated these experiments with sorted T cells from spleen and/or mLN and found that only the Vbeta8.1/8.2/6+DN/CD4low T cell population from V{alpha}19iC{alpha}–/–MR1+/+ mice secreted significant levels of IL-4, IL-5, and IL-10 following TCR ligation (Fig. 5). These data were confirmed by staining mLN or splenic T cells for intracellular cytokines following TCR ligation. Only T cells from V{alpha}19iC{alpha}–/–MR1+/+ mice secreted IL-4 and IL-10, and we were able to detect a small population secreting both IFN-{gamma} and IL-4 and another IFN-{gamma} and IL-10 (Fig. 6).


Figure 4
View larger version (29K):
[in this window]
[in a new window]
  		FIGURE 4. Cytokine secretion by T cells from B6 and V{alpha}19i2 transgenic mice. A, mLN cells isolated from B6 and V{alpha}19i2 transgenic mice (1 x 105 TCRbeta+ cells per well) were cultured for 72 h with plate-bound anti-CD3 and anti-CD28, and cytokine secretion was measured using a cytometric bead assay. Data shown are representative of four independent experiments. B, Total IEL or LPL isolated from V{alpha}19i10 (1 x 105 TCRbeta+ cells per well) were cultured for 72 h with plate-bound anti-CD3 and anti-CD28, and cytokine secretion was measured using a cytometric bead assay. Data shown are representative of two independent experiments.

 

Figure 5
View larger version (34K):
[in this window]
[in a new window]
  		FIGURE 5. Cytokine secretion from sorted T cells. mLN T cells from B6 and V{alpha}19i10 transgenic mice were sorted as shown in the panels above. A total of 1 x 105 cells per well were stimulated with anti-CD3 and anti-CD28 for 72 h, and cytokine secretion was measured by cytometric bead assay. IL-4, IL-5, and IL-10 secretion from each of the three sorted populations from B6 and transgenic mice is shown below. Data shown are representative of three independent experiments with T cells sorted as shown. Similar results for the transgenic T cells were obtained using anti-CD3 alone in these assays.

 

Figure 6
View larger version (32K):
[in this window]
[in a new window]
  		FIGURE 6. Cytokine secretion by V{alpha}19i transgenic T cells. mLN cells from V{alpha}19itg2 mice were stimulated for 48 h with anti-CD3; PMA and ionomycin were added for the last 6 h and monensin for the last 4 h of culture. Cells were stained for IL-10, IL-4, and IFN-{gamma} as described in Materials and Methods.

 
IL-4, IL-5, and IFN-{gamma} secretion following TCR ligation in vitro is characteristic of V{alpha}14i NK T cells. In this respect, MR1-restricted V{alpha}19i T cells appear functionally analogous to CD1d-restricted V{alpha}14i NKT cells. To further test this, we injected V{alpha}19i transgenic mice with an anti-CD3 mAb; rapid secretion of IL-4 and IFN-{gamma} following in vivo activation with anti-CD3 in the absence of further stimulation is a hallmark of NK T cell function (25). Ninety minutes after i.v. injection of an anti-CD3 mAb, splenocytes from V{alpha}19iC{alpha}–/–MR1+/+ mice secreted high levels of IL-4 and IFN-{gamma}, whereas splenocytes from identically treated V{alpha}19iC{alpha}–/–MR1–/– mice secreted very little of these cytokines (Fig. 7). This indicates that, like V{alpha}14i NK T cells, MR1-restricted V{alpha}19i transgenic T cells rapidly secrete IL-4 and IFN-{gamma} following CD3 ligation in vivo. Cumulatively, these data demonstrate that at least a proportion of MR1-restricted transgenic MAIT cells are functionally equivalent to V{alpha}14i NK T cells.


Figure 7
View larger version (23K):
[in this window]
[in a new window]
  		FIGURE 7. Cytokine secretion after in vivo injection of anti-CD3. One microgram of anti-CD3 (2C11 in 200 µl of PBS) was injected i.v. into V{alpha}19i and WT controls. Ninety minutes later, spleen cell suspensions were prepared and washed two times. Cells were incubated at 5 x 106 cells/ml in 24-well plates (107 cells per well) for 2 h at 37°C. Cytokine secretion in cell supernatants was measured by cytometric bead array. Each sample was done in triplicate, and these data are representative of three independent experiments.

