Invariant NKT (iNKT) cells recognize glycolipid Ags, such as the marine sponge-derived glycosphingolipid ␣-galactosylceramide (␣GalCer) presented by the CD1d protein. In vivo activation of iNKT cells with ␣GalCer results in robust cytokine production, followed by the acquisition of an anergic phenotype. Here we have investigated mechanisms responsible for the establishment of ␣GalCer-induced iNKT cell anergy. We found that ␣GalCer-activated iNKT cells rapidly up-regulated expression of the inhibitory costimulatory receptor programmed death (PD)-1 at their cell surface, and this increased expression was retained for at least one month. Blockade of the interaction between PD-1 and its ligands, PD-L1 and PD-L2, at the time of ␣GalCer treatment prevented the induction iNKT cell anergy, but was unable to reverse established iNKT cell anergy. Consistently, injection of ␣GalCer into PD-1-deficient mice failed to induce iNKT cell anergy. However, blockade of the PD-1/PD-L pathway failed to prevent bacterial-or sulfatide-induced iNKT cell anergy, suggesting additional mechanisms of iNKT cell tolerance. Finally, we showed that blockade of PD-1/PD-L interactions enhanced the antimetastatic activities of ␣GalCer. Collectively, our findings reveal a critical role for the PD-1/PD-L costimulatory pathway in the ␣GalCer-mediated induction of iNKT cell anergy that can be targeted for the development of immunotherapies.

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Intestinal intraepithelial lymphocytes (IEL) bear a partially activated phenotype that permits them to rapidly respond to antigenic insults. However, this phenotype also implies that IEL must be highly controlled to prevent misdirected immune reactions. It has been suggested that IEL are regulated through the interaction of the CD8␣␣ homodimer with the thymus leukemia (TL) antigen expressed by intestinal epithelial cells. We have generated and characterized mice genetically-deficient in TL expression. Our findings show that TL expression has a critical role in maintaining IEL effector functions. Also, TL deficiency accelerated colitis in a genetic model of inflammatory bowel disease. These findings reveal an important regulatory role of TL in controlling IEL function and intestinal inflammation.CD8␣␣ ͉ mucosal immunity ͉ colitis ͉ nonclassical MHC ͉ immunoregulation T he intestinal mucosa represents one of the major entry points for antigens into the body, and thus requires a refined immunological system that can prevent the invasion and dissemination of both commensal and pathogenic microorganisms. One of the main immunological compartments involved in regulating mucosal immune responses is comprised by the intraepithelial lymphocytes (IEL). IEL are a population of lymphocytes that reside within the intestinal epithelium, and constitute one of the largest populations of lymphocytes in the body. In mice, 3 main subpopulations of IEL have been identified: TCR␥␦ ϩ T cells expressing mostly the CD8␣␣ homodimer, TCR␣␤ ϩ T cells expressing either CD4 or CD8␣␤ and sometimes coexpressing CD8␣␣, and TCR␣␤ ϩ T cells expressing CD8␣␣ (1). Many reports indicate that these populations function in the recognition of stress signals (2), are involved in the recovery from tissue damage (3, 4), function as conventional memory cells (1), or have natural autoreactivity, suggesting a regulatory role (5, 6). Despite their varied function and phenotype, the great majority of IEL are characterized by a ''partial activation'' state (7). This phenotype suggests that IEL are capable of rapidly responding to stimuli, and therefore, must be held in tight check to prevent unwanted reactions. Thus, an intriguing aspect is the regulation of the effector functions of IEL, which remains incompletely understood.The thymus leukemia (TL) antigen is a nonclassical MHC class I molecule encoded by a locus within the MHC complex (8). TL expression is confined to the surface of intestinal epithelial cells (IEC) (9, 10) and it does not appear to bind an antigenic moiety (11,12). Recently, it has been demonstrated that TL binds preferentially to the CD8␣␣ homodimer (11-17), and it has been suggested that this interaction, at least in vitro, modulates IEL responses.Considering that CD8␣␣ is a prevalent surface marker on IEL, and that these cells reside in close proximity to TL-expressing IEC, we hypothesized that TL has a key role in regulating IEL effector functions. In the present study, we have analyzed mice deficient in the expression of TL and report...

The transcription factor c‐Maf is critical for IL‐4 production and the development of Th2 cells, which promote humoral immunity and protect against extracellular parasites. Yet, little else is known of c‐Maf function in CD4 cells. Here, we identify a novel role for c‐Maf in regulating susceptibility to apoptosis. Overexpression of c‐Maf results in increased susceptibility of CD4 cells to apoptosis induced by multiple stimuli, including growth factor withdrawal, dexamethasone, irradiation, and TCR engagement. This effect is independent of Fas or p53; however, Bcl‐2 expression is reduced in c‐Maf Tg CD4 cells. Immunoprecipitation and Western blot analyses demonstrate that c‐Maf–c‐Myb complex formation is enhanced among T cells from c‐Maf Tg mice compared to non‐Tg littermates following TCR engagement. Unlike non‐Tg T cells, c‐Myb binding to the Bcl‐2 promoter is not detectable in c‐Maf Tg T cells by chromatin immunoprecipitation. In reporter assays, Bcl‐2 promoter activity is reduced by c‐Maf in a dose‐dependent manner. Furthermore, transgene‐mediated Bcl‐2 expression corrects the apoptosis defect observed among c‐Maf Tg CD4 cells. These data suggest that c‐Maf can interact with c‐Myb to reduce Bcl‐2 expression, thereby limiting CD4 cell survival following TCR engagement.

Invariant natural killer T (iNKT) cells are a subset of T lymphocytes that recognizes glycolipid antigens presented by the major histocompatibility complex class I-related protein CD1d. Although iNKT cells have received a lot of attention as targets for the development of immunotherapies, few studies have investigated the in vivo response of iNKT cells to glycolipid antigen activation. Accumulating evidence indicates that iNKT cells generate a dynamic response to in vivo activation by glycolipid antigens that is characterized by surface receptor downmodulation, expansion, cytokine production, cross talk with other cells, homeostatic contraction, and acquisition of an anergic phenotype. These studies provide new insight into the biology of iNKT cells and have important implications for designing safe and effective iNKT cell-based vaccines and therapies.