Huabao Xiong

The Tim-3 ligand galectin-9 negatively regulates T helper type 1 immunity

Tim-3 is a T helper type 1 (TH1)–specific cell surface molecule that seems to regulate TH1 responses and the induction of peripheral tolerance. However, the identity of the Tim-3 ligand and the mechanism by which this ligand inhibits the function of effector TH1 cells remain unknown. Here we show that galectin-9 is the Tim-3 ligand. Galectin-9-induced intracellular calcium flux, aggregation and death of TH1 cells were Tim-3-dependent in vitro, and administration of galectin-9 in vivo resulted in selective loss of interferon-γ-producing cells and suppression of TH1 autoimmunity. These data suggest that the Tim-3–galectin-9 pathway may have evolved to ensure effective termination of effector TH1 cells.

Toll-Like Receptors on Tumor Cells Facilitate Evasion of Immune Surveillance

The signal pathways that trigger tumor cell escape from immune surveillance are incompletely understood. Toll-like receptors (TLRs), which activate innate and adaptive immune responses, are thought to be restricted to immune cells. We show here that TLRs, including TLR4, are expressed on tumor cells from a wide variety of tissues, suggesting that TLR activation may be an important event in tumor cell immune evasion. Activation of TLR4 signaling in tumor cells by lipopolysaccharide induces the synthesis of various soluble factors and proteins including interleukin-6, inducible nitric oxide synthase, interleukin-12, B7-H1, and B7-H2, and results in resistance of tumor cells to CTL attack. In addition, lipopolysaccharide-stimulated tumor cell supernatants inhibit both T cell proliferation and natural killer cell activity. Blockade of the TLR4 pathway by either TLR4 short interfering RNA or a cell-permeable TLR4 inhibitory peptide reverses tumor-mediated suppression of T cell proliferation and natural killer cell activity in vitro, and in vivo, delays tumor growth and thus prolongs the survival of tumor-bearing mice. These findings indicate that TLR signaling results in a cascade leading to tumor evasion from immune surveillance. These novel functions of TLRs in tumor biology suggest a new class of therapeutic targets for cancer therapy.

Mucus Enhances Gut Homeostasis and Oral Tolerance by Delivering Immunoregulatory Signals

Guardian of the Gut The intestine is able to tolerate continual exposure to large amounts of commensal bacteria and foreign food antigens without triggering an inappropriate inflammatory immune response. In the large intestine, this immunological tolerance is thought to occur via a physical separation between environment and host imposed by a continuous mucous layer built up from the secreted mucin protein, MUC2. However, in the small intestine, this mucous layer is porous, necessitating an additional layer of immune control. Shan et al. (p. 447, published online 26 September; see the Perspective by Belkaid and Grainger) now report that in the small intestine, MUC2 plays an active role in immunological tolerance by activating a transcription factor in resident dendritic cells, thereby selectively blocking their ability to launch an inflammatory response. This work identifies MUC2 as a central mediator of immune tolerance to maintain homeostasis in the gut and possibly at other mucosal surfaces in the body. Mucus not only forms a physical barrier in the intestine but also promotes immunological tolerance of bacteria and foods. [Also see Perspective by Belkaid and Grainger] A dense mucus layer in the large intestine prevents inflammation by shielding the underlying epithelium from luminal bacteria and food antigens. This mucus barrier is organized around the hyperglycosylated mucin MUC2. Here we show that the small intestine has a porous mucus layer, which permitted the uptake of MUC2 by antigen-sampling dendritic cells (DCs). Glycans associated with MUC2 imprinted DCs with anti-inflammatory properties by assembling a galectin-3–Dectin-1–FcγRIIB receptor complex that activated β-catenin. This transcription factor interfered with DC expression of inflammatory but not tolerogenic cytokines by inhibiting gene transcription through nuclear factor κB. MUC2 induced additional conditioning signals in intestinal epithelial cells. Thus, mucus does not merely form a nonspecific physical barrier, but also constrains the immunogenicity of gut antigens by delivering tolerogenic signals.