 
Finally, we asked whether NK1.1+ and NK1.1neg MR1-restricted T cells are functionally distinct. In experiments with sorted cells, NK1.1+ V{alpha}19i transgenic T cells from MR1+/+ mice consistently secreted IL-4 and IL-5, but neither V{alpha}19i transgenic nor WT NK1.1+ T cells secreted high levels of IL-10 in these assays (Fig. 8 and data not shown). Conversely, NK1.1negVbeta8.1/8.2/6+ T cells from MR1+/+ mice consistently secreted high levels of IL-10 and variable amounts of IL-4 and IL-5 (Fig 8). Therefore, MR1-restricted NK1.1+ V{alpha}19i transgenic T cells are functionally as well as phenotypically comparable to V{alpha}14i NK T cells. The NK1.1neg population is more heterogeneous and contains most of the IL-10-producing T cells. This may be related to differences in cell surface expression of ICOS on these two populations. In a functional analysis of T cells isolated from unmanipulated mice, Löhning et al. (23) found that T cells expressing intermediate levels of ICOS secreted IL-4, IL-5, and IL-13, whereas those expressing high levels of ICOS secreted primarily IL-10. Both B6 and MR1-restricted V{alpha}19i NK1.1+ T cells express intermediate levels of ICOS; very few NK1.1+ T cells are ICOShigh (Fig. 3B). However, the NK1.1neg MR1-restricted V{alpha}19i transgenic T cell population contains both ICOS-intermediate and -high cells. Thus, cytokine secretion by NK1.1+ and NK1.1neg MR1-restricted V{alpha}19i T cells correlates well with cell surface expression of ICOS.


Figure 8
View larger version (30K):
[in this window]
[in a new window]
  		FIGURE 8. Cytokine secretion from NK1.1+ and NK1.1neg V{alpha}19i2 transgenic T cells. CD8neg splenic and mLN T cells were sorted as shown in the panels above. Cells were stimulated with anti-CD3 and anti-CD28 for 24–72 h as indicated, and cytokine secretion was measured by cytometric bead assay. IL-4, IL-5, IL-10, and IFN-{gamma} secretion from each population is shown below. These data are representative of four independent experiments with cells sorted to distinguish NK1.1+ and NK1.1neg MR1-restricted V{alpha}19i transgenic cells; the amounts of IL-4 and IL-5 secreted by the NK1.1neg population varied in these assays.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
These V{alpha}19i transgenic mice recapitulate what is known about MAIT cell development: MR1-restricted transgenic V{alpha}19i T cells are DN or CD4low; preferentially express Vbeta8.1, Vbeta8.2, and Vbeta6; and are most abundant in the intestinal LP. In addition, 25–50% of transgenic MAIT cells express NK1.1, and most have a cell surface phenotype similar to that of V{alpha}14i NK T cells. This suggests that selection of V{alpha}19i T cells by MR1 closely mimics that of V{alpha}14i T cells by CD1d; both invariant TCR{alpha} chains direct the development of NK T cells in the presence of appropriate MHC class Ib molecules. V{alpha}14i NK T cells are the product of unique selection events mediated by CD1+ double-positive cells in the thymus (26). Interestingly, functional NK T cells also can develop in transgenic mice expressing TCRs specific for MHC class Ia or class II/peptide in the absence of endogenous TCRV{alpha} rearrangements (27, 28, 29). This suggests that selection of NK T cells does not require V{alpha}14i and CD1d per se. Given this, generation of V{alpha}19i transgenic NK1.1+ T cells probably reflects this mode of selection, which is clearly favored in mice expressing MR1. Consistent with this, Shimamura and Huang (30) reported that ~50% of T cell hybridomas established from NK1.1+ T cells from CD1d–/– mice express the V{alpha}19i MAIT invariant TCR{alpha} chain. In addition, the frequency of NK1.1+ T cells in the V{alpha}19i transgenic mice is similar to that seen in transgenic mice expressing a V{alpha}14i TCR{alpha} chain driven by a V{alpha} promoter and Ig enhancer (31).

How do MAIT cells fit in with the known NK T cell subsets in WT mice?

