bluuebunny/crossref_metadata_2025 huggingface dataset:
A Critical Role for Allograft Inflammatory Factor-1 in the Pathogenesis of Rheumatoid Arthritis
Authors: Kimura Mizuho , Kawahito Yutaka , Obayashi Hiroshi , Ohta Mitsuhiro , Hara Hirokazu , Adachi Tetsuo , Tokunaga Daisaku , Hojo Tatsuya , Hamaguchi Masahide , Omoto Atsushi , Ishino Hidetaka , Wada Makoto , Kohno Masataka , Tsubouchi Yasunori , Yoshikawa Toshikazu
Published: April 2014

Rheumatoid arthritis (RA) is characterized by massive synovial proliferation, angiogenesis, subintimal infiltration of inflammatory cells and the production of cytokines such as TNF-α and IL-6. Allograft inflammatory factor-1 (AIF-1) has been identified in chronic rejection of rat cardiac allografts as well as tissue inflammation in various autoimmune diseases. AIF-1 is thought to play an important role in chronic immune inflammatory processes, especially those involving macrophages. In the current work, we examined the expression of AIF-1 in synovial tissues and measured AIF-1 in synovial fluid (SF) derived from patients with either RA or osteoarthritis (OA). We also examined the proliferation of synovial cells and induction of IL-6 following AIF-1 stimulation. Immunohistochemical staining showed that AIF-1 was strongly expressed in infiltrating mononuclear cells and synovial fibroblasts in RA compared with OA. Western blot analysis and semiquantitative RT-PCR analysis demonstrated that synovial expression of AIF-1 in RA was significantly greater than the expression in OA. AIF-1 induced the proliferation of cultured synovial cells in a dose-dependent manner and increased the IL-6 production of synovial fibroblasts and PBMC. The levels of AIF-1 protein were higher in synovial fluid from patients with RA compared with patients with OA (p < 0.05). Furthermore, the concentration of AIF-1 significantly correlated with the IL-6 concentration (r = 0.618, p < 0.01). These findings suggest that AIF-1 is closely associated with the pathogenesis of RA and is a novel member of the cytokine network involved in the immunological processes underlying RA.
C3d Binding to the Myelin Oligodendrocyte Glycoprotein Results in an Exacerbated Experimental Autoimmune Encephalomyelitis
Authors: Jégou Jean-François , Chan Philippe , Schouft Marie-Thérèse , Griffiths Mark R. , Neal James W. , Gasque Philippe , Vaudry Hubert , Fontaine Marc
Published: April 2014

The complement system is known to contribute to demyelination in multiple sclerosis and experimental autoimmune encephalomyelitis. However, there are few data concerning the natural adjuvant effect of C3d on the humoral response when it binds to myelin Ags. This study addresses the effect of C3d binding to the myelin oligodendrocyte glycoprotein (MOG) in the induction of experimental autoimmune encephalomyelitis in C57BL/6J mice. Immunization with human MOG coupled to C3d was found to accelerate the appearance of clinical signs of the disease and to enhance its severity compared with MOG-immunized mice. This finding was correlated with an increased infiltration of leukocytes into the central nervous system accompanied by increased complement activation and associated with areas of demyelination and axonal loss. Furthermore, B cell participation in the pathogenesis of the disease was determined by their increased capacity to act as APCs and to form germinal centers. Consistent with this, the production of MOG-specific Abs was found to be enhanced following MOG/C3d immunization. These results suggest that binding of C3d to self-Ags could increase the severity of an autoimmune disease by enhancing the adaptive autoimmune response.
Maternal Acceptance of the Fetus: True Human Tolerance
Authors: Guleria Indira , Sayegh Mohamed H.
Published: April 2014

Induction and maintenance of immunologic tolerance in humans remains a desirable but elusive goal. Therefore, understanding the physiologic mechanisms of regulation of immune responses is highly clinically relevant for immune-mediated diseases (e.g., autoimmunity and asthma/allergy) and for cell and organ transplantation. Acceptance of the fetus, which expresses paternally inherited alloantigens, by the mother during pregnancy is a unique example of how the immune system reshapes a destructive alloimmune response to a state of tolerance. Understanding the complex mechanisms of fetomaternal tolerance has important implications for developing novel strategies to induce immunologic tolerance in humans in general and for prevention of spontaneous abortion in at-risk populations in particular.
Cutting Edge: Acute and Chronic Exposure of Immature B Cells to Antigen Leads to Impaired Homing and SHIP1-Dependent Reduction in Stromal Cell-Derived Factor-1 Responsiveness
Authors: Brauweiler Anne , Merrell Kevin , Gauld Stephen B. , Cambier John C.
Published: April 2014

An encounter of B cells with cognate self Ags in the periphery can lead to anergy, a condition characterized by altered anatomical localization, shortened life span, and refractility to Ag stimulation. We recently reported that an immature B cell encounter with cognate self-Ag in the bone marrow can also lead to anergy. In this study we show that anergic as well as acutely Ag-stimulated immature B cells are defective in stromal cell-derived factor-1 (SDF-1)-induced calcium mobilization and migration and do not localize to bone marrow following adoptive transfer. This hyporesponsiveness does not involve CXCR4 modulation. However, BCR signal-mediated hyporesponsiveness to SDF-1 is associated with phosphorylation of the 5-inositol phosphatase SHIP1 and requires SHIP1 expression. Therefore, an encounter with cognate Ag may, by preventing SDF-1-induced phosphatidylinositol 3,4,5-triphosphate accumulation, trigger premature emigration of immature B cells from bone marrow.
Cutting Edge: Chemokine Receptor CCR4 Is Necessary for Antigen-Driven Cutaneous Accumulation of CD4 T Cells under Physiological Conditions
Authors: Campbell James J. , O’Connell Daniel J. , Wurbel Marc-André
Published: April 2014

