Shyr-Te Ju

A Major Lung CD103 (αE)-β7 Integrin-Positive Epithelial Dendritic Cell Population Expressing Langerin and Tight Junction Proteins

Dendritic cells (DC) mediate airway Ag presentation and play key roles in asthma and infections. Although DC subsets are known to perform different functions, their occurrence in mouse lungs has not been clearly defined. In this study, three major lung DC populations have been found. Two of them are the myeloid and plasmacytoid DC (PDC) well-characterized in other lymphoid organs. The third and largest DC population is the integrin αE (CD103) β7-positive and I-AhighCD11chigh-DC population. This population was found to reside in the lung mucosa and the vascular wall, express a wide variety of adhesion and costimulation molecules, endocytose avidly, present Ag efficiently, and produce IL-12. Integrin αEβ7+ DC (αE-DC) were distinct from intraepithelial lymphocytes and distinguishable from CD11bhigh myeloid and mPDCA-1+B220+Gr-1+ PDC populations in surface marker phenotype, cellular functions, and tissue localization. Importantly, this epithelial DC population expressed high levels of the Langerhans cell marker Langerin and the tight junction proteins Claudin-1, Claudin-7, and ZO-2. In mice with induced airway hyperresponsiveness and eosinophilia, αE-DC numbers were increased in lungs, and their costimulation and adhesion molecules were up-regulated. These studies show that αE-DC is a major and distinct lung DC population and a prime candidate APC with the requisite surface proteins for migrating across the airway epithelia for Ag and pathogen capture, transport, and presentation. They exhibit an activated phenotype in allergen-induced lung inflammation and may play significant roles in asthma pathogenesis.

Regulatory T Cells Suppress Innate Immunity in Kidney Ischemia-Reperfusion Injury

Both innate and adaptive mechanisms participate in the pathogenesis of kidney ischemia-reperfusion injury (IRI), but the role of regulatory immune mechanisms is unknown. We hypothesized that the anti-inflammatory effects of CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) protect against renal IRI. Partial depletion of Tregs with an anti-CD25 mAb potentiated kidney damage induced by IRI. Reducing the number of Tregs resulted in more neutrophils, macrophages, and innate cytokine transcription in the kidney after IRI but did not affect CD4(+) T cells or B cells. We performed adoptive transfer of lymph node cells from wild-type mice or FoxP3-deficient Scurfy mice into T cell- and B cell-deficient RAG-1 knockout mice to generate mice with and without FoxP3(+) Tregs, respectively. FoxP3(+) Treg-deficient mice accumulated a greater number of inflammatory leukocytes after renal IRI than mice containing Tregs. To confirm that a lack of Tregs potentiated renal injury, we co-transferred isolated Tregs and Scurfy lymph node cells; Treg repletion significantly attenuated IRI-induced renal injury and leukocyte accumulation. Furthermore, although adoptive transfer of wild-type Tregs into RAG-1 knockout mice was sufficient to prevent kidney IRI, transfer of IL-10-deficient Tregs was not. Taken together, these results demonstrate that Tregs modulate injury after kidney IRI through IL-10-mediated suppression of the innate immune system.

Fas Ligand Engagement of Resident Peritoneal Macrophages In Vivo Induces Apoptosis and the Production of Neutrophil Chemotactic Factors

Fas ligand (FasL) is a potent proapoptotic type-II transmembrane protein that can cause cell death in Fas+ target populations. Despite the presumed “silent” nature of apoptotic cell death, forced expression of FasL can induce a dramatic inflammatory response. To elucidate the in vivo mechanism(s) linking FasL and inflammation, we used a membrane-bound cell-free form of FasL (mFasL-vesicle preparation (VP)). We found that i.p. injection of FasL-microvesicles led to the rapid activation and subsequent demise of Mac1high resident peritoneal macrophages. Apoptosis of Mac1high peritoneal macrophages was observed within 0.5 h of mFasL-VP injection and correlated with the detection of increased macrophage inflammatory protein (MIP)-2 levels in peritoneal lavage fluid as well as induced RNA expression of IL-1β, MIP-2, MIP-1α, and MIP-1β. In vitro culture of purified peritoneal populations identified Mac1high cells as the major cytokine/chemokine producers in response to mFasL-VP. Purified Mac1high cells exposed to FasL could restore the ability of Fas-deficient mice to mount an inflammatory response. Our data demonstrate that the FasL-mediated inflammatory response starts with the production of proinflammatory mediators by preapoptotic resident tissue macrophages and suggest a general mechanism responsible for neutrophil inflammation seen in cases of FasL-expressing allografts.