Eight-five percent of murine NK T cells express V{alpha}14i; almost all thymic and liver NK T cells are CD1d dependent and express the V{alpha}14i chain, whereas CD1d- and thymus-independent NKT cells are enriched in the spleen and BM (9, 10, 11). Extensive repertoire analysis of DN NK T cells from the spleen and BM revealed two distinct populations: one strikingly skewed toward V{alpha}14, Vbeta7, Vbeta8.1/2/3, and Vbeta2 (V{alpha}14i NK T cells), and the other with an essentially unbiased TCRV{alpha} and -Vbeta repertoire (32). This indicates that V{alpha}19i T cells do not comprise a significant proportion of the V{alpha}14i-negative NK1.1+ T cell population in the spleen and BM of normal mice. This is not surprising, given that MAIT cells are detected primarily in the gut LP. The large population of NK1.1+ T cells residing in the LP of V{alpha}19iC{alpha}–/–MR1+/+ mice may correspond to a functionally significant population of T cells in the LP of WT mice. In addition, the few NK1.1+ T cells present in the mLN of WT mice may include MAIT cells. Thus, although MAIT cells share many characteristics of NK T cells, they are a distinct T cell lineage that intersects very little with the known NK T cell subsets (33).

Furthermore, we observed what may be two functionally distinct populations of MR1-restricted V{alpha}19i T cells. One subset appears virtually identical with V{alpha}14i NK T cells in that it secretes IFN-{gamma}, IL-4, and IL-5 following in vitro TCR ligation and responds rapidly to in vivo injection of anti-CD3 mAb. More unique are the NK1.1neg MR1-restricted V{alpha}19i T cells that secrete IL-10 following TCR ligation in vitro. V{alpha}14i NK T cells can secrete IL-10, but this has been shown only after in vivo manipulation: IL-10-secreting NK T cells have been isolated from mice that have been inoculated with Ag in the anterior chamber of the eye and from mice that have been injected with the potent CD1d ligand {alpha}-galactosylceramide and then infected with Toxoplasma gondii (34, 35). In both cases, the IL-10-secreting NK T cells were critical for inducing or recruiting Treg. MR1-restricted V{alpha}19i transgenic T cells more closely resemble Treg induced by encounter with self Ag in the periphery; for example, DO11.1 transgenic T cells specific for OVA323–339 isolated from mice expressing high levels of OVA under control of the rat insulin promoter secrete IL-10 after in vitro activation, as do T cells from transgenic mice expressing a TCR specific for a peptide epitope of glutamic acid decarboxylase 65 (24, 36). V{alpha}14i NK T cells have the capacity to secrete Th1 and/or Th2 cytokines depending on their mode of stimulation. It is quite possible that MAIT cells share this phenotype, and what we have observed are different aspects of MAIT cell differentiation. However, clearly, the capacity to secrete IL-10 is favored more by V{alpha}19i/MR1 than by V{alpha}14i/CD1d interactions in naive mice

The main differences between MAIT cells and V{alpha}14i NK T cells are their restriction elements and anatomical location. V{alpha}14i NK T cells are most frequent in internal organs, including the thymus, liver, BM, and spleen, and recognize endogenous and exogenous glycosphingolipids bound to CD1d; unlike MAIT cells, they are present in germfree mice (37). MAIT cells accumulate primarily in the gut LP and mLN and are dependent on MR1 and commensal flora for selection/expansion. They may be present in other mucosal tissues and have been reported to accumulate in some of the CNS lesions of multiple sclerosis patients as well as in peripheral nerve samples from patients with chronic inflammatory demyelinating polyneuropathy (38). It is probable that they recognize MR1 in the context of several related ligands, some of which are likely to be derived from or induced by commensal flora. Our data demonstrate that V{alpha}19i MAIT cells are phenotypically and functionally very similar to NK T cells and, hence, may influence the gut and perhaps other immune responses in a variety of ways, depending on their mode of stimulation. In addition, at least a proportion of these cells are primed for IL-10 production. As MAIT cells are relatively rare, even in the gut LP, one of their primary roles may be to induce or recruit other Treg, as has been shown for V{alpha}14i NK T cells. The V{alpha}19i transgenic mice clearly illustrate the striking parallels between V{alpha}19i MAIT cells and V{alpha}14i NK T cells and provide a valuable model for further probing MAIT cell function as well as a possible means to identify MR1 ligands.


