Xiongfei Xu

The E3 ubiquitin ligase Nrdp1 'preferentially' promotes TLR-mediated production of type I interferon

E3 ubiquitin ligases are important in both innate and adaptive immunity. Here we report that Nrdp1, an E3 ubiquitin ligase, inhibited the production of proinflammatory cytokines but increased interferon-β production in Toll-like receptor–triggered macrophages by suppressing adaptor MyD88–dependent activation of transcription factors NF-κB and AP-1 while promoting activation of the kinase TBK1 and transcription factor IRF3. Nrdp1 directly bound and polyubiquitinated MyD88 and TBK1, which led to degradation of MyD88 and activation of TBK1. Knockdown of Nrdp1 inhibited the degradation of MyD88 and the activation of TBK1 and IRF3. Nrdp1-transgenic mice showed resistance to lipopolysaccharide-induced endotoxin shock and to infection with vesicular stomatitis virus. Our data suggest that Nrdp1 functions as both an adaptor protein and an E3 unbiquitin ligase to regulate TLR responses in different ways.

Hepatic microenvironment programs hematopoietic progenitor differentiation into regulatory dendritic cells, maintaining liver tolerance

The liver has been generally considered an organ prone to tolerance induction and maintenance. However, whether and how the unique liver microenvironment contributes to tolerance maintenance is largely unknown. Here, we used liver fibroblastic stromal cells to mimic the liver microenvironment and found that liver stroma could induce Lin(-)CD117(+) progenitors to differentiate into dendritic cells (DCs) with low CD11c, MHC II but high CD11b expression, high IL-10, but low IL-12 secretion. Such regulatory DCs could inhibit T-cell proliferation in vitro and in vivo, induce apoptosis of the activated T cells, and alleviate the damage of autoimmune hepatitis. Furthermore, liver stroma-derived macrophage colony-stimulating factor (M-CSF) was found to contribute to the generation of such regulatory DCs. Regulatory DC-derived PGE2 and T cell-derived IFN-gamma were responsible for the regulatory function. The natural counterpart of regulatory DCs was phenotypically and functionally identified in the liver. Importantly, Lin(-)CD117(+) progenitors could be differentiated into regulatory DCs in the liver once transferred into the liver. Infusion with liver regulatory DCs alleviated experimental autoimmune hepatitis. Therefore, we demonstrate that the liver microenvironment is highly important to program progenitors to differentiate into regulatory DCs in situ, which contributes to the maintenance of liver tolerance.

IL-17A–Producing γδT Cells Promote CTL Responses against Listeria monocytogenes Infection by Enhancing Dendritic Cell Cross-Presentation

Interleukin-17A–producing T cells, especially Th17, have been shown to be involved in inflammatory autoimmune diseases and host defense against extracellular infections. However, whether and how IL-17A or IL-17A–producing cells can help protection against intracellular bacteria remains controversial, especially how it regulates the adaptive immunity besides recruitment of neutrophils in the innate immune system. By infecting IL-17A–deficient mice with Listeria monocytogenes, we show in this study that IL-17A is required for the generation of Ag-specific CD8+ CTL response against primary infection, but not for the generation of memory CD8+ T cells against secondary challenge. Interestingly, we identify γδT cells, but not conventional CD4+ Th17 cells, as the main cells for innate IL-17A production during L. monocytogenes infection. Furthermore, γδT cells are found to promote Ag-specific CD8+ T cell proliferation by enhancing cross-presentation of dendritic cells through IL-17A. Adoptive transfer of Il17a+/+ γδT cells, but not Il17a−/− γδT cells or Il17a+/+ CD4+ T cells, were sufficient to recover dendritic cells cross-presentation and defective CD8+ T cell response in Il17a−/− mice. Our findings indicate an important role of infection-inducible IL-17A–producing γδT cells and their derived IL-17A against intracellular bacterial infection, providing a mechanism of IL-17A for regulation of innate and adaptive immunity.

Pulmonary stromal cells induce the generation of regulatory DC attenuating T-cell-mediated lung inflammation.

