参考文献

Type I IFNs are key mediators of immune defense against viruses and bacteria. Type I IFNs were also previously implicated in protection against fungal infection,but their roles in antifungal immunity have not been thoroughly investigated. A recent study demonstrated that bacterial and fungal β-glucans stimulate IFN-β production by dendritic cells (DCs) following detection by the Dectin-1 receptor,but the effects of β-glucan-induced type I IFNs have not been defined. We investigated whether type I IFNs regulate CD8 T cell activation by fungal β-glucan particle-stimulated DCs. We demonstrate that β-glucan-stimulated DCs induce CD8 T cell proliferation,activation marker (CD44 and CD69) expression,and production of IFN-γ,IL-2,and granzyme B. Moreover,we show that type I IFNs support robust CD8 T cell activation (proliferation and IFN-γ and granzyme B production) by β-glucan-stimulated DCs in vitro and in vivo due to autocrine effects on the DCs. Specifically,type I IFNs promote Ag presentation on MHC I molecules,CD86 and CD40 expression,and the production of IL-12 p70,IL-2,IL-6,and TNF-α by β-glucan-stimulated DCs. We also demonstrate a role for autocrine type I IFN signaling in bacterial LPS-induced DC maturation,although,in the context of LPS stimulation,this mechanism is not so critical for CD8 T cell activation (promotes IFN-γ production but not proliferation or granzyme B production). This study provides insight into the mechanisms underlying CD8 T cell activation during infection,which may be useful in the rational design of vaccines directed against pathogens and tumors. View Publication

Macroautophagy plays an important role in the regulation of cell survival,metabolism,and the lysosomal degradation of cytoplasmic material. In the immune system,autophagy contributes to the clearance of intracellular pathogens,MHCII cross-presentation of endogenous Ags,as well as cell survival. We and others have demonstrated that autophagy occurs in T lymphocytes and contributes to the regulation of their cellular function,including survival and proliferation. Here we show that the essential autophagy gene Atg7 is required in a cell-intrinsic manner for the survival of mature primary T lymphocytes. We also find that mitochondrial content is developmentally regulated in T but not in B cells,with exit from the thymus marking a transition from high mitochondrial content in thymocytes to lower mitochondrial content in mature T cells. Macroautophagy has been proposed to play an important role in the clearance of intracellular organelles,and autophagy-deficient mature T cells fail to reduce their mitochondrial content in vivo. Consistent with alterations in mitochondrial content,autophagy-deficient T cells have increased reactive oxygen species production as well as an imbalance in pro- and antiapoptotic protein expression. With much recent interest in the possibility of autophagy-dependent developmentally programmed clearance of organelles in lens epithelial cells and erythrocytes,our data demonstrate that autophagy may have a physiologically significant role in the clearance of superfluous mitochondria in T lymphocytes as part of normal T cell homeostasis. View Publication

Myasthenia gravis (MG) is a prototypical autoimmune disease that is among the few for which the target Ag and the pathogenic autoantibodies are clearly defined. The pathology of the disease is affected by autoantibodies directed toward the acetylcholine receptor (AChR). Mature,Ag-experienced B cells rely on the action of Th cells to produce these pathogenic Abs. The phenotype of the MG Ag-reactive T cell compartment is not well defined; thus,we sought to determine whether such cells exhibit both a proinflammatory and a pathogenic phenotype. A novel T cell library assay that affords multiparameter interrogation of rare Ag-reactive CD4(+) T cells was applied. Proliferation and cytokine production in response to both AChR and control Ags were measured from 3120 T cell libraries derived from 11 MG patients and paired healthy control subjects. The frequency of CCR6(+) memory T cells from MG patients proliferating in response to AChR-derived peptides was significantly higher than that of healthy control subjects. Production of both IFN-γ and IL-17,in response to AChR,was also restricted to the CCR6(+) memory T cell compartment in the MG cohort,indicating a proinflammatory phenotype. These T cells also included an elevated expression of GM-CSF and absence of IL-10 expression,indicating a proinflammatory and pathogenic phenotype. This component of the autoimmune response in MG is of particular importance when considering the durability of MG treatment strategies that eliminate B cells,because the autoreactive T cells could renew autoimmunity in the reconstituted B cell compartment with ensuing clinical manifestations. View Publication

