Antigen receptor engagement on T lymphocytes activates transcription factors important for stimulating cytokine gene expression. This is critical for clonal expansion of antigen-specific T cells and propagation of immune responses. Additionally, under some conditions antigen receptor stimulation initiates apoptosis of T lymphocytes through the induced expression of CD95 ligand and its receptor. Here we demonstrate that the transcription factor, NFAT, which is critical for the inducible expression of many cytokine genes, also plays a critical role in the regulation of T cell receptor-mediated CD95 ligand expression. Two sites within the CD95 ligand promoter, identified through DNase I footprinting, bind NFAT proteins from nuclear extracts of activated T cells. Although both sites appear important for optimal expression of CD95 ligand in activated T cells, mutational analysis suggests that the distal NFAT site plays a more significant role. Furthermore, these sites do not appear to be required for constitutive CD95 ligand expression in Sertoli cells.Adaptive immune responses require the activation of T lymphocytes through antigen-specific T cell receptor (TCR) 1 stimulation. Signaling events initiated by TCR ligation lead to the activation of transcription factors that regulate expression of cytokine genes, such as IL-2. This is important for the clonal expansion of antigen-specific T cells and propagation of immune responses (1, 2). However, once the antigenic stimulus has been cleared, the expanded population of cells must be eliminated to prevent accumulation of excessive lymphocytes (2, 3). Recently, it has been proposed that one mechanism by which this occurs is through the induced expression of CD95 ligand (4 -7). Once expressed, CD95 ligand engages its receptor, CD95, also expressed on the population of activated lymphocytes (8). In the absence of costimulatory signals that can delay apoptosis (9 -12), CD95 ligation rapidly initiates the programmed cell death machinery thus efficiently eliminating excessive activated lymphocytes (8). Additionally, autoreactive T cells that are inappropriately activated in the periphery are believed to undergo apoptosis through a similar process (13).The significance of CD95 ligand expression in the process of activation-induced cell death has been highlighted by recent studies that demonstrate that blocking CD95/CD95 ligand interactions prevents apoptosis of TCR-stimulated lymphocytes (5, 7). Interestingly, in addition to its inducible expression on activated lymphocytes, CD95 ligand is constitutively expressed on epithelial cells within the eye and Sertoli cells within the testes (14 -16). Constitutive CD95 ligand expression participates in maintenance of the "immune privileged" status of these tissues by inducing apoptosis in infiltrating, CD95-bearing, activated lymphocytes (17). Despite improved understanding of the important physiological roles for CD95 ligand in immune privileged sites and in controlling T cell homeostasis, little is yet known about the regulation of ...

Dendritic cells (DC) have a critical effect on the outcome of adaptive immune responses against growing tumors. Whereas it is generally assumed that the presence of phenotypically mature DCs should promote protective antitumor immunity, evidence to the contrary does exist. We describe here a novel mechanism by which tumor-infiltrating dendritic cells (TIDC) actively contribute to the suppression of protective CD8