The tissue microenvironment may affect the development and function of immune cells such as DC. Whether and how the pulmonary stromal microenvironment can affect the development and function of lung DC need to be investigated. Regulatory DC (DCreg) can regulate T‐cell response. We wondered whether such regulatory DC exist in the lung and what is the effect of the pulmonary stromal microenvironment on the generation of DCreg. Here we demonstrate that murine pulmonary stromal cells can drive immature DC, which are regarded as being widely distributed in the lung, to proliferate and differentiate into a distinct subset of DCreg, which express high levels of CD11b but low levels of MHC class II (I‐A), CD11c, secrete high amounts of IL‐10, NO and prostaglandin E2 (PGE2) and suppress T‐cell proliferation. The natural counterpart of DCreg in the lung with similar phenotype and regulatory function has been identified. Pulmonary stroma‐derived TGF‐β is responsible for the differentiation of immature DC to DCreg, and DCreg‐derived PGE2 contributes to their suppression of T‐cell proliferation. Moreover, DCreg can induce the generation of CD4+CD25+Foxp3+ Treg. Importantly, infusion with DCreg attenuates T‐cell‐mediated eosinophilic airway inflammation in vivo. Therefore, the pulmonary microenvironment may drive the generation of DCreg, thus contributing to the maintenance of immune homoeostasis and the control of inflammation in the lung.

The lectin Siglec-G inhibits dendritic cell cross-presentation by impairing MHC class I–peptide complex formation

CD8α+ dendritic cells (DCs) are specialized at cross-presenting extracellular antigens on major histocompatibility complex (MHC) class I molecules to initiate cytotoxic T lymphocyte (CTL) responses; however, details of the mechanisms that regulate cross-presentation remain unknown. We found lower expression of the lectin family member Siglec-G in CD8α+ DCs, and Siglec-G deficient (Siglecg−/−) mice generated more antigen-specific CTLs to inhibit intracellular bacterial infection and tumor growth. MHC class I–peptide complexes were more abundant on Siglecg−/− CD8α+ DCs than on Siglecg+/+ CD8α+ DCs. Mechanistically, phagosome-expressed Siglec-G recruited the phosphatase SHP-1, which dephosphorylated the NADPH oxidase component p47phox and inhibited the activation of NOX2 on phagosomes. This resulted in excessive hydrolysis of exogenous antigens, which led to diminished formation of MHC class I–peptide complexes for cross-presentation. Therefore, Siglec-G inhibited DC cross-presentation by impairing such complex formation, and our results add insight into the regulation of cross-presentation in adaptive immunity.

Splenic Stroma-Educated Regulatory Dendritic Cells Induce Apoptosis of Activated CD4 T Cells via Fas Ligand-Enhanced IFN-γ and Nitric Oxide

Stromal microenvironments of bone marrow, lymph nodes, and spleen have been shown to be able to regulate immune cell differentiation and function. Our previous studies demonstrate that splenic stroma could drive mature dendritic cells (DC) to further proliferate and differentiate into regulatory DC subset that could inhibit T cell response via NO. However, how splenic stroma-educated regulatory DC release NO and whether other molecules are involved in the suppression of T cell response remain unclear. In this study, we show that splenic stroma educates regulatory DC to express high level of Fas ligand (FasL) by TGF-β via ERK activation. The findings, that inhibition of CD4 T cell proliferation by regulatory DC required cell-to-cell contact and FasL deficiency impaired inhibitory effect of regulatory DC, indicate that regulatory DC inhibit CD4 T cell proliferation via FasL. Then, regulatory DC have been found to be able to induce apoptosis of activated CD4 T cells via FasL in caspase 8- and caspase 3-dependent manner. Interestingly, FasL on regulatory DC enhanced IFN-γ production from activated CD4 T cells, and in turn T cell-derived IFN-γ induced NO production from regulatory DC, working jointly to induce apoptosis of activated CD4 T cells. Blockade of IFN-γ and NO could reduce the apoptosis induction. Therefore, our results demonstrated that splenic stroma-educated regulatory DC induced T cell apoptosis via FasL-enhanced T cell IFN-γ and DC NO production, thus outlining a new way for negative regulation of T cell responses and maintenance of immune homeostasis by regulatory DC and splenic stromal microenvironment.