   Acknowledgments
 
We thank Drs. D. Mathis and C. Benoist for the TCR{alpha} expression cassette; M. White for embryo injection; B. Eades and J. Hughes for cell sorting; and M. Colonna, T. Hansen, M. Cella, C. Kemper, and B. Sleckman for helpful discussions and/or critically reading the manuscript.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by National Institutes of Health Grants 1R21AI059631-01 and 1R21AI062889-01 (to S.G.). Mice were housed in a facility supported by National Center for Research Resources Grant C06 RR012466. I.K. was supported by a fellowship from the Sankyo Foundation of Life Science. Back

2 Address correspondence and reprint requests to Dr. Susan Gilfillan, Department of Pathology and Immunology, Washington University School of Medicine, Campus Box 8118, 660 South Euclid Avenue, St. Louis, MO 63110. E-mail address: susang{at}pathbox.wustl.edu Back

3 Abbreviations used in this paper: MAIT, mucosal-associated invariant T; DN, double negative; mLN, mesenteric lymph node; LP, lamina propria; BM, bone marrow; WT, wild type; C{alpha}–/–, TCR{alpha}–/–; LPL, LP lymphocyte; IEL, intraepithelial lymphocyte; Treg, T regulatory cell. Back

Received for publication October 6, 2005. Accepted for publication November 15, 2005.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Porcelli, S., C. E. Yockey, M. B. Brenner, S. P. Balk. 1993. Analysis of T cell antigen receptor (TCR) expression by human peripheral blood CD4–8– {alpha}/beta T cells demonstrates preferential use of several Vbeta genes and an invariant TCR {alpha}-chain. J. Exp. Med. 178: 1-16. [Abstract/Free Full Text]
  2. Tilloy, F., E. Treiner, S. H. Park, C. Garcia, F. Lemonnier, H. de la Salle, A. Bendelac, M. Bonneville, O. Lantz. 1999. An invariant T cell receptor {alpha}-chain defines a novel TAP-independent major histocompatibility complex class Ib-restricted {alpha}beta T cell subpopulation in mammals. J. Exp. Med. 189: 1907-1921. [Abstract/Free Full Text]
  3. Treiner, E., L. Duban, S. Bahram, M. Radosavljevic, V. Wanner, F. Tilloy, P. Affaticati, S. Gilfillan, O. Lantz. 2003. Selection of evolutionarily conserved mucosal-associated invariant T cells by MR1. Nature 422: 164-169. [Medline]
  4. Riegert, P., V. Wanner, S. Bahram. 1998. Genomics, isoforms, expression, and phylogeny of the MHC class I-related MR1 gene. J. Immunol. 161: 4066-4077. [Abstract/Free Full Text]
  5. Yamaguchi, H., M. Hirai, Y. Kurosawa, K. Hashimoto. 1997. A highly conserved major histocompatibility complex class I-related gene in mammals. Biochem. Biophys. Res. Commun. 238: 697-702. [Medline]
  6. Yamaguchi, H., K. Hashimoto. 2002. Association of MR1 protein, an MHC class I-related molecule, with beta2-microglobulin. Biochem. Biophys. Res. Commun. 290: 722-729. [Medline]
  7. Miley, M. J., S. M. Truscott, Y. Y. Yu, S. Gilfillan, D. H. Fremont, T. H. Hansen, L. Lybarger. 2003. Biochemical features of the MHC-related protein 1 consistent with an immunological function. J. Immunol. 170: 6090-6098. [Abstract/Free Full Text]
  8. Huang, S., S. Gilfillan, M. Cella, M. J. Miley, O. Lantz, L. Lybarger, D. H. Fremont, T. H. Hansen. 2005. Evidence for MR1 antigen presentation to mucosal-associated invariant T cells. J. Biol. Chem. 280: 21183-21193. [Abstract/Free Full Text]
  9. Godfrey, D. I., K. J. Hammond, L. D. Poulton, M. J. Smyth, A. G. Baxter. 2000. NKT cells: facts, functions and fallacies. Immunol. Today 21: 573-583. [Medline]
  10. Kronenberg, M., L. Gapin. 2002. The unconventional lifestyle of NKT cells. Nat. Rev. Immunol. 2: 557-568. [Medline]