Broadly neutralizing HIV-1 antibodies (bNAbs) isolated from infected subjects display protective potential in animal models. Their elicitation by immunization is thus highly desirable. The HIV-1 envelope glycoprotein (Env) is the sole viral target of bnAbs,but is also targeted by binding,non-neutralizing antibodies. Env-based immunogens tested so far in various animal species and humans have elicited binding and autologous neutralizing antibodies but not bNAbs (with a few notable exceptions). The underlying reasons for this are not well understood despite intensive efforts to characterize the binding specificities of the elicited antibodies; mostly by employing serologic methodologies and monoclonal antibody isolation and characterization. These approaches provide limited information on the ontogenies and clonal B cell lineages that expand following Env-immunization. Thus,our current understanding on how the expansion of particular B cell lineages by Env may be linked to the development of non-neutralizing antibodies is limited. Here,in addition to serological analysis,we employed high-throughput BCR sequence analysis from the periphery,lymph nodes and bone marrow,as well as B cell- and antibody-isolation and characterization methods,to compare in great detail the B cell and antibody responses elicited in non-human primates by two forms of the clade C HIV Env 426c: one representing the full length extracellular portion of Env while the other lacking the variable domains 1,2 and 3 and three conserved N-linked glycosylation sites. The two forms were equally immunogenic,but only the latter elicited neutralizing antibodies by stimulating a more restricted expansion of B cells to a narrower set of IGH/IGK/IGL-V genes that represented a small fraction (0.003-0.02{\%}) of total B cells. Our study provides new information on how Env antigenic differences drastically affect the expansion of particular B cell lineages and supports immunogen-design efforts aiming at stimulating the expansion of cells expressing particular B cell receptors. View Publication

Class switch DNA recombination (CSR) is central to the maturation of the Ab response because it diversifies Ab effector functions. Like somatic hypermutation,CSR requires activation-induced cytidine deaminase (AID),whose expression is restricted to B cells,as induced by CD40 engagement or dual TLR-BCR engagement (primary CSR-inducing stimuli). By constructing conditional knockout Igh(+/C)γ(1-cre)Rab7(fl/fl) mice,we identified a B cell-intrinsic role for Rab7,a small GTPase involved in intracellular membrane functions,in mediating AID induction and CSR. Igh(+/C)γ(1-cre)Rab7(fl/fl) mice displayed normal B and T cell development and were deficient in Rab7 only in B cells undergoing Igh(C)γ(1-cre) Iγ1-Sγ1-Cγ1-cre transcription,as induced--like Igh germline Iγ1-Sγ1-Cγ1 and Iε-Sε-Cε transcription--by IL-4 in conjunction with a primary CSR-inducing stimulus. These mice could not mount T-independent or T-dependent class-switched IgG1 or IgE responses while maintaining normal IgM levels. Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells showed,in vivo and in vitro,normal proliferation and survival,normal Blimp-1 expression and plasma cell differentiation,as well as intact activation of the noncanonical NF-κB,p38 kinase,and ERK1/2 kinase pathways. They,however,were defective in AID expression and CSR in vivo and in vitro,as induced by CD40 engagement or dual TLR1/2-,TLR4-,TLR7-,or TLR9-BCR engagement. In Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells,CSR was rescued by enforced AID expression. These findings,together with our demonstration that Rab7-mediated canonical NF-κB activation,as critical to AID induction,outline a novel role of Rab7 in signaling pathways that lead to AID expression and CSR,likely by promoting assembly of signaling complexes along intracellular membranes. View Publication

Wild-type (WT) NOD.H-2h4 mice develop spontaneous autoimmune thyroiditis (SAT) when given 0.05% NaI in their drinking water,whereas B cell-deficient NOD.H-2h4 mice are SAT resistant. To test the hypothesis that resistance of B cell-deficient mice to SAT was due to the activity of regulatory CD4+CD25+ T (T reg) cells activated if autoantigen was initially presented on non-B cells,CD25+ T reg cells were transiently depleted in vivo using anti-CD25. B cell-deficient NOD.H-2h4 mice given three weekly injections of anti-CD25 developed SAT 8 wk after NaI water. Thyroid lesions were similar to those in WT mice except there were no B cells in thyroid infiltrates. WT and B cell-deficient mice had similar numbers of CD4+CD25+Foxp3+ cells. Mice with transgenic nitrophenyl-specific B cells unable to secrete immunoglobulin were also resistant to SAT,and transient depletion of T reg cells resulted in severe SAT with both T and B cells in thyroid infiltrates. T reg cells that inhibit SAT were eliminated by day 3 thymectomy,indicating they belong to the subset of naturally occurring T reg cells. However,T reg cell depletion did not increase SAT severity in WT mice,suggesting that T reg cells may be nonfunctional when effector T cells are activated; i.e.,by autoantigen-presenting B cells. View Publication