Dual expression of chemokine receptor CCR4 and E-selectin ligand is characteristic of skin-tropic CD4 T cells from blood, lymphoid organs, and the skin itself. A strong and specific correlation exists among CCR4, its ligand CCL17/TARC, and the cutaneous lymphocyte-homing process. Nevertheless, whether CCR4 function is required for skin-specific trafficking remains an open question, which we address in this study. We developed an Ag-specific, TCR-transgenic, murine CD4 T cell adoptive transfer model that induces a mixed Th1 and Th17 cutaneous response. Within the hosts, both CCR4+/+ and CCR4−/− donor CD4 T cells contribute equally well to the circulating E-selectin ligand+ pool in response to Ag. However, only CCR4+/+ donor cells accumulate efficiently within the skin. CCR4−/− cells home normally to the peritoneum, showing that they do not have a general defect in lymphocyte trafficking. We conclude that under physiological conditions, CCR4 is a nonredundant, necessary component of skin-specific lymphocyte trafficking.
Cutting Edge: Proinflammatory and Th2 Cytokines Synergize to Induce Thymic Stromal Lymphopoietin Production by Human Skin Keratinocytes
Authors: Bogiatzi Sofia I. , Fernandez Isabel , Bichet Jean-Christophe , Marloie-Provost Marie-Annick , Volpe Elisabetta , Sastre Xavier , Soumelis Vassili
Published: April 2014

Thymic stromal lymphopoietin (TSLP) is an epithelial cell-derived cytokine that strongly activates dendritic cells (DC) and can initiate allergic inflammation. The factors inducing the production of human TSLP are not known. In this study, we show that proinflammatory (TNF-α or IL-1α) and Th2 (IL-4 or IL-13) cytokines synergized to induce the production of TSLP in human skin explants. TSLP production in situ was restricted to epidermal keratinocytes of the suprabasal layer. TSLP production could not be inhibited by factors regulating Th2 inflammation, such as IL-10, TGF-β, or IFN-γ. Cytokine-treated skin culture supernatants induced the maturation of blood CD11c+ DC in a TSLP-dependent manner. Our data provide the first evidence of TSLP induction and subsequent DC activation in human skin. Blocking TSLP-inducing cytokines could represent a novel strategy for the treatment of allergic diseases.
The Fas/Fas Ligand System Inhibits Differentiation of Murine Osteoblasts but Has a Limited Role in Osteoblast and Osteoclast Apoptosis
Authors: Kovac̆ić Nataša , Lukić Ivan Krešimir , Grc̆ević Danka , Katavić Vedran , Croucher Peter , Marušić Ana
Published: April 2014

Apoptosis through Fas/Fas ligand (FasL) is an important regulator of immune system homeostasis but its role in bone homeostasis is elusive. We systematically analyzed: 1) the expression of Fas/FasL during osteoblastogenesis and osteoclastogenesis in vitro, 2) the effect of FasL on apoptosis and osteoblastic/osteoclastic differentiation, and 3) osteoblastogenesis and osteoclastogenesis in mice deficient in Fas or FasL. The expression of Fas increased with osteoblastic differentiation. Addition of FasL weakly increased the proportion of apoptotic cells in both osteoclastogenic and osteoblastogenic cultures. In a CFU assay, FasL decreased the proportion of osteoblast colonies but did not affect the total number of colonies, indicating specific inhibitory effect of Fas/FasL on osteoblastic differentiation. The effect depended on the activation of caspase 8 and was specific, as addition of FasL to osteoblastogenic cultures significantly decreased gene expression for runt-related transcription factor 2 (Runx2) required for osteoblastic differentiation. Bone marrow from mice without functional Fas or FasL had similar osteoclastogenic potential as bone marrow from wild-type mice, but generated more osteoblast colonies ex vivo. These colonies had increased expression of the osteoblast genes Runx2, osteopontin, alkaline phosphatase, bone sialoprotein, osteocalcin, and osteoprotegerin. Our results indicate that Fas/FasL system primarily controls osteoblastic differentiation by inhibiting progenitor differentiation and not by inducing apoptosis. During osteoclastogenesis, the Fas/FasL system may have a limited effect on osteoclast progenitor apoptosis. The study suggests that Fas/FasL system plays a key role in osteoblastic differentiation and provides novel insight into the interactions between the immune system and bone.
Serpin-6 Expression Protects Embryonic Stem Cells from Lysis by Antigen-Specific CTL
Authors: Abdullah Zeinab , Saric Tomo , Kashkar Hamid , Baschuk Nikola , Yazdanpanah Benjamin , Fleischmann Bernd K. , Hescheler Jürgen , Krönke Martin , Utermöhlen Olaf
Published: April 2014

The immune response to embryonic stem (ES) cells is still poorly understood. In this study, we addressed the adaptive cellular immune response to undifferentiated and differentiated ES cells infected with lymphocytic choriomeningitis virus (LCMV), a vertically transmitted pathogen in mice and humans. In contrast to the prevailing view, we found that undifferentiated and differentiated murine ES cells express MHC class I molecules, although at low levels. When cocultured with LCMV-infected ES cells, syngeneic but not allogeneic LCMV-specific CTL secrete IFN-γ. Strikingly, LCMV-specific CTL do not efficiently kill LCMV-infected ES cells. ES cells showed high-level expression of the serine protease inhibitor 6, an endogenous inhibitor of the CTL-derived cytotoxic effector molecule granzyme B. Down-regulation of serpin-6 by RNA interference sensitized ES cells for CTL-induced cell death. The results of this study suggest that LCMV-infected murine ES cells present viral Ags and are recognized by LCMV-specific CTL in a MHC class I-restricted manner, yet resist CTL-mediated lysis through high-level expression of serine protease inhibitor 6.
Treatment-Enhanced CD4+Foxp3+ Glucocorticoid-Induced TNF Receptor Family RelatedHigh Regulatory Tumor-Infiltrating T Cells Limit the Effectiveness of Cytokine-Based Immunotherapy
Authors: Berhanu Aklile , Huang Jian , Watkins Simon C. , Okada Hideho , Storkus Walter J.
Published: April 2014