Regulatory dendritic cells program generation of interleukin-4–producing alternative memory CD4 T cells with suppressive activity

The heterogeneity and mechanisms for the generation of CD4 memory T (CD4 Tm) cells remain elusive. Distinct subsets of dendritic cells (DCs) have been found to regulate a distinct T-helper (Th)-cell subset differentiation by influencing cytokine cues around CD4 T cells; however, whether and how the regulatory DC subset can regulate Tm-cell differentiation remains unknown. Further, there is no ideal in vitro experimental system with which to mimic the 3 phases of the CD4 T-cell immune response (expansion, contraction, memory generation) and/or to culture CD4 Tm cells for more than a month. By analyzing CD4 T cells programmed by long-term coculture with regulatory DCs, we identified a population of long-lived CD4 T cells with a CD44(hi)CD62L(-)CCR7(-) effector memory phenotype and rapid, preferential secretion of the Th2 cytokines interleukin-4 (IL-4), IL-5, IL-10, and IL-13 after antigenic stimulation. These regulatory DC-programmed Tm cells suppress CD4 T-cell activation and proliferation in vitro via IL-10 and inhibit the delayed-type hypersensitivity response once infused in vivo. We also identify their natural counterpart, which is up-regulated by regulatory DC transfusion and negatively regulates the recall response in vivo. Different from interferon-γ-producing conventional Tm cells, these IL-4-producing CD4 Tm cells act as alternative Tm cells with a regulatory function, suggesting a new way of negative immune regulation by memory T cells.

Blockade of CD47 ameliorates autoimmune inflammation in CNS by suppressing IL-1-triggered infiltration of pathogenic Th17 cells

The migration of Th17 cells into central nervous system (CNS) tissue is the key pathogenic step in experimental autoimmune encephalomyelitis (EAE) model. However, the mechanism underlying the pathogenic Th17 cell migration remains elusive. Here we report that blockade of CD47 with CD47-Fc fusion protein is effective in preventing and curing EAE by impairing infiltration of Th17 cells into CNS. However, CD47 deficiency does not directly impair the migration of Th17 cells. Mechanistic studies showed that CD47 deficiency inhibited degradation of inducible nitric oxide synthase (iNOS) in proteasome of macrophages by Src activation and led to the increased nitric oxide (NO) production. Then NO suppressed inflammasome activation-induced IL-1β production. This lower IL-1β reduces the expression of IL-1R1 and migration-related chemokine receptors on CD47(-/-) Th17 cells, inhibiting the ability of Th17 cells to infiltrate into the CNS of CD47(-/-) mice and therefore suppressing EAE development. In vivo administration of exogenous IL-1β indeed promoted the infiltration CD47(-/-) Th17 cells into CNS and antagonized the protective role of CD47 deficiency in EAE pathogenesis. Our results demonstrate a potential preventive and therapeutic application of CD47 blockade in controlling EAE development.

Liver stroma enhances activation of TLR3-triggered NK cells through fibronectin

Recent studies demonstrate that tissue microenvironments may regulate development and functions of immune cells including dendritic cells, monocytes and T cells. However, there is no report about functional regulation of innate NK cells by tissue microenvironment. The liver is an organ abundant in NK cells and susceptible to virus infection. The number and cytotoxicity of liver NK cells have been shown to be increased during pathogenesis of viral hepatitis and contribute to liver damage. So, whether and how liver stromal microenvironment regulates NK cells need to be fully investigated. By preparing liver fibroblast stromal cells to mimic stromal microenvironment in liver, in this study we demonstrate that liver stroma could chemoattract and adhere TLR3-triggered NK cells, and further augment the cytotoxicity and IFN-gamma production of TLR3-triggered NK cells via fibronectin. Furthermore, the autocrined IFN-gamma from NK cells is required for the enhancement of TLR3-triggered NK cell activation by liver stroma. Our results suggest that liver stroma can recruit NK cells and promote activation of NK cells during viral infection, thus providing a mechanistic explanation for the increased number and cytotoxicity of liver NK cells which may cause liver damage during pathogenesis of viral hepatitis.