Jane Tian

Toll-like receptor 9–dependent activation by DNA-containing immune complexes is mediated by HMGB1 and RAGE

Increased concentrations of DNA-containing immune complexes in the serum are associated with systemic autoimmune diseases such as lupus. Stimulation of Toll-like receptor 9 (TLR9) by DNA is important in the activation of plasmacytoid dendritic cells and B cells. Here we show that HMGB1, a nuclear DNA-binding protein released from necrotic cells, was an essential component of DNA-containing immune complexes that stimulated cytokine production through a TLR9–MyD88 pathway involving the multivalent receptor RAGE. Moreover, binding of HMGB1 to class A CpG oligodeoxynucleotides considerably augmented cytokine production by means of TLR9 and RAGE. Our data demonstrate a mechanism by which HMGB1 and RAGE activate plasmacytoid dendritic cells and B cells in response to DNA and contribute to autoimmune pathogenesis.

The CD28-Related Molecule ICOS Is Required for Effective T Cell-Dependent Immune Responses

While CD28 is critical for expansion of naive T cells, recent evidence suggests that the activation of effector T cells is largely independent of CD28/B7. We suggest that ICOS, the third member of the CD28/CTLA-4 family, plays an important role in production of IL-2, IL-4, IL-5, and IFNgamma from recently activated T cells and contributes to T cell-dependent B help in vivo. Inhibition of ICOS attenuates lung mucosal inflammation induced by Th2 but not Th1 effector populations. Our data indicate a critical function for the third member of the CD28 family in T cell-dependent immune responses.

Tim-3 inhibits T helper type 1-mediated auto- and alloimmune responses and promotes immunological tolerance.

Although T helper (TH) cell–mediated immunity is required to effectively eliminate pathogens, unrestrained TH activity also contributes to tissue injury in many inflammatory and autoimmune diseases. We report here that the TH type 1 (TH1)-specific Tim-3 (T cell immunoglobulin domain, mucin domain) protein functions to inhibit aggressive TH1-mediated auto- and alloimmune responses. Tim-3 pathway blockade accelerated diabetes in nonobese diabetic mice and prevented acquisition of transplantation tolerance induced by costimulation blockade. These effects were mediated, at least in part, by dampening of the antigen-specific immunosuppressive function of CD4+CD25+ regulatory T cell populations. Our data indicate that the Tim-3 pathway provides an important mechanism to down-regulate TH1-dependent immune responses and to facilitate the development of immunological tolerance.

ICOS is critical for T helper cell–mediated lung mucosal inflammatory responses

We examined the requirement for and cooperation between CD28 and inducible costimulator (ICOS) in effective T helper (TH) cell responses in vivo. We found that both CD28 and ICOS were critical in determining the outcome of an immune response; cytolytic T lymphocyte–associated antigen 4–immunoglobulin (CTLA-4–Ig), ICOS-Ig and/or a neutralizing ICOS monoclonal antibody attenuated T cell expansion, TH2 cytokine production and eosinophilic inflammation. CD28-dependent signaling was essential during priming, whereas ICOS–B7RP-1 regulated TH effector responses, and the up-regulation of chemokine receptors that determine T cell migration. Our data suggests a scenario whereby both molecules regulate the outcome of the immune response but play separate key roles: CD28 primes T cells and ICOS regulates effector responses.

Mature T Cells Depend on Signaling through the IKK Complex

The transcription factor NF-kappaB is implicated in various aspects of T cell development and function. The IkappaB kinase (IKK) complex, consisting of two kinases, IKK1/alpha and IKK2/beta, and the NEMO/IKKgamma regulatory subunit, mediates NF-kappaB activation by most known stimuli. Adoptive transfer experiments had demonstrated that IKK1 and IKK2 are dispensable for T cell development. We show here that T lineage-specific deletion of IKK2 allows survival of naive peripheral T cells but interferes with the generation of regulatory and memory T cells. T cell-specific ablation of NEMO or replacement of IKK2 with a kinase-dead mutant prevent development of peripheral T cells altogether. Thus, IKK-induced NF-kappaB activation, mediated by either IKK1 or IKK2, is essential for the generation and survival of mature T cells, and IKK2 has an additional role in regulatory and memory T cell development.

Noncanonical Autophagy Is Required for Type I Interferon Secretion in Response to DNA-Immune Complexes

Toll-like receptor-9 (TLR9) is largely responsible for discriminating self from pathogenic DNA. However, association of host DNA with autoantibodies activates TLR9, inducing the pathogenic secretion of type I interferons (IFNs) from plasmacytoid dendritic cells (pDCs). Here, we found that in response to DNA-containing immune complexes (DNA-IC), but not to soluble ligands, IFN-α production depended upon the convergence of the phagocytic and autophagic pathways, a process called microtubule-associated protein 1A/1B-light chain 3 (LC3)-associated phagocytosis (LAP). LAP was required for TLR9 trafficking into a specialized interferon signaling compartment by a mechanism that involved autophagy-related proteins, but not the conventional autophagic preinitiation complex, or adaptor protein-3 (AP-3). Our findings unveil a new role for nonconventional autophagy in inflammation and provide one mechanism by which anti-DNA autoantibodies, such as those found in several autoimmune disorders, bypass the controls that normally restrict the apportionment of pathogenic DNA and TLR9 to the interferon signaling compartment.