FasL promoter activation by IL-2 through SP1 and NFAT but not Egr-2 and Egr-3

Recently activated peripheral T cells treated with IL‐2 for 4 days expressed Fas ligand (FasL)‐mediated cytotoxicity. These IL‐2‐treated T cells had high nuclear expression of SP1 and NFAT, but lacked the Egr‐2 and Egr‐3 that could be induced by anti‐CD3 stimulation and had been implicated in FasL gene activation. A minimal promoter region that responded to IL‐2 was identified by transient transfection assays using deletion mutants. The data suggests that the GGGCGGAAA site present in the 5′ end of the minimal FasL promoter is critical to IL‐2‐induced FasL gene activation. The GGGCGGAAA sequence contains an overlapping site used by two transcription factor families, one (GGGCGG) for the SP1 family and the other (GGAAA) for the NFAT family. FasL promoter activity was partially but statistically significantly reduced with constructs mutated at either site. More activity was lost with a construct mutated at both sites. In contrast, mutation at the Egr site had no effect on IL‐2‐induced FasL promoter activity. Our study identified a new FasL promoter site responding to IL‐2‐induced SP1 and NFAT factors. Furthermore, the nuclei of IL‐2‐treated cells express SP1 and NFAT, but not Egr‐2 and Egr‐3, for FasL gene activation.

Molecular and Cellular Mechanisms RegulatingTand B Cell Apoptosis through Fas/FasL Interaction

Fas (CD95) and Fas ligand (FasL) are a receptor/ligand pair critically involved in lymphocyte homeostasis and peripheral tolerance such that genetic defect in either Fas or FasL results in an autoimmune lymphoproliferative syndrome. Fas is a type I transmembrane protein and a member of the tumor necrosis factor receptor (TNFR) family whereas FasL is a type II transmembrane protein and a member of TNF family. Binding of Fas by FasL induces apoptosis of the Fas-expressing cells. In the past few years, Fas/FasL interaction has been connected to a series of important phenomena previously viewed as independent immune processes. The activation-induced T cell death (AICD) and the FasL-mediated cytotoxicity by activated T cells are two critical mechanisms that can account for most of these phenomena. It is in the context of the two mechanisms that we discuss in this review the molecular and cellular events that occur during T/T and T/B interactions that account for the down-regulation of the immune response. We have also discussed recent advances in the areas of FasL gene regulation, lymphokine regulation of AICD, and regulation of B cell susceptibility to FasL. Investigation in these areas should help elucidate the role of Fas/FasL in the complex network of regulatory mechanisms that control immune response and autoimmunity.

IL-2: A Two-Faced Master Regulator of Autoimmunity

CD4(+) T-cell (Th) cytokines provide important regulatory and effector functions of T-cells. Among them, IL-2 plays a unique role. IL-2 is required for the generation and maintenance of regulatory T-cells (Treg) to provide lifelong protection from autoimmune disease. Whether IL-2 is also required for autoimmune disease development is less clear as Il2(-/)(-) mice themselves spontaneously develop multi-organ inflammation (MOI). In this communication, we discuss evidence that support the thesis that IL-2 is required for the development of autoimmune response, although some aspects of autoimmune response are not regulated by IL-2. Potential IL-2-dependent mechanisms operating at specific stages of the inflammation process are presented. The interplays among Treg, IL-2, autoimmune response and adaptive immunity are discussed. Overall, available information indicates that IL-2 is a two-faced master regulator of autoimmunity: one to prevent autoimmunity while the other promotes autoimmune response. The latter is an unfortunate consequence of IL-2 function that is used to promote the adaptive immune response against foreign antigens and pathogens.

