Rosana Gonzalez-Quintial

Commitment to the Regulatory T Cell Lineage Requires CARMA1 in the Thymus but Not in the Periphery

Regulatory T (Treg) cells expressing forkhead box P3 (Foxp3) arise during thymic selection among thymocytes with modestly self-reactive T cell receptors. In vitro studies suggest Foxp3 can also be induced among peripheral CD4+ T cells in a cytokine dependent manner. Treg cells of thymic or peripheral origin may serve different functions in vivo, but both populations are phenotypically indistinguishable in wild-type mice. Here we show that mice with a Carma1 point mutation lack thymic CD4+Foxp3+ Treg cells and demonstrate a cell-intrinsic requirement for CARMA1 in thymic Foxp3 induction. However, peripheral Carma1-deficient Treg cells could be generated and expanded in vitro in response to the cytokines transforming growth factor beta (TGFβ) and interleukin-2 (IL-2). In vivo, a small peripheral Treg pool existed that was enriched at mucosal sites and could expand systemically after infection with mouse cytomegalovirus (MCMV). Our data provide genetic evidence for two distinct mechanisms controlling regulatory T cell lineage commitment. Furthermore, we show that peripheral Treg cells are a dynamic population that may expand to limit immunopathology or promote chronic infection.

Sensors of the innate immune system: their mode of action

The discovery of molecular sensors that enable eukaryotes to recognize microbial pathogens and their products has been a key advance in our understanding of innate immunity. A tripartite sensing apparatus has developed to detect danger signals from infectious agents and damaged tissues, resulting in an immediate but short-lived defense response. This apparatus includes Toll-like receptors, retinoid acid-inducible gene-I-like receptors and other cytosolic nucleic acid sensors, and nucleotide-binding and oligomerization domain-like receptors; adaptors, kinases and other signaling molecules are required to elicit effective responses. Although this sensing is beneficial to the host, excessive activation and/or engagement by self molecules might induce autoimmune and other inflammatory disorders.

Essential requirement for IRF8 and SLC15A4 implicates plasmacytoid dendritic cells in the pathogenesis of lupus

In vitro evidence suggests that plasmacytoid dendritic cells (pDCs) are intimately involved in the pathogenesis of lupus. However, it remains to be determined whether these cells are required in vivo for disease development, and whether their contribution is restricted to hyperproduction of type I IFNs. To address these issues, we created lupus-predisposed mice lacking the IFN regulatory factor 8 (IRF8) or carrying a mutation that impairs the peptide/histidine transporter solute carrier family 15, member 4 (SLC15A4). IRF8-deficient NZB mice, lacking pDCs, showed almost complete absence of anti-nuclear, anti-chromatin, and anti-erythrocyte autoantibodies, along with reduced kidney disease. These effects were observed despite normal B-cell responses to Toll-like receptor (TLR) 7 and TLR9 stimuli and intact humoral responses to conventional T-dependent and -independent antigens. Moreover, Slc15a4 mutant C57BL/6-Faslpr mice, in which pDCs are present but unable to produce type I IFNs in response to endosomal TLR ligands, also showed an absence of autoantibodies, reduced lymphadenopathy and splenomegaly, and extended survival. Taken together, our results demonstrate that pDCs and the production of type I IFNs by these cells are critical contributors to the pathogenesis of lupus-like autoimmunity in these models. Thus, IRF8 and SLC15A4 may provide important targets for therapeutic intervention in human lupus.

Sensors of the innate immune system: their link to rheumatic diseases

Evidence strongly suggests that excessive or protracted signaling, or both, by cell-surface or intracellular innate immune receptors is central to the pathogenesis of most autoimmune and autoinflammatory rheumatic diseases. The initiation of aberrant innate and adaptive immune responses in autoimmune diseases can be triggered by microbes and, at times, by endogenous molecules—particularly nucleic acids and related immune complexes—under sterile conditions. By contrast, most autoinflammatory syndromes are generally dependent on germline or de novo gene mutations that cause or facilitate inflammasome assembly. The consequent production of proinflammatory cytokines, principally interferon-α/β and tumor necrosis factor in autoimmune diseases, and interleukin-1β in autoinflammatory diseases, leads to the creation of autoamplification feedback loops and chronicity of these syndromes. These findings have resulted in a critical reappraisal of pathogenetic mechanisms, and provide a basis for the development of novel diagnostic and therapeutic modalities for these diseases.

γδ T Cell Homeostasis Is Controlled by IL-7 and IL-15 Together with Subset-Specific Factors

Among T cell subsets, γδ T cells uniquely display an Ag receptor-based tissue distribution, but what defines their preferential homing and homeostasis is unknown. To address this question, we studied the resources that control γδ T cell homeostasis in secondary lymphoid organs. We found that γδ and αβ T cells are controlled by partially overlapping resources, because acute homeostatic proliferation of γδ T cells was inhibited by an intact αβ T cell compartment, and both populations were dependent on IL-7 and IL-15. Significantly, to undergo acute homeostatic proliferation, γδ T cells also required their own depletion. Thus, γδ T cell homeostasis is maintained by trophic cytokines commonly used by other types of lymphoid cells, as well as by additional, as yet unidentified, γδ-specific factors.