  11. Benlagha, K., A. Bendelac. 2000. CD1d-restricted mouse V{alpha}14 and human V{alpha}24 T cells: lymphocytes of innate immunity. Semin. Immunol. 12: 537-542. [Medline]
  12. Zhou, D., J. Mattner, C. Cantu, III, N. Schrantz, N. Yin, Y. Gao, Y. Sagiv, K. Hudspeth, Y. P. Wu, T. Yamashita, et al 2004. Lysosomal glycosphingolipid recognition by NKT cells. Science 306: 1786-1789. [Abstract/Free Full Text]
  13. Kinjo, Y., D. Wu, G. Kim, G. W. Xing, M. A. Poles, D. D. Ho, M. Tsuji, K. Kawahara, C. H. Wong, M. Kronenberg. 2005. Recognition of bacterial glycosphingolipids by natural killer T cells. Nature 434: 520-525. [Medline]
  14. Mattner, J., K. L. Debord, N. Ismail, R. D. Goff, C. Cantu, III, D. Zhou, P. Saint-Mezard, V. Wang, Y. Gao, N. Yin, et al 2005. Exogenous and endogenous glycolipid antigens activate NKT cells during microbial infections. Nature 434: 525-529. [Medline]
  15. Lisbonne, M., S. Diem, A. de Castro Keller, J. Lefort, L. M. Araujo, P. Hachem, J. M. Fourneau, S. Sidobre, M. Kronenberg, M. Taniguchi, et al 2003. Cutting edge: invariant V{alpha}14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model. J. Immunol. 171: 1637-1641. [Abstract/Free Full Text]
  16. Akbari, O., P. Stock, E. Meyer, M. Kronenberg, S. Sidobre, T. Nakayama, M. Taniguchi, M. J. Grusby, R. H. DeKruyff, D. T. Umetsu. 2003. Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity. Nat. Med. 9: 582-588. [Medline]
  17. Heller, F., I. J. Fuss, E. E. Nieuwenhuis, R. S. Blumberg, W. Strober. 2002. Oxazolone colitis, a Th2 colitis model resembling ulcerative colitis, is mediated by IL-13-producing NK-T cells. Immunity 17: 629-638. [Medline]
  18. Van Kaer, L., S. Joyce. 2005. Innate immunity: NKT cells in the spotlight. Curr. Biol. 15: R429-R431. [Medline]
  19. Kouskoff, V., K. Signorelli, C. Benoist, D. Mathis. 1995. Cassette vectors directing expression of T cell receptor genes in transgenic mice. J. Immunol. Methods 180: 273-280. [Medline]
  20. Baldwin, T. A., M. M. Sandau, S. C. Jameson, K. A. Hogquist. 2005. The timing of TCR{alpha} expression critically influences T cell development and selection. J. Exp. Med. 202: 111-121. [Abstract/Free Full Text]
  21. Huang, C. Y., O. Kanagawa. 2004. Impact of early expression of TCR {alpha}-chain on thymocyte development. Eur. J. Immunol. 34: 1532-1541. [Medline]
  22. Kronenberg, M., A. Rudensky. 2005. Regulation of immunity by self-reactive T cells. Nature 435: 598-604. [Medline]
  23. Löhning, M., A. Hutloff, T. Kallinich, H. W. Mages, K. Bonhagen, A. Radbruch, E. Hamelmann, R. A. Kroczek. 2003. Expression of ICOS in vivo defines CD4+ effector T cells with high inflammatory potential and a strong bias for secretion of interleukin 10. J. Exp. Med. 197: 181-193. [Abstract/Free Full Text]
  24. Kohyama, M., D. Sugahara, S. Sugiyama, H. Yagita, K. Okumura, N. Hozumi. 2004. Inducible costimulator-dependent IL-10 production by regulatory T cells specific for self-antigen. Proc. Natl. Acad. Sci. USA 101: 4192-4197. [Abstract/Free Full Text]
  25. Yoshimoto, T., W. E. Paul. 1994. CD4pos, NK1.1pos T cells promptly produce interleukin 4 in response to in vivo challenge with anti-CD3. J. Exp. Med. 179: 1285-1295. [Abstract/Free Full Text]
  26. Wei, D. G., H. Lee, S. H. Park, L. Beaudoin, L. Teyton, A. Lehuen, A. Bendelac. 2005. Expansion and long-range differentiation of the NKT cell lineage in mice expressing CD1d exclusively on cortical thymocytes. J. Exp. Med. 202: 239-248. [Abstract/Free Full Text]