Regulatory T cells can suppress activated CD4+ and CD8+ T effector cells and may serve as an impediment to spontaneous or therapeutic type 1 antitumor immunity. In a previous study, we observed minimal therapeutic impact, but significantly enhanced T cell cross-priming and lesional infiltration of tumor-reactive CD8+ T cells into established CMS4 sarcomas after combined treatment of BALB/c mice with rFLt3 ligand (rFL) and recombinant GM-CSF (rGM-CSF). In this study, we show that this cytokine regimen also results in the profound enhancement of CD4+ tumor-infiltrating lymphocytes (TIL) expressing FoxP3, IL-10, and TGF-β mRNA, with 50 or 90% of CD4+ TIL coexpressing the CD25 and glucocorticoid-induced TNFR family related molecules, respectively. Intracellular staining for Foxp3 protein revealed that combined treatment with rFL plus rGM-CSF results in a significant increase in CD4+Foxp3+ T cells in the spleen of both control and tumor-bearing mice, and that nearly half of CD4+ TIL expressed this marker. In addition, CD4+ TIL cells were of an activated/memory (ICOShighCD62LlowCD45RBlow) phenotype and were capable of suppressing allospecific T cell proliferation and IFN-γ production from (in vivo cross-primed) anti-CMS4 CD8+ T cells in vitro, via a mechanism at least partially dependent on IL-10 and TGF-β. Importantly, in vivo depletion of CD4+ T cells resulted in the ability of previously ineffective, rFL plus rGM-CSF therapy-induced CD8+ T cells to now mediate tumor regression.
Targeting of the Transcription Factor STAT4 by Antisense Phosphorothioate Oligonucleotides Suppresses Collagen-Induced Arthritis
Authors: Hildner Kai M. , Schirmacher Peter , Atreya Imke , Dittmayer Maria , Bartsch Brigitte , Galle Peter R. , Wirtz Stefan , Neurath Markus F.
Published: April 2014

The transcription factor STAT4 mediates signals of various proinflammatory cytokines, such as IL-12, IL-15, and IL-23, that initiate and stabilize Th1 cytokine production. Although Th1 cytokine production has been suggested to play a major pathogenic role in rheumatoid arthritis, the role of STAT4 in this disease is poorly understood. In this study, we demonstrate a key functional role of STAT4 in murine collagen-induced arthritis (CIA). In initial studies we found that STAT4 expression is strongly induced in CD4+ T cells and to a lesser extent in CD11b+ APCs during CIA. To analyze the role of STAT4 for arthritis manifestation, we next investigated the outcome of interfering with STAT4 gene expression in CIA by using STAT4-deficient mice. Interestingly, STAT4-deficient mice developed significantly less severe arthritis than wild-type control mice and the T cells from such mice produced less IL-6, TNF, and IL-17. In addition, the targeting of STAT4 expression by a specific antisense phosphorothioate oligonucleotide directed at the translation start site suppressed STAT4 levels and signs of CIA even when applied during the onset of disease manifestation. These data suggest a key regulatory role of STAT4 in the pathogenesis and manifestation of murine collagen-induced arthritis. Furthermore, the targeting of STAT4 emerges as a novel approach to therapy for chronic arthritis.
Migration of CD4 T Cells and Dendritic Cells toward Sphingosine 1-Phosphate (S1P) Is Mediated by Different Receptor Subtypes: S1P Regulates the Functions of Murine Mature Dendritic Cells via S1P Receptor Type 3
Authors: Maeda Yasuhiro , Matsuyuki Hirofumi , Shimano Kyoko , Kataoka Hirotoshi , Sugahara Kunio , Chiba Kenji
Published: April 2014

Dendritic cells (DCs) and lymphocytes are known to show a migratory response to the phospholipid mediator, sphingosine 1-phosphate (S1P). However, it is unclear whether the same S1P receptor subtype mediates the migration of lymphocytes and DCs toward S1P. In this study, we investigated the involvement of S1P receptor subtypes in S1P-induced migration of CD4 T cells and bone marrow-derived DCs in mice. A potent S1P receptor agonist, the (S)-enantiomer of FTY720-phosphate [(S)-FTY720-P], at 0.1 nM or higher and a selective S1P receptor type 1 (S1P1) agonist, SEW2871, at 0.1 μM or higher induced a dose-dependent down-regulation of S1P1. The pretreatment with these compounds resulted in a significant inhibition of mouse CD4 T cell migration toward S1P. Thus, it is revealed that CD4 T cell migration toward S1P is highly dependent on S1P1. Mature DCs, when compared with CD4 T cells or immature DCs, expressed a relatively higher level of S1P3 mRNA. S1P at 10–1000 nM induced a marked migration and significantly enhanced the endocytosis of FITC-dextran in mature but not immature DCs. Pretreatment with (S)-FTY720-P at 0.1 μM or higher resulted in a significant inhibition of S1P-induced migration and endocytosis in mature DCs, whereas SEW2871 up to 100 μM did not show any clear effect. Moreover, we found that S1P-induced migration and endocytosis were at an extremely low level in mature DCs prepared from S1P3-knockout mice. These results indicate that S1P regulates migration and endocytosis of murine mature DCs via S1P3 but not S1P1.
B Cell Regulation of CD4+CD25+ T Regulatory Cells and IL-10 Via B7 is Essential for Recovery From Experimental Autoimmune Encephalomyelitis
Authors: Mann Monica K. , Maresz Katarzyna , Shriver Leah P. , Tan Yanping , Dittel Bonnie N.
Published: April 2014