RAGE is a nucleic acid receptor that promotes inflammatory responses to DNA

Receptor for advanced glycation end-products (RAGE) detects nucleic acids and promotes DNA uptake into endosomes, which in turn lowers the immune recognition threshold for TLR9 activation.

Participation of Rip2 in lipopolysaccharide signaling is independent of its kinase activity.

Rip2 (Rick, Cardiak, CCK2, and CARD3) is a serine/threonine kinase containing a caspase recruitment domain (CARD) at the C terminus. Previous reports have shown that Rip2 is involved in multiple receptor signaling pathways that are important for innate and adaptive immune responses. However, it is not known whether Rip2 kinase activity is required for its function. Here we confirm that Rip2 participates in lipopolysaccharide (LPS)/Toll-like receptor (TLR4) signaling and demonstrate that its kinase activity is not required. Upon LPS stimulation, Rip2 was transiently recruited to the TLR4 receptor complex and associated with key TLR signaling mediators IRAK1 and TRAF6. Furthermore, Rip2 kinase activity was induced by LPS treatment. These data indicate that Rip2 is directly involved in the LPS/TLR4 signaling. Whereas macrophages from Rip2-deficient mice showed impaired NF-κB and p38 mitogen-activated protein kinase activation and reduced cytokine production in response to LPS stimulation, LPS signaling was intact in macrophages from mice that express Rip2 kinase-dead mutant. These results demonstrate that Rip2-mediated LPS signaling is independent of its kinase activity. Our findings strongly suggest that Rip2 functions as an adaptor molecule in transducing signals from immune receptors.

Resolution of Bronchial Hyperresponsiveness and Pulmonary Inflammation Is Associated with IL-3 and Tissue Leukocyte Apoptosis

We have used two models of murine pulmonary inflammation to investigate the signals responsible for the resolution of bronchial hyperresponsiveness (BHR). Both protocols involved two sensitizations with OVA followed by serial aerosolized challenge with OVA. We determined that administration of the second sensitization by aerosol (model A) was associated with a transient response, whereas administration by the i.p. route (model B) induced a sustained response, in the form of BHR and eosinophilia. This difference in kinetics was due solely to the route of the second Ag administration and was not associated with Ag dose or adjuvant. Differences in kinetics of lung eosinophilia/BHR were shown to be independent of IgE levels and IL-4 or IL-5. However, IL-3 levels in model A closely correlated with the rate of leukocyte clearance by apoptosis and were observed concomitant with a decline in BHR. Blockage of IL-3 in model B increased leukocyte apoptosis but reduced tissue eosinophilia and BHR. The use of mouse models in which a single different administration of allergen is associated with a failure/success to resolve inflammation and BHR by 72 h postchallenge indicates a link between IL-3 production, leukocyte apoptosis, and BHR responses.

Cell Type-Specific Recognition of Human Metapneumoviruses (HMPVs) by Retinoic Acid-Inducible Gene I (RIG-I) and TLR7 and Viral Interference of RIG-I Ligand Recognition by HMPV-B1 Phosphoprotein

Human metapneumoviruses (HMPVs) are recently identified Paramyxoviridae that contribute to respiratory tract infections in children. No effective treatments or vaccines are available. Successful defense against virus infection relies on early detection by germ line-encoded pattern recognition receptors and activation of cytokine and type I IFN genes. Recently, the RNA helicase retinoic acid-inducible gene I (RIG-I) has been shown to sense HMPV. In this study, we investigated the abilities of two prototype strains of HMPV (A1 [NL\1\00] and B1 [NL\1\99]) to activate RIG-I and induce type I IFNs. Despite the abilities of both HMPV-A1 and HMPV-B1 to infect and replicate in cell lines and primary cells, only the HMPV-A1 strain triggered RIG-I to induce IFNA/B gene transcription. The failure of the HMPV-B1 strain to elicit type I IFN production was dependent on the B1 phosphoprotein, which specifically prevented RIG-I–mediated sensing of HMPV viral 5′ triphosphate RNA. In contrast to most cell types, plasmacytoid dendritic cells displayed a unique ability to sense both HMPV-A1 and HMPV-B1 and in this case sensing was via TLR7 rather than RIG-I. Collectively, these data reveal differential mechanisms of sensing for two closely related viruses, which operate in cell type‑specific manners.