The transmembrane activator TACI triggers immunoglobulin class switching by activating B cells through the adaptor MyD88

BAFF and APRIL are innate immune mediators that trigger immunoglobulin (Ig) G and IgA class switch recombination (CSR) in B cells by engaging the receptor TACI. The mechanism underlying CSR signaling by TACI remains unknown. Here, we found that the cytoplasmic domain of TACI encompasses a conserved motif that bound MyD88, an adaptor protein that activates NF-κB signaling pathways via a Toll-interleukin-1 receptor (TIR) domain. TACI lacks a TIR domain, yet triggered CSR via the DNA-editing enzyme AID by activating NF-κB through a TLR-like MyD88–IRAK-1-IRAK-4–TRAF6–TAK1 pathway. TACI-induced CSR was impaired in mice and humans lacking MyD88 or IRAK-4, indicating that MyD88 controls a B cell-intrinsic, TIR-independent, TACI-dependent pathway for Ig diversification.BAFF and APRIL are innate immune mediators that trigger immunoglobulin G (IgG) and IgA class-switch recombination (CSR) in B cells by engaging the receptor TACI. The mechanism that underlies CSR signaling by TACI remains unknown. Here we found that the cytoplasmic domain of TACI encompasses a conserved motif that bound MyD88, an adaptor that activates transcription factor NF-κB signaling pathways via a Toll–interleukin 1 (IL-1) receptor (TIR) domain. TACI lacks a TIR domain, yet triggered CSR via the DNA-editing enzyme AID by activating NF-κB through a Toll-like receptor (TLR)-like MyD88-IRAK1-IRAK4-TRAF6-TAK1 pathway. TACI-induced CSR was impaired in mice and humans lacking MyD88 or the kinase IRAK4, which indicates that MyD88 controls a B cell–intrinsic, TIR-independent, TACI-dependent pathway for immunoglobulin diversification.

TLR signaling by tumor and immune cells: a double-edged sword

The tumor cell signaling pathways that trigger the uncontrolled proliferation, resistance to apoptosis, metastasis and escape from immune surveillance are partially understood. Toll-like receptors (TLRs), which recognize a variety of pathogen-associated molecular patterns, are centrally involved in the initiation of the innate and adaptive immune responses. However, recent evidence shows that functional TLRs are also expressed on a wide variety of tumors suggesting that TLRs may play important roles in tumor biology. Activation of tumor cell TLRs not only promotes tumor cell proliferation and resistance to apoptosis, but also enhances tumor cell invasion and metastasis by regulating metalloproteinases and integrins. Moreover, the activation of TLR signaling in tumor cells induces the synthesis of proinflammatory factors and immunosuppressive molecules, which enhance the resistance of tumor cells to cytotoxic lymphocyte attack and lead to immune evasion. Thus, the neoplastic process may usurp TLR signaling pathways to advance cancer progression, which suggests that targeting tumor TLR signaling pathways may open novel therapeutic avenues.

SCF-mediated mast cell infiltration and activation exacerbate the inflammation and immunosuppression in tumor microenvironment

Despite the evidence for the role of inflammation in cancer initiation, promotion, and progression, the precise mechanism by which the inflammation within tumor is orchestrated by inflammatory cells remains to be determined. Here, we report that tumor-infiltrating mast cells remodel tumor microenvironment and promote tumor growth. Mast cell infiltration and activation in tumors were mainly mediated by tumor-derived stem cell factor (SCF) and its receptor c-Kit on mast cells. Low concentrations of SCF efficiently induced the chemotactic migration of mast cells. Tumor-infiltrating mast cells, activated by higher concentrations of SCF, expressed multiple proinflammatory factors and increased IL-17 expression in tumors. The activity of NF-kappaB and AP-1 in tumor cells was intensified in the mast cell-remodeled inflammatory microenvironment. SCF-activated mast cells also exacerbated tumor immunosuppression by releasing adenosine and increasing T regulatory cells, which augmented the suppression of T cells and natural killer cells in tumors. These findings emphasize that the remodeling of the tumor microenvironment can actually be initiated by tumor cell-released SCF and suggest that mast cells are not only a participator but also a critical regulator of inflammation and immunosuppression in the tumor microenvironment.

Interleukin 10 suppresses Th17 cytokines secreted by macrophages and T cells

IL‐17 and IL‐22 are typical cytokines produced by the Th17 T cell subset, but it is unclear if Th17 cytokines can be produced by other cell types. We demonstrate that IL‐10‐deficient and IL‐10R‐deficient macrophages stimulated with lipopolysaccharide produce high levels of IL‐17 and IL‐22. Addition of exogenous IL‐10 to IL‐10‐deficient macrophages abolished IL‐17 production. When IL‐10‐deficient and IL‐10R‐deficient splenocytes were cultured under Th17 polarizing conditions, the population of IL‐17‐producing cells was increased and the cultures produced significantly higher levels of IL‐17 and IL‐22. The addition of recombinant IL‐10 to IL‐10‐deficient splenocytes significantly decreased the percentage of IL‐17‐producing CD4+ T cells. Finally, the mRNA for the Th17 transcription factor retinoic acid‐related orphan receptor (ROR)γt was significantly elevated in IL‐10‐deficient spleen cells and macrophages. These data demonstrate that Th17 cytokines and RORγt are also expressed in macrophages and that IL‐10 negatively regulates the expression of Th17 cytokines and RORγt by both macrophages and T cells.