CD4+CD25+ T regulatory (Treg) cells expressing the Foxp3 transcription factor have been shown to be present in the CNS during the autoimmune disease experimental autoimmune encephalomyelitis (EAE) and can inhibit EAE clinical disease by an IL-10-dependent mechanism. In addition, IL-10 expression in the CNS late in the EAE disease course has been attributed to recovery. However, it is not known how Treg cells and IL-10 expressions are regulated during EAE. We have previously shown a requirement for B cells in recovery from EAE and here investigated whether this was due to a deficiency in Treg cells and IL-10 in the CNS. We found that B cell deficiency resulted in a delay in the emergence of Foxp3-expressing Treg cells and IL-10 in the CNS during EAE, but not in the periphery. Reconstitution with wild-type B cells resulted in disease recovery and normalized IL-10 and Foxp3 expression. However, reconstitution with B7-deficient B cells did not. Furthermore, we show that IL-10 and Foxp3 expression is enhanced in CNS nonencephalitogenic T cells. These data suggest a novel mechanism whereby B cells regulate CD4+CD25+ Treg cells via B7 and subsequently enter the CNS and suppress autoimmune inflammation, mediating recovery.
Temporal Dissection of T-bet Functions
Authors: Matsuda Jennifer L. , George Thaddeus C. , Hagman James , Gapin Laurent
Published: April 2014

T-bet is a transcription factor of the T-box family that regulates the expression of numerous immune system-associated genes. T-bet directs the acquisition of the Th1-associated genetic program in differentiating CD4+ lymphocytes. It also influences the development of NK and NKT cells through its regulation of the IL-2/IL-15Rβ-chain (CD122) and the trafficking of these lymphocytes through CxCR3. The temporal requirements of T-bet activity for the production of IFN-γ and the regulation of CD122 and CxCR3 expression remain undefined. We produced an ectopically controllable form of T-bet by fusing its C-terminal domain with a mutated ligand-binding domain of human estrogen receptor α. By temporally controlling the expression of T-bet-estrogen receptor α by the addition or removal of 4-hydroxytamoxifen (4-HT), we show that IFN-γ, CD122, and CxCR3 are direct gene targets of T-bet whose expression are acutely regulated by T-bet activity.
Differential Survival of Cytotoxic T Cells and Memory Cell Precursors
Authors: Zhang Manling , Byrne Susan , Liu Ni , Wang Yue , Oxenius Annette , Ashton-Rickardt Philip G.
Published: April 2014

It is widely assumed that the development of memory CD8 T cells requires the escape of CTLs from programmed cell death. We show in this study that although serine protease inhibitor 6 (Spi6) is required to protect clonal bursts of CTLs from granzyme B-induced programmed cell death, it is not required for the development of memory cells. This conclusion is reached because memory cell precursors down-regulate both Spi6 and granzyme B, unlike CTLs, and they do not require Spi6 for survival. These findings suggest that memory CD8 T cells are derived from progenitors that are refractory to self-inflicted damage, rather than derived from fully differentiated CTLs.
Interleukin-15 but Not Interleukin-7 Abrogates Vaccine-Induced Decrease in Virus Level in Simian Immunodeficiency Virusmac251-Infected Macaques
Authors: Hryniewicz Anna , Price David A. , Moniuszko Marcin , Boasso Adriano , Edghill-Spano Yvette , West Sadie M. , Venzon David , Vaccari Monica , Tsai Wen-Po , Tryniszewska Elzbieta , Nacsa Janos , Villinger Francois , Ansari Aftab A. , Trindade Christopher J. , Morre Michel , Brooks David , Arlen Philip , Brown Helen J. , Kitchen Christina M. R. , Zack Jerome A. , Douek Daniel C. , Shearer Gene M. , Lewis Mark G. , Koup Richard A. , Franchini Genoveffa
Published: April 2014

The loss of CD4+ T cells and the impairment of CD8+ T cell function in HIV infection suggest that pharmacological treatment with IL-7 and IL-15, cytokines that increase the homeostatic proliferation of T cells and improve effector function, may be beneficial. However, these cytokines could also have a detrimental effect in HIV-1-infected individuals, because both cytokines increase HIV replication in vitro. We assessed the impact of IL-7 and IL-15 treatment on viral replication and the immunogenicity of live poxvirus vaccines in SIVmac251-infected macaques (Macaca mulatta). Neither cytokine augmented the frequency of vaccine-expanded CD4+ or CD8+ memory T cells, clonal recruitment to the SIV-specific CD8+ T cell pool, or CD8+ T cell function. Vaccination alone transiently decreased the viral set point following antiretroviral therapy suspension. IL-15 induced massive proliferation of CD4+ effector T cells and abrogated the ability of vaccination to decrease set point viremia. In contrast, IL-7 neither augmented nor decreased the vaccine effect and was associated with a decrease in TGF-β expression. These results underscore the importance of testing immunomodulatory approaches in vivo to assess potential risks and benefits for HIV-1-infected individuals.
Identification of Adiponectin as a Novel Hemopoietic Stem Cell Growth Factor
Authors: DiMascio Leah , Voermans Carlijn , Uqoezwa Mweia , Duncan Andrew , Lu Danhong , Wu Judy , Sankar Uma , Reya Tannishtha
Published: April 2014

