IFN-γ plays a central role in antitumor immunity. T cell Ig and mucin domain (Tim-3) is expressed on IFN-γ–producing Th1 cells; on interaction with its ligand, galectin-9, Th1 immunity is terminated. In this study, we show that transgenic overexpression of Tim-3 on T cells results in an increase in CD11b+Ly-6G+ cells and inhibition of immune responses. Molecular characterization of CD11b+Ly-6G+ cells reveals a phenotype consistent with granulocytic myeloid-derived suppressor cells. Accordingly, we find that modulation of the Tim-3/galectin-9 (Gal-9) pathway impacts on tumor growth. Similarly, overexpression of Tim-3 ligand, Gal-9, results in an increase in CD11b+Ly-6G+ cells and inhibition of immune responses. Loss of Tim-3 restores normal levels of CD11b+Ly-6G+ cells and normal immune responses in Gal-9 transgenic mice. Our data uncover a novel mechanism by which the Tim-3/Gal-9 pathway regulates immune responses and identifies this pathway as a therapeutic target in diseases where myeloid-derived suppressor cells are disadvantageous.

T-cell immunoglobulin, mucin domain-3 (Tim-3) is a membrane protein expressed at late stages of IFN-c secreting CD4 1 Th1 cell differentiation and constitutively on DC. Ligation of Tim-3 on Th1 cells terminates Th1 immune responses. In addition, Tim-3 plays a role in tolerance induction, although the mechanism by which this is accomplished has yet to be elucidated. While it is clear that Tim-3 plays an important role in the immune system, little is known regarding the molecular pathways that regulate Tim-3 expression. In the current study, we examine the role of Th1-associated transcription factors in regulating Tim-3 expression. Our experiments reveal that Tim-3 expression is regulated by the Th1-specific transcription factor T-bet. This introduces a novel paradigm into the generation of a Th1 response, whereby a transcription factor responsible for effector Th1 cell differentiation also increases the expression of a specific counter-regulatory molecule to ensure appropriate termination of pro-inflammatory Th1 immune responses.Key words: Autoimmunity . T cells . Transcription factors Supporting Information available online Introduction T-cell immunoglobulin, mucin domain-3 (Tim-3) is a type I transmembrane protein, originally identified as specifically and selectively induced upon the differentiation of naive CD4 1 Th cells into IFN-g-secreting Th1 effector cells [1], where it serves to promote termination of Th1 cells upon interaction with its ligand, galectin-9 [2]. Modulation of the Tim-3:galectin-9 pathway has potent effects in vivo as treatment with anti-Tim-3 antibody exacerbates the induction of EAE, an animal model of MS [1], and administration of Tim-3 fusion protein to immunized mice results in an uncontrolled Th1 response [3]. In addition, WT mice treated with Tim-3 fusion protein, as well as Tim-3-deficient mice, are refractory to tolerance induction [3,4]. Taken together, these data demonstrate the importance of the Tim-3:galectin-9 pathway in regulating inflammatory T-cell responses and states of immunological tolerance. Recently, Tim-3 expression has been demonstrated on DC and on cells of the innate immune system [5]. However, the factors that regulate Tim-3 expression in both T cells and innate immune cells have not been identified.Ã These authors contributed equally to this work. 859Recent evidence shows a correlation between mRNA expression levels of the Th1-specific transcription factor T-bet (Tbx21; T-box expressed in T cells), IFN-g and Tim-3 in T-cell clones from both patients with MS and healthy controls [6]. In this study, the linked expression of T-bet, IFN-g and Tim-3 could simply be due to the fact that T-bet is required for IFN-g transcription and IFN-gproducing Th1 cells predominantly express Tim-3 on their cell surface. Direct evidence that T-bet controls Tim-3 expression was lacking from this study. Interestingly, like Tim-3, T-bet is also expressed in DC where it controls IFN-g production [7] and is required for optimal TLR responsiveness [8]. Given that T-bet is crucial for in...

Experimental autoimmune encephalomyelitis (EAE) is a widely used model of multiple sclerosis (MS). In NOD mice, EAE develops as a relapsing-remitting disease that transitions to a chronic progressive disease, making the NOD model the only mouse model that recapitulates the full clinical disease course observed in most MS patients. We have generated a T cell receptor (TCR) transgenic mouse that expresses the alpha and beta chains of a myelin oligodendrocyte glycoprotein (MOG) 35–55-reactive TCR (1C6) on the NOD background. 1C6 TCR transgenic mice spontaneously generate both CD4+ and CD8+ T cells that recognize MOG and produce pro-inflammatory cytokines, allowing for the first time the simultaneous examination of myelin-reactive CD4+ and CD8+ T cells in the same host. 1C6 CD8+ T cells alone can induce optic neuritis and mild EAE with delayed onset; however, 1C6 CD4+ T cells alone induce severe EAE and predominate in driving disease when both cell types are present. When 1C6 mice are crossed with mice bearing an immunoglobulin heavy chain specific for MOG, the mice develop spontaneous EAE with high incidence but surprisingly the disease pattern does not resemble the Neuromyelitis optica (NMO)-like disease observed in mice bearing CD4+ T cells and B cells reactive to MOG on the C57BL/6 background. Collectively our data show that while myelin-reactive CD8+ T cells contribute to disease, disease is primarily driven by myelin-reactive CD4+ T cells and that the co-existence of myelin-reactive T and B cells does not necessarily result in a distinct pathological phenotype.