Functional analysis of macrophage hybridomas: II. Antibody-independent tumor cytotoxicity and its dissociation from IL-1 production and Ia expression☆

Cloned macrophage hybridoma cells derived from fusions between splenic adherent cells of CKB mice and the drug-marked P388D1 cell line were studied to further understand macrophage-mediated tumoricidal activity at the clonal level. Most of the macrophage hybridomas were specifically activated by a combination of lymphokine and lipopolysaccharide (LPS), but were not activated by either agent alone. There is no apparent correlation between macrophage-mediated tumor cytotoxicity and the ability of LPS to induce interleukin 1 (IL-1) secretion or of concanavalin A (Con A) supernatants containing interferon-gamma to increase H-2 and Ia expression. Thus, one hybridoma clone which was unable to kill tumor cells was still sensitive to LPS and lymphokine activation by the latter criterion.

Role of Activator Protein-1 in TCR-Mediated Regulation of the Murine fasl Promoter

The present study demonstrates that transcription factor interactions are important in regulating the murine fasl promoter following TCR-mediated activation. We used DNase I-footprinting, EMSAs, and transient transfection assays to identify the minimal TCR signal-responsive region within the fasl promoter. This region contains the previously identified binding sites for NF-κB and Egr and the AP-1 site identified in this study. We found that TCR signaling induces AP-1 binding to this site and regulates the fasl promoter function in a fashion dependent on NF-κB binding. However, mutation in the AP-1 site alone did not show a significant effect on the promoter function. The data suggest that the minimal promoter required at least two transcription factors to function.

A novel role of IL-2 in organ-specific autoimmune inflammation beyond regulatory T cell checkpoint: both IL-2 knockout and Fas mutation prolong lifespan of Scurfy mice but by different mechanisms.

Mutation of the Foxp3 transcription factor in Scurfy (Sf) mice results in complete absence of the CD4+Foxp3+ regulatory T cells (Tregs), severe multiorgan autoimmune syndrome, and early death at 4 wk of age. However, Sf mice simultaneously bearing the Il2−/− (Sf.Il2−/−) or Faslpr/lpr gene (Sf.Faslpr/lpr) have extended lifespan despite totally lacking Tregs, indicating a role of IL-2 and CD95 (Fas) signaling pathways in the multiorgan autoimmune syndrome beyond the Treg checkpoint. IL-2 has been implicated in regulating lymphoproliferation and CD178 (FasL) expression. However, Sf.Il2−/− mice have increased lymphoproliferation and FasL expression. Importantly, the pattern of organ-specific autoimmune response of Sf.Il2−/−mice resembled IL-2 knockout mice whereas that of Sf.Faslpr/lpr was similar to Sf mice, indicating that the distinct and weakened autoimmune manifestation in IL-2 knockout mice was not caused by the residual Tregs. Our study demonstrated a novel role of IL-2 in regulating multiorgan autoimmune inflammation beyond the Treg checkpoint and indicated that both Il2−/− and Faslpr/lpr genes prolong the lifespan of Sf mice but by different mechanisms.

Cutting Edge: A Regulatory T Cell-Dependent Novel Function of CD25 (IL-2Rα) Controlling Memory CD8+ T Cell Homeostasis

A massive systemic expansion of CD8+ memory T (TM) cells and a remarkable increase in circulating IL-2 were observed only in IL-2Rα (CD25) knockout (KO) mice but not in IL-2 KO and scurfy mice, although all three mutants lack regulatory T (Treg) cells. However, both phenotypes were suppressed by the transfer of Treg cells. The data presented indicate that Treg cell deficiency drives naive T cells to TM cells. The lack of high-affinity IL-2R in IL-2Rα KO mice increases circulating IL-2 that is then preferentially used by CD8+ TM cells through its abundant low-affinity IL-2R, resulting in systemic CD8+ TM cell dominance. Our study demonstrates the critical control of CD8+ TM cell homeostasis by a Treg cell-dependent novel function of CD25 and resolves its mechanism of action.