The hemopoietic microenvironment consists of a diverse repertoire of cells capable of providing signals that influence hemopoietic stem cell function. Although the role of osteoblasts and vascular endothelial cells has recently been characterized, the function of the most abundant cell type in the bone marrow, the adipocyte, is less defined. Given the emergence of a growing number of adipokines, it is possible that these factors may also play a role in regulating hematopoiesis. Here, we investigated the role of adiponectin, a secreted molecule derived from adipocytes, in hemopoietic stem cell (HSC) function. We show that adiponectin is expressed by components of the HSC niche and its’ receptors AdipoR1 and AdipoR2 are expressed by HSCs. At a functional level, adiponectin influences HSCs by increasing their proliferation, while retaining the cells in a functionally immature state as determined by in vitro and in vivo assays. We also demonstrate that adiponectin signaling is required for optimal HSC proliferation both in vitro and in long term hemopoietic reconstitution in vivo. Finally we show that adiponectin stimulation activates p38 MAPK, and that inhibition of this pathway abrogates adiponectin’s proliferative effect on HSCs. These studies collectively identify adiponectin as a novel regulator of HSC function and suggest that it acts through a p38 dependent pathway.
Accelerated Notch-Dependent Degradation of E47 Proteins in Aged B Cell Precursors Is Associated with Increased ERK MAPK Activation
Authors: King Anne M. , Van der Put Elaine , Blomberg Bonnie B. , Riley Richard L.
Published: April 2014

The transcriptional regulator E47, encoded by the E2A gene, is crucial to B lymphopoiesis. In BALB/c senescent mice (∼2 years old), the incidence of E47-expressing pro-B cells in vivo and E47 protein steady state levels in B cell precursors in vitro were reduced. Poor expression of E47 protein was a consequence of accelerated proteasome-mediated turnover and was associated with heightened ubiquitin modification of E2A-encoded proteins in aged B cell precursors. Both MAPK and Notch activity have been previously associated with E2A-encoded protein stability in lymphocytes. Aged B cell precursors exhibited heightened levels of MAPK activity reflected in increased levels of phospho-ERK proteins. Phosphorylation of E2A-encoded proteins was also increased in aged B cell precursors and pharmacologic inhibition of MEK-1 resulted in a partial restoration of their E47 protein. Both Notch proteins and their Delta-like ligands were detected comparably in young and aged B cell precursors. Either inhibition of Notch activation via gamma-secretase or Ab blockade of Notch-Delta-like ligand interactions partially restored E47 expression in aged B cell precursors. We hypothesize that increased MAPK activity promotes phosphorylation of E2A-encoded protein in aged B cell precursors. Subsequently, E2A-encoded proteins undergo ubiquitination and accelerated degradation in a Notch-dependent process. The dysregulation of E2A-encoded protein expression may contribute to the reductions seen in early B lymphopoiesis during murine senescence.
Discrete T Cell Populations with Specificity for a Neo-Self-Antigen Bear Distinct Imprints of Tolerance
Authors: Standifer Nathan E. , Stacy Sue , Kraig Ellen , Infante Anthony J.
Published: April 2014

Mice expressing the Torpedo acetylcholine receptor α-chain as a neo-self-Ag exhibit a reduced frequency of T cells responding to the immunodominant epitope Tα146–162 indicating a degree of tolerance. We characterized tolerance induction in these animals by analyzing the residual Tα146–162-responsive T cell population and comparing it to that of nontransgenic littermates. Using CD4high sorting, we isolated the vast majority of Ag-reactive T cells from both strains of mice. Quantitative studies of the CD4high populations in transgenic mice following immunization with Tα146–162 revealed a diminished expansion of cells expressing the canonical TCRBV6 but not other TCRBV gene segments when compared with nontransgenic littermates. In addition, CD4high cells from transgenic mice were functionally hyporesponsive to Tα146–162 in terms of proliferation and cytokine secretion regardless of TCRBV gene segment use. TCR sequence analysis of transgenic Vβ6+CD4high cells revealed a reduced frequency of cells expressing a conserved motif within the TCRβ CDR3. Thus, the canonical Tα146–162 responsive, Vβ6+ population demonstrates both quantitative and qualitative deficits that correlate with an altered TCR repertoire whereas the non-Vβ6 population in transgenic mice exhibits only a reduction in peptide responsiveness, a qualitative defect. These data demonstrate that discrete autoreactive T cell populations with identical peptide/MHC specificity in Torpedo acetylcholine receptor-α-transgenic animals bear distinct tolerance imprints.
IL-12 Controls Cytotoxicity of a Novel Subset of Self-Antigen-Specific Human CD28+ Cytolytic T Cells
Authors: Barbey Catherine , Baumgaertner Petra , Devevre Estelle , Rubio-Godoy Verena , Derre Laurent , Bricard Gabriel , Guillaume Philippe , Luescher Immanuel F. , Liénard Danielle , Cerottini Jean-Charles , Romero Pedro , Rufer Nathalie , Speiser Daniel E.
Published: April 2014