Anti–IFN-α/β Receptor Antibody Treatment Ameliorates Disease in Lupus-Predisposed Mice

The demonstration in humans and mice that nucleic acid-sensing TLRs and type I IFNs are essential disease mediators is a milestone in delineating the mechanisms of lupus pathogenesis. In this study, we show that Ifnb gene deletion does not modify disease progression in NZB mice, thereby strongly implicating IFN-α subtypes as the principal pathogenic effectors. We further document that long-term treatment of male BXSB mice with an anti–IFN-α/β receptor Ab of mouse origin reduced serologic, cellular, and histologic disease manifestations and extended survival, suggesting that disease acceleration by the Tlr7 gene duplication in this model is mediated by type I IFN signaling. The efficacy of this treatment in BXSB mice was clearly evident when applied early in the disease process, but only partial reductions in some disease characteristics were observed when treatment was initiated at later stages. A transient therapeutic effect was also noted in the MRL-Faslpr model, although overall mortality was unaffected. The combined findings suggest that IFN-α/β receptor blockade, particularly when started at early disease stages, may be a useful treatment approach for human systemic lupus erythematosus and other autoimmune syndromes.

Identification of clonally expanded T cells in rheumatoid arthritis using a sequence enrichment nuclease assay.

Identification of expanded clones engaged in immune and autoimmune responses is still imperfect, since they are often diluted by irrelevant cells expressing diverse specificities. To efficiently characterize T cell receptors expressed by clonally expanded lymphocytes in rheumatoid arthritis (RA) and other inflammatory conditions, we developed an assay system, termed sequence enrichment nuclease assay (SENA). Key elements of SENA are the efficiency of heat-denatured DNA strand reassociation, which increases exponentially with concentration, and the elimination of unhybridized sequences by single-strand-specific DNase. T cell clonal expansions were identified primarily in synovial fluids, but also in peripheral blood of RA patients. Synovial fluids had more prominent expansions in the CD8 than the CD4 subset, whereas clonal expansions in the CD4 subset predominated among peripheral blood lymphocytes. Dominant clones exhibited diverse sequences with no clear conservation of junctional motifs, although the same amino acid sequence was identified in two patients. In most instances, dominant clones in the blood were discordant to those in the corresponding synovial fluid, suggesting local stimulation or preferential sequestration of T cells displaying particular specifities.

Systemic Autoimmunity and Lymphoproliferation Are Associated with Excess IL-7 and Inhibited by IL-7Rα Blockade

Lupus is characterized by disturbances in lymphocyte homeostasis, as demonstrated by the marked accumulation of activated/memory T cells. Here, we provide evidence that proliferation of the CD8+ precursors for the accumulating CD4–CD8– T cells in MRL-Faslpr lupus-predisposed mice is, in part, driven by commensal antigens. The ensuing lymphadenopathy is associated with increased production of IL-7 due to expansion of fibroblastic reticular cells, the primary source of this cytokine. The excess IL-7 is not, however, consumed by CD4–CD8– T cells due to permanent down-regulation of IL-7Rα (CD127), but instead supports proliferation of autoreactive T cells and progression of autoimmunity. Accordingly, IL-7R blockade reduced T cell activation and autoimmune manifestations even when applied at advanced disease stage. These findings indicate that an imbalance favoring production over consumption of IL-7 may contribute to systemic autoimmunity, and correction of this imbalance may be a novel therapeutic approach in lymphoproliferative and autoimmune syndromes.

Type I interferon is a therapeutic target for virus-induced lethal vascular damage

Significance Lassa virus is, after dengue virus, the second most common cause of viral hemorrhagic fever. In susceptible individuals, Lassa virus infection is associated with vascular permeability, leading to tissue edema, organ failure, and death. Hemorrhagic fever viruses efficiently infect vascular endothelial cells, but are generally considered noncytopathic. Thus, the mechanism of virus-induced vascular injury remains unclear. Using the lymphocytic choriomeningitis virus variant clone 13, a prototype of Lassa virus, we show here that lethal vascular leakage in susceptible mice was completely prevented by type I IFN receptor blockade. Therefore, approaches that target type I IFNs or effector molecules induced by these cytokines may be considered for the treatment of Lassa fever and other severe hemorrhagic viral illnesses. The outcome of a viral infection reflects the balance between virus virulence and host susceptibility. The clone 13 (Cl13) variant of lymphocytic choriomeningitis virus—a prototype of Old World arenaviruses closely related to Lassa fever virus—elicits in C57BL/6 and BALB/c mice abundant negative immunoregulatory molecules, associated with T-cell exhaustion, negligible T-cell–mediated injury, and high virus titers that persist. Conversely, here we report that in NZB mice, despite the efficient induction of immunoregulatory molecules and high viremia, Cl13 generated a robust cytotoxic T-cell response, resulting in thrombocytopenia, pulmonary endothelial cell loss, vascular leakage, and death within 6–8 d. These pathogenic events required type I IFN (IFN-I) signaling on nonhematopoietic cells and were completely abrogated by IFN-I receptor blockade. Thus, IFN-I may play a prominent role in hemorrhagic fevers and other acute virus infections associated with severe vascular pathology, and targeting IFN-I or downstream effector molecules may be an effective therapeutic approach.

Tumor immunity via homeostatic T cell proliferation: mechanistic aspects and clinical perspectives.

Efforts to develop effective anti-tumor immunotherapies are hampered by the difficulty of overcoming tolerance against tumor antigens, which in most instances are normal gene products that are over-expressed, preferentially expressed or re-expressed in cancer cells. Considering that lymphopenia-induced homeostatic T cell proliferation is mediated by self-peptide/MHC recognition and that the expanded cells acquire some effector functions, we hypothesized that this process could be used to break tolerance against tumor antigens. Studies by us and others in several mouse models demonstrated that availability of tumor antigens during homeostatic T cell proliferation indeed leads to effective anti-tumor autoimmunity with specificity and memory. This effect appears to be mediated by reduction in the activation threshold of low-affinity tumor-specific T cells, leading to their preferential engagement and expansion. In its simplicity, this approach is likely to have application in humans, since it relies on conventional lymphopenia-inducing cancer therapies, infusion of autologous lymphocytes and, optimally, tumor-specific vaccination.