The immunodeficiency disorder,X-linked agammaglobulinemia (XLA),results from mutations in the gene encoding Bruton tyrosine kinase (Btk). Btk is required for pre-B cell clonal expansion and B-cell antigen receptor signaling. XLA patients lack mature B cells and immunoglobulin and experience recurrent bacterial infections only partially mitigated by life-long antibody replacement therapy. In pursuit of definitive therapy for XLA,we tested ex vivo gene therapy using a lentiviral vector (LV) containing the immunoglobulin enhancer (Emu) and Igbeta (B29) minimal promoter to drive B lineage-specific human Btk expression in Btk/Tec(-/-) mice,a strain that reproduces the features of human XLA. After transplantation of EmuB29-Btk-LV-transduced stem cells,treated mice showed significant,albeit incomplete,rescue of mature B cells in the bone marrow,peripheral blood,spleen,and peritoneal cavity,and improved responses to T-independent and T-dependent antigens. LV-treated B cells exhibited enhanced B-cell antigen receptor signaling and an in vivo selective advantage in the peripheral versus central B-cell compartment. Secondary transplantation showed sustained Btk expression,viral integration,and partial functional responses,consistent with long-term stem cell marking; and serial transplantation revealed no evidence for cellular or systemic toxicity. These findings strongly support pursuit of B lineage-targeted LV gene therapy in human XLA. View Publication

B lymphocyte stimulator (BLyS) is a well-known direct costimulator of adaptive immune cells,particularly B lineage cells. However,we have reported recently that BLyS is also able to activate monocytes. Other innate immune cells,such as dendritic cells (DCs),play a key role in the initiation of adaptive immune responses and the purpose of the current study was to assess whether there is a direct role for BLyS in modulating human DC functions. In this study,we show that BLyS induces DC activation and maturation. Thus,BLyS strongly induced up-regulation of surface costimulatory molecule expression and secretion of specific cytokines and chemokines in DCs. BLyS-stimulated DCs (BLyS-DCs) were also able to augment allogeneic CD4 T cell proliferation to a greater extent than control DCs. BLyS-DCs secreted elevated levels of the major Th1-polarizing cytokine,IL-12p70,and they promoted naive CD4 T cell differentiation into Th1 T cells. Regarding BLyS receptor expression,DCs primarily express cytoplasmic transmembrane activator and CAML interactor; however,low levels of cell surface transmembrane activator and CAML interactor are expressed as well. Collectively,our data suggest that BLyS may modulate adaptive immune cells indirectly by inducing DC maturation. View Publication

Bacteriophage are abundant at sites of bacterial infection,but their effects on mammalian hosts are unclear. We have identified pathogenic roles for filamentous Pf bacteriophage produced by Pseudomonas aeruginosa (Pa) in suppression of immunity against bacterial infection. Pf promote Pa wound infection in mice and are associated with chronic human Pa wound infections. Murine and human leukocytes endocytose Pf,and internalization of this single-stranded DNA virus results in phage RNA production. This triggers Toll-like receptor 3 (TLR3)- and TIR domain-containing adapter-inducing interferon-beta (TRIF)-dependent type I interferon production,inhibition of tumor necrosis factor (TNF),and the suppression of phagocytosis. Conversely,immunization of mice against Pf prevents Pa wound infection. Thus,Pf triggers maladaptive innate viral pattern-recognition responses,which impair bacterial clearance. Vaccination against phage virions represents a potential strategy to prevent bacterial infection. View Publication

The AP-1 factor basic leucine zipper transcription factor,ATF-like (BATF) is important for CD4(+) Th17,Th9,and follicular Th cell development. However,its precise role in Th2 differentiation and function remains unclear,and the requirement for BATF in nonallergic settings of type-2 immunity has not been explored. In this article,we show that,in response to parasitic helminths,Batf(-/-) mice are unable to generate follicular Th and Th2 cells. As a consequence,they fail to establish productive type-2 immunity during primary and secondary infection. Batf(-/-) CD4(+) T cells do not achieve type-2 cytokine competency,which implies that BATF plays a key role in the regulation of IL-4 and IL-13. In contrast to Th17 and Th9 cell subsets in which BATF binds directly to promoter and enhancer regions to regulate cytokine expression,our results show that BATF is significantly enriched at Rad50 hypersensitivity site (RHS)6 and RHS7 of the locus control region relative to AP-1 sites surrounding type-2 cytokine loci in Th2 cells. Indeed,Batf(-/-) CD4(+) T cells do not obtain permissive epigenetic modifications within the Th2 locus,which were linked to RHS6 and RHS7 function. In sum,these findings reveal BATF as a central modulator of peripheral and humoral hallmarks of type-2 immunity and begin to elucidate a novel mechanism by which it regulates type-2 cytokine production through its modification of the Th2 locus control region. View Publication