Activated CD8 T cells develop cytotoxicity against autologous cells bearing foreign Ags and self/tumor Ags. However, self-specific cytolysis needs to be kept under control to avoid overwhelming immunopathology. After peptide vaccination of melanoma patients, we studied molecular and functional properties of T cell subsets specific for the self/tumor Ag Melan-A/MART-1. Ex vivo analysis revealed three Ag-specific effector memory (EM) populations, as follows: CD28-negative EM (EM28−) T cells strongly expressing granzyme/perforin, and two EM28+ subsets, one with high and the other with low level expression of these cytotoxic proteins. For further functional characterization, we generated 117 stable CD8 T cell clones by ex vivo flow cytometry-based sorting of these subsets. All EM28−-derived clones lysed target cells with high efficacy. In contrast, EM28+-derived clones were heterogenous, and could be classified in two groups, one with high and the other with low killing capacity, correlating with granzyme/perforin expression. High and low killer phenotypes remained surprisingly stable for several months. However, strongly increased granzyme expression and cytotoxicity were observed after exposure to IL-12. Thus, the data reveal a newly identified subset of CD28+ conditional killer T cells. Because CD28 can mediate strong costimulatory signals, tight cytotoxicity control, as shown in this study through IL-12, may be particularly important for subsets of T cells expressing CD28.
T-Independent Activation-Induced Cytidine Deaminase Expression, Class-Switch Recombination, and Antibody Production by Immature/Transitional 1 B Cells
Authors: Ueda Yoshihiro , Liao Dongmei , Yang Kaiyong , Patel Anjali , Kelsoe Garnett
Published: April 2014

Inflammation elicits a splenic lymphopoiesis of unknown physiologic significance but one that juxtaposes developing B cells and exogenous Ag. We show that immature and transitional 1 (immature/T1) B cells constitutively express activation-induced cytidine deaminase and B lymphocyte-induced maturation protein 1 in amounts that support accelerated plasmacytic differentiation and limited class-switch recombination. In vivo, activation of immature/T1 B cells by TLR ligands or bacterial vaccine rapidly induces T1 cells to divide, proliferate, and secrete IgM, IgG, or IgA Ab; in vitro, proliferation and differentiation are substantially enhanced by B cell-activating factor. We propose that inflammation-induced extramedullary lymphopoiesis represents a specialized mechanism for innate Ab responses to microbial pathogens.
IFN Regulatory Factor-2 Regulates Macrophage Apoptosis through a STAT1/3- and Caspase-1-Dependent Mechanism
Authors: Cuesta Natalia , Nhu Quan M. , Zudaire Enrique , Polumuri Swamy , Cuttitta Frank , Vogel Stefanie N.
Published: April 2014

IFN regulatory factor (IRF)-2−/− mice are significantly more resistant to LPS challenge than wild-type littermates, and this was correlated with increased numbers of apoptotic Kupffer cells. To assess the generality of this observation, and to understand the role of IRF-2 in apoptosis, responses of peritoneal macrophages from IRF-2+/+ and IRF-2−/− mice to apoptotic stimuli, including the fungal metabolite, gliotoxin, were compared. IRF-2−/− macrophages exhibited a consistently higher incidence of apoptosis that failed to correlate with caspase-3/7 activity. Using microarray gene expression profiling of liver RNA samples derived from IRF-2+/+ and IRF-2−/− mice treated with saline or LPS, we identified >40 genes that were significantly down-regulated in IRF-2−/− mice, including Stat3, which has been reported to regulate apoptosis. Compared with IRF-2+/+ macrophages, STAT3α mRNA was up-regulated constitutively or after gliotoxin treatment of IRF-2−/− macrophages, whereas STAT3β mRNA was down-regulated. Phospho-Y705-STAT3, phospho-S727-STAT1, and phospho-p38 protein levels were also significantly higher in IRF-2−/− than control macrophages. Activation of the STAT signaling pathway has been shown to elicit expression of CASP1 and apoptosis. IRF-2−/− macrophages exhibited increased basal and gliotoxin-induced caspase-1 mRNA expression and enhanced caspase-1 activity. Pharmacologic inhibition of STAT3 and caspase-1 abolished gliotoxin-induced apoptosis in IRF-2−/− macrophages. A novel IFN-stimulated response element, identified within the murine promoter of Casp1, was determined to be functional by EMSA and supershift analysis. Collectively, these data support the hypothesis that IRF-2 acts as a transcriptional repressor of Casp1, and that the absence of IRF-2 renders macrophages more sensitive to apoptotic stimuli in a caspase-1-dependent process.
Stimulation of an Unfolded Protein Response Impairs MHC Class I Expression
Authors: de Almeida Sérgio F. , Fleming John V. , Azevedo Jorge E. , Carmo-Fonseca Maria , de Sousa Maria
Published: April 2014

HFE C282Y is an example of a mutant protein that does not fold correctly, is retained in the endoplasmic reticulum, and was found previously to diminish surface expression of MHC class I (MHC-I). We now show that its expression in 293T cells triggers an unfolded protein response (UPR), as revealed by the increased levels of H chain binding protein, GRP94, and C/EBP homologous protein. Elevated levels of these proteins were also found in HFE C282Y homozygous PBMCs. Following the UPR induction, a decrease in MHC-I cell surface expression was observed. This defect in MHC-I could be mimicked, however, by overexpression of transcriptionally active isoforms of activating transcription factor-6 and X box-binding protein-1, which induced the UPR, and reversed in HFE C282Y-expressing cells by using dominant-negative constructs that block UPR signaling. The present results provide evidence to the finding that stimulation of an UPR affects MHC-I expression.
CD1-Restricted Recognition of Exogenous and Self-Lipid Antigens by Duodenal γδ+ T Lymphocytes
Authors: Russano Anna M. , Bassotti Gabrio , Agea Elisabetta , Bistoni Onelia , Mazzocchi Alessandro , Morelli Antonio , Porcelli Steven A. , Spinozzi Fabrizio
Published: April 2014