Listeria monocytogenes Promotes Tumor Growth via Tumor Cell Toll-Like Receptor 2 Signaling

The contribution of bacterial infection to tumorigenesis is usually ascribed to infection-associated inflammation. An alternate view is that direct interaction of bacteria with tumor cells promotes tumor progression. Here, we show that the microenvironment of large tumors favors bacterial survival, which in turn directly accelerates tumor growth by activating tumor cell Toll-like receptors (TLR). Listeria monocytogenes (Lm) survives in the microenvironment of large but not small tumors, resulting in the promotion of tumor growth. Lm did not affect the percentage of regulatory T cells or myeloid suppressor cells in the tumor. Through TLR2 signaling, Lm activated mitogen-activated protein kinases and nuclear factor-kappaB in tumor cells, resulting in the increased production of nitric oxide and interleukin-6 and increased proliferation of tumor cells. All of these effects were abrogated by silencing expression of TLR2, but not TLR4. The interaction of Helicobacter pylori with tumor cells from gastric carcinoma patients resulted in similar effects. These findings provide a new insight into infection-associated tumorigenesis and illustrate the importance of antibiotic therapy to treat tumors with bacterial infiltration.

IRF-8/Interferon (IFN) Consensus Sequence-binding Protein Is Involved in Toll-like Receptor (TLR) Signaling and Contributes to the Cross-talk between TLR and IFN-γ Signaling Pathways

Toll-like receptor (TLR) and interferon-γ (IFN-γ) signaling pathways are important for both innate and adaptive immune responses. However, the cross-talk between these two signaling pathways is incompletely understood. Here we show that IFN-γ and LPS synergistically induce the expression of proinflammatory factors, including interleukin-1 (IL-1), IL-6, IL-12, NO, and tumor necrosis factor-α (TNF-α). Comparable synergism was observed between IFN-γ and peptidoglycan (PGN; a TLR2 ligand) and poly(I:C) (a TLR3 ligand) in the induction of IL-12 promoter activity. IFN-γ enhanced lipopolysaccharide (LPS)-induced ERK and JNK phosphorylation but had no effect on LPS-induced NF-κB activation. Interestingly, we found that IRF-8–/– macrophages were impaired in the activation of LPS-induced ERK and JNK and the production of proinflammatory cytokines induced by LPS or IFN-γ plus LPS. Retroviral transduction of IRF-8 into IRF-8–/– macrophages rescued ERK and JNK activation. Furthermore, co-immunoprecipitation experiments show that IRF-8 physically interacts with TRAF6 at a binding site between amino acid residues 356 and 305 of IRF-8. Transfection of IRF-8 enhanced TRAF6 ubiquitination, which is consistent with a physical interaction of IRF-8 with TRAF6. Taken together, the results suggest that the interaction of IRF-8 with TRAF6 modulates TLR signaling and may contribute to the cross-talk between IFN-γ and TLR signal pathways.

Activation of the murine interleukin-12 p40 promoter by functional interactions between NFAT and ICSBP

Interleukin (IL)-12 is a heterodimeric cytokine that is critical for the development of a T-helper-1 immune response and immunity against intracellular pathogens. The IL-12 p40 gene product, expressed specifically in macrophages and dendritic cells, heterodimerizes with p35 to form bioactive IL-12, and heterodimerizes with p19 to comprise the cytokine IL-23. Regulation of the murine IL-12 p40 promoter is complex. Multiple cis-acting elements have been characterized that are involved in activation by bacterial products. However, molecular mechanisms through which interferon (IFN)-γ and bacterial products synergistically activate IL-12 p40 gene expression are less clear. In this study, a composite NFAT/ICSBP binding site at –68 to –54 is identified that is functionally important for p40 promoter activation by lipopolysaccharide (LPS) and LPS plus IFN-γ. DNA binding of NFAT and ICSBP is demonstrated on the endogenous promoter by chromatin immunoprecipitation. NFAT is required for ICSBP binding to this region. Overexpression of NFAT and ICSBP synergistically activates the p40 promoter. A dominant negative NFAT molecule attenuates LPS- and IFN-γ-activated endogenous IL-12 p40 mRNA expression. A physical association between NFAT and ICSBP in the absence of DNA is detected by co-immunoprecipitation of endogenous proteins. Three NFAT domains are required for ICSBP interaction. Finally, in LPS- and IFN-γ-activated RAW-264.7 cells, the association between NFAT and ICSBP is abrogated by IL-10 priming.