BackgroundThe constellation of human inflammatory bowel disease (IBD) includes ulcerative colitis and Crohn's disease, which both display a wide spectrum in the severity of pathology. One theory is that multiple genetic hits to the host immune system may contribute to the susceptibility and severity of IBD. However, experimental proof of this concept is still lacking. Several genetic mouse models that each recapitulate some aspects of human IBD have utilized a single gene defect to induce colitis. However, none have produced pathology clearly distinguishable as either ulcerative colitis or Crohn's disease, in part because none of them reproduce the most severe forms of disease that are observed in human patients. This lack of severe IBD models has posed a challenge for research into pathogenic mechanisms and development of new treatments. We hypothesized that multiple genetic hits to the regulatory machinery that normally inhibits immune activation in the intestine would generate more severe, reproducible pathology that would mimic either ulcerative colitis or Crohn's disease.Methods and FindingsWe generated a novel mouse line (dnKO) that possessed defects in both TGFβRII and IL-10R2 signaling. These mice rapidly and reproducibly developed a disease resembling fulminant human ulcerative colitis that was quite distinct from the much longer and more variable course of pathology observed previously in mice possessing only single defects. Pathogenesis was driven by uncontrolled production of proinflammatory cytokines resulting in large part from T cell activation. The disease process could be significantly ameliorated by administration of antibodies against IFNγ and TNFα and was completely inhibited by a combination of broad-spectrum antibiotics.ConclusionsHere, we develop to our knowledge the first mouse model of fulminant ulcerative colitis by combining multiple genetic hits in immune regulation and demonstrate that the resulting disease is sensitive to both anticytokine therapy and broad-spectrum antibiotics. These findings indicated the IL-10 and TGFβ pathways synergize to inhibit microbially induced production of proinflammatory cytokines, including IFNγ and TNFα, which are known to play a role in the pathogenesis of human ulcerative colitis. Our findings also provide evidence that broad-spectrum antibiotics may have an application in the treatment of patients with ulcerative colitis. This model system will be useful in the future to explore the microbial factors that induce immune activation and characterize how these interactions produce disease.

Obesity is a mounting health concern in the United States, and is associated with an increased risk for developing several cancers, including renal cell carcinoma (RCC). Despite this, little is known regarding the impact of obesity on antitumor immunity. As dendritic cells (DC) are critical regulators of antitumor immunity, we examined the combined effects of obesity and tumor outgrowth on DC function. Using a diet-induced obesity (DIO) model, DC function was evaluated in mice bearing orthotopic RCC and in tumor-free controls. Tumor-free DIO mice had profoundly altered serum cytokine and chemokine profiles, with upregulation of 15 proteins, including IL-1α, IL-17, and LIF. Tumor-free DIO mice had elevated percentages of conventional splenic DC that were impaired in their ability to stimulate naive T cell expansion, although they were phenotypically similar to normal weight (NW) controls. In DIO mice, intra-renal RCC tumor challenge in the absence of therapy led to increased local infiltration by T cell-suppressive DC and accelerated early tumor outgrowth. Following administration of a DC-dependent immunotherapy, established RCC tumors regressed in NW mice. The same immunotherapy was ineffective in DIO mice, and was characterized by an accumulation of regulatory DC in tumor-bearing kidneys, decreased local infiltration by IFNγ-producing CD8 T cells, and progressive tumor outgrowth. Our results suggest that the presence of obesity as a co-morbidity can impair the efficacy of DC-dependent antitumor immunotherapies.

Glioblastoma (GBM) is a lethal disease with no effective therapies available. We previously observed upregulation of the TAM (Tyro-3, Axl, and Mer) receptor tyrosine kinase family member AXL in mesenchymal GBM and showed that knockdown of AXL induced apoptosis of mesenchymal, but not proneural, glioma sphere cultures (GSC). In this study, we report that BGB324, a novel small molecule inhibitor of AXL, prolongs the survival of immunocompromised mice bearing GSC-derived mesenchymal GBM-like tumors. We show that protein S (PROS1), a known ligand of other TAM receptors, was secreted by tumor-associated macrophages/microglia and subsequently physically associated with and activated AXL in mesenchymal GSC. PROS1-driven phosphorylation of AXL (pAXL) induced NFκB activation in mesenchymal GSC, which was inhibited by BGB324 treatment. We also found that treatment of GSC-derived mouse GBM tumors with nivolumab, a blocking antibody against the immune checkpoint protein PD-1, increased intratumoral macrophages/microglia and activation of AXL. Combinatorial therapy with nivolumab plus BGB324 effectively prolonged the survival of mice bearing GBM tumors. Clinically, expression of AXL or PROS1 was associated with poor prognosis for patients with GBM. Our results suggest that the PROS1-AXL pathway regulates intrinsic mesenchymal signaling and the extrinsic immune microenvironment, contributing to the growth of aggressive GBM tumors. These findings suggest that development of combination treatments of AXL and immune checkpoint inhibitors may provide benefit to patients with GBM. .