γδ T cells are present in the mucosal intestinal epithelia and secrete factors necessary to maintain tissue integrity. Ags recognized by these cells are poorly defined, although in mice non-classical MHC class I molecules have been implicated. Since MHC class I-like CD1 receptors are widely expressed at the surface of epithelial and dendritic intestinal cells and have the capacity to present lipid Ags to T cells, we hypothesized that these molecules might present autologous and/or exogenous phospholipids to intestinal γδ T lymphocytes. Intraepithelial T lymphocytes from normal human duodenal mucosal biopsies were cloned and exposed to natural and synthetic phospholipids using CD1a-, CD1b-, CD1c- or CD1d-transfected C1R lymphoblastoid or HeLa cell lines as APCs. Their cytolytic properties and regulatory cytokine secretion were also examined. Most clones obtained from duodenal mucosa (up to 70%) were TCRαβ+, and either CD4+ or CD8+, whereas 20% were CD4−CD8− (6 clones) or TCRγδ+ (12 clones). A relevant percentage (up to 66%) of TCRγδ+ but few (<5%) TCRαβ+ T cell clones responded to synthetic and/or natural phospholipids presented by CD1 molecules, as measured by both [3H]thymidine incorporation and IL-4 release assays. A Th1-like cytolytic and functional activity along with the ability to secrete regulatory cytokines was observed in most phospholipid-specific γδ T cell clones. Thus, a substantial percentage of TCRγδ+ but few TCRαβ+ from human duodenal mucosa recognize exogenous phospholipids in a CD1-restricted fashion. This adaptive response could contribute to mucosal homeostasis, but could also favor the emergence of inflammatory or allergic intestinal diseases.
Superantigen Presentation by Airway Smooth Muscle to CD4+ T Lymphocytes Elicits Reciprocal Proasthmatic Changes in Airway Function
Authors: Veler Haviva , Hu Aihua , Fatma Sumbul , Grunstein Judith S. , DeStephan Christine M. , Campbell Donald , Orange Jordan S. , Grunstein Michael M.
Published: April 2014

Microbial products serving as superantigens (SAgs) have been implicated in triggering various T cell-mediated chronic inflammatory disorders, including severe asthma. Given earlier evidence demonstrating that airway smooth muscle (ASM) cells express MHC class II molecules, we investigated whether ASM can present SAg to resting CD4+ T cells, and further examined whether this action reciprocally elicits proasthmatic changes in ASM responsiveness. Coincubation of CD4+ T cells with human ASM cells pulsed with the SAg, staphylococcal enterotoxin A (SEA), elicited adherence and clustering of class II and CD3 molecules at the ASM/T cell interface, indicative of immunological synapse formation, in association with T cell activation. This ASM/T cell interaction evoked up-regulated mRNA expression and pronounced release of the Th2-type cytokine, IL-13, into the coculture medium, which was MHC class II dependent. Moreover, when administering the conditioned medium from the SEA-stimulated ASM/T cell cocultures to isolated naive rabbit ASM tissues, the latter exhibited proasthmatic-like changes in their constrictor and relaxation responsiveness that were prevented by pretreating the tissues with an anti-IL-13 neutralizing Ab. Collectively, these observations are the first to demonstrate that ASM can present SAg to CD4+ T cells, and that this MHC class II-mediated cooperative ASM/T cell interaction elicits release of IL-13 that, in turn, evokes proasthmatic changes in ASM constrictor and relaxant responsiveness. Thus, a new immuno-regulatory role for ASM is identified that potentially contributes to the pathogenesis of nonallergic (intrinsic) asthma and, accordingly, may underlie the reported association between microbial SAg exposure, T cell activation, and severe asthma.
Capture of Target Cell Membrane Components via Trogocytosis Is Triggered by a Selected Set of Surface Molecules on T or B Cells
Authors: Hudrisier Denis , Aucher Anne , Puaux Anne-Laure , Bordier Christine , Joly Etienne
Published: April 2014

Key events of T and B cell biology are regulated through direct interaction with APC or target cells. Trogocytosis is a process whereby CD4+ T, CD8+ T, and B cells capture their specific membrane-bound Ag through the acquisition of plasma membrane fragments from their cellular targets. With the aim of investigating whether the ability to trigger trogocytosis was a selective property of Ag receptors, we set up an assay that allowed us to test the ability of many different cell surface molecules to trigger trogocytosis. On the basis of the analysis of a series of surface molecules on CD4+ T, CD8+ T, and B cells, we conclude that a set of cell type-specific surface determinants, including but not limited to Ag receptors, do trigger trogocytosis. On T cells, these determinants include components of the TCR/CD3 as well as that of coreceptors and of several costimulatory molecules. On B cells, we identified only the BCR and MHC molecules as potentials triggers of trogocytosis. Remarkably, latrunculin, which prevents actin polymerization, impaired trogocytosis by T cells, but not by B cells. This was true even when the same Abs were used to trigger trogocytosis in T or B cells. Altogether, our results indicate that although trogocytosis is performed by all hemopoietic cells tested thus far, both the receptors and the mechanisms involved can differ depending on the lineage of the cell acquiring membrane materials from other cells. This could therefore account for the different biological consequences of Ag capture via trogocytosis proposed for different types of cells.
Genome-Wide Identification of Novel Genes Involved in Early Th1 and Th2 Cell Differentiation
Authors: Lund Riikka J. , Löytömäki Maritta , Naumanen Tiina , Dixon Craig , Chen Zhi , Ahlfors Helena , Tuomela Soile , Tahvanainen Johanna , Scheinin Joonas , Henttinen Tiina , Rasool Omid , Lahesmaa Riitta
Published: April 2014

Th cell subtypes, Th1 and Th2, are involved in the pathogenesis or progression of many immune-mediated diseases, such as type 1 diabetes and asthma, respectively. Defining the molecular networks and factors that direct Th1 and Th2 cell differentiation will help to understand the pathogenic mechanisms causing these diseases. Some of the key factors regulating this differentiation have been identified, however, they alone do not explain the process in detail. To identify novel factors directing the early differentiation, we have studied the transcriptomes of human Th1 and Th2 cells after 2, 6, and 48 h of polarization at the genome scale. Based on our current and previous studies, 288 genes or expressed sequence tags, representing ∼1–1.5% of the human genome, are regulated in the process during the first 2 days. These transcriptional profiles revealed genes coding for components of certain pathways, such as RAS oncogene family and G protein-coupled receptor signaling, to be differentially regulated during the early Th1 and Th2 cell differentiation. Importantly, numerous novel genes with unknown functions were identified. By using short-hairpin RNA knockdown, we show that a subset of these genes is regulated by IL-4 through STAT6 signaling. Furthermore, we demonstrate that one of the IL-4 regulated genes, NDFIP2, promotes IFN-γ production by the polarized human Th1 lymphocytes. Among the novel genes identified, there may be many factors that play a crucial role in the regulation of the differentiation process together with the previously known factors and are potential targets for developing therapeutics to modulate Th1 and Th2 responses.
Characterization of B7S3 as a Novel Negative Regulator of T Cells
Authors: Yang Yang , Liu Xikui K. , Nguyen Thang , Bishop Caroline , Graf Daniel , Dong Chen
Published: April 2014

T cell activation by APCs is regulated by B7-like costimulatory molecules. In this study, we describe a new B7 superfamily member, B7S3, with two differentially spliced isoforms expressed in lymphoid and nonlymphoid tissues. A soluble B7S3-Ig protein bound to professional APC constitutively as well as to activated but not naive T cells. B7S3-Ig treatment greatly inhibited T cell proliferation and IL-2 production. B7S3-Ig also reduced cytokine production by effector T cells. Interestingly, although human genome appears to contain a single-copy B7S3 homolog, the mouse B7S3 gene has 10 relatives within a 2-Mb region constituting a B7S3 gene family. This study identifies B7S3 as a novel negative regulator of T cells, and suggests evolutionarily divergent T cell regulation mechanisms in mammals.
Importance of the CD3γ Ectodomain Terminal β-Strand and Membrane Proximal Stalk in Thymic Development and Receptor Assembly
Authors: Touma Maki , Sun Zhen-Yu J. , Clayton Linda K. , Marissen Wilfred E. , Kruisbeek Ada M. , Wagner Gerhard , Reinherz Ellis L.
Published: April 2014

CD3εγ and CD3εδ are noncovalent heterodimers; each consists of Ig-like extracellular domains associated side-to-side via paired terminal β-strands that are linked to individual subunit membrane proximal stalk segments. CD3ε, CD3γ, and CD3δ stalks contain the RxCxxCxE motif. To investigate the functional importance of a CD3 stalk and terminal β-strand, we created a CD3γ double mutant CD3γC82S/C85S and a CD3γ β-strand triple mutant CD3γQ76S/Y78A/Y79A for use in retroviral transduction of lymphoid progenitors for comparison with CD3γwt. Although both mutant CD3γ molecules reduced association with CD3ε in CD3εγ heterodimers, CD3γQ76S/Y78A/Y79A abrogated surface TCR expression whereas CD3γC82S/C85S did not. Furthermore, CD3γC82S/C85S rescued thymic development in CD3γ−/− fetal thymic organ culture. However, the numbers of double-positive and single-positive thymocytes after CD3γC82S/C85S transduction were significantly reduced despite surface pre-TCR and TCR expression comparable to that of CD3γ−/− thymocytes transduced in fetal thymic organ culture with a retrovirus harboring CD3γwt cDNA. Furthermore, double-negative thymocyte development was perturbed with attenuated double-negative 3/double-negative 4 maturation and altered surface-expressed CD3εγ, as evidenced by the loss of reactivity with CD3γ N terminus-specific antisera. Single histidine substitution of either CD3γ stalk cysteine failed to restore CD3εγ association and conformation in transient COS-7 cell transfection studies. Thus, CD3γC82 and CD3γC85 residues likely are either reduced or form a tight intrachain disulfide loop rather than contribute to a metal coordination site in conjunction with CD3εC80 and CD3εC83. The implications of these results for CD3εγ and TCR structure and signaling function are discussed.
Messenger RNA Expression of IL-8, FOXP3, and IL-12β Differentiates Latent Tuberculosis Infection from Disease
Authors: Wu Bo , Huang Chunhong , Kato-Maeda Midori , Hopewell Philip C. , Daley Charles L. , Krensky Alan M. , Clayberger Carol
Published: April 2014

Differentiation of active from latent tuberculosis (TB) is a major challenge in the control of TB. In this study, PBMC from latent TB-infected subjects, TB patients, and tuberculin skin test-negative donors stimulated with the Mycobacterium tuberculosis (Mtb)-specific Ag, early secretory antigenic target 6, and mRNA for 45 immune-related genes was measured by quantitative real-time PCR. Univariate analysis showed significant differences in the expression of 10 genes (IFN-γ, FOXP3, IL-1α, IL-1β, IL-2, IL-6, IL-8, IL-12α, IL-12β, and IL-24) in PBMC from TB patients vs latent TB-infected subjects (p < 0.01). Multivariate logistic regression and classification and regression tree analyses revealed that expression of three genes, IL-8, FOXP3, and IL-12β, is predictive for TB vs latent Mtb infection. Thus, measurement of Ag-specific expression of these three genes may offer a specific and noninvasive means of differentiating between latent Mtb infection and TB.
Induction of Protective Immunity to Listeria monocytogenes in Neonates
Authors: Kollmann Tobias R. , Reikie Brian