  27. Legendre, V., C. Boyer, S. Guerder, B. Arnold, G. Hammerling, A. M. Schmitt-Verhulst. 1999. Selection of phenotypically distinct NK1.1+ T cells upon antigen expression in the thymus or in the liver. Eur. J. Immunol. 29: 2330-2343. [Medline]
  28. Schulz, R. J., A. Parkes, E. Mizoguchi, A. K. Bhan, S. Koyasu. 1996. Development of CD4–CD8– {alpha}beta TCR+NK1.1+ T lymphocytes: thymic selection by self antigen. J. Immunol. 157: 4379-4389. [Abstract]
  29. Iwabuchi, C., K. Iwabuchi, K. Nakagawa, T. Takayanagi, H. Nishihori, S. Tone, K. Ogasawara, R. A. Good, K. Onoe. 1998. Intrathymic selection of NK1.1+{alpha}beta T cell antigen receptor (TCR)+ cells in transgenic mice bearing TCR specific for chicken ovalbumin and restricted to I-Ad. Proc. Natl. Acad. Sci. USA 95: 8199-8204. [Abstract/Free Full Text]
  30. Shimamura, M., Y. Y. Huang. 2002. Presence of a novel subset of NKT cells bearing an invariant V{alpha}19.1-J{alpha}26 TCR {alpha}-chain. FEBS Lett. 516: 97-100. [Medline]
  31. Bendelac, A., R. D. Hunziker, O. Lantz. 1996. Increased interleukin 4 and immunoglobulin E production in transgenic mice overexpressing NK1 T cells. J. Exp. Med. 184: 1285-1293. [Abstract/Free Full Text]
  32. Apostolou, I., A. Cumano, G. Gachelin, P. Kourilsky. 2000. Evidence for two subgroups of CD4–CD8– NKT cells with distinct TCR {alpha}beta repertoires and differential distribution in lymphoid tissues. J. Immunol. 165: 2481-2490. [Abstract/Free Full Text]
  33. Godfrey, D. I., H. R. MacDonald, M. Kronenberg, M. J. Smyth, L. Van Kaer. 2004. NKT cells: what’s in a name?. Nat. Rev. Immunol. 4: 231-237. [Medline]
  34. Ronet, C., S. Darche, M. Leite de Moraes, S. Miyake, T. Yamamura, J. A. Louis, L. H. Kasper, D. Buzoni-Gatel. 2005. NKT cells are critical for the initiation of an inflammatory bowel response against Toxoplasma gondii. J. Immunol. 175: 899-908. [Abstract/Free Full Text]
  35. Sonoda, K. H., D. E. Faunce, M. Taniguchi, M. Exley, S. Balk, J. Stein-Streilein. 2001. NK T cell-derived IL-10 is essential for the differentiation of antigen-specific T regulatory cells in systemic tolerance. J. Immunol. 166: 42-50. [Abstract/Free Full Text]
  36. Tarbell, K. V., M. Lee, E. Ranheim, C. C. Chao, M. Sanna, S. K. Kim, P. Dickie, L. Teyton, M. Davis, H. McDevitt. 2002. CD4+ T cells from glutamic acid decarboxylase (GAD) 65-specific T cell receptor transgenic mice are not diabetogenic and can delay diabetes transfer. J. Exp. Med. 196: 481-492. [Abstract/Free Full Text]
  37. Park, S. H., K. Benlagha, D. Lee, E. Balish, A. Bendelac. 2000. Unaltered phenotype, tissue distribution and function of V{alpha}14+ NKT cells in germ-free mice. Eur. J. Immunol. 30: 620-625. [Medline]
  38. Illes, Z., M. Shimamura, J. Newcombe, N. Oka, T. Yamamura. 2004. Accumulation of V{alpha}7.2-J{alpha}33 invariant T cells in human autoimmune inflammatory lesions in the nervous system. Int. Immunol. 16: 223-230. [Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 J. Exp. Med.Home page
S. Huang, S. Gilfillan, S. Kim, B. Thompson, X. Wang, A. J. Sant, D. H. Fremont, O. Lantz, and T. H. Hansen
MR1 uses an endocytic pathway to activate mucosal-associated invariant T cells
J. Exp. Med., May 12, 2008; 205(5): 1201 - 1211.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Gastrointest. Liver Physiol.Home page
G. Wingender and M. Kronenberg
Role of NKT cells in the digestive system. IV. The role of canonical natural killer T cells in mucosal immunity and inflammation
Am J Physiol Gastrointest Liver Physiol, January 1, 2008; 294(1): G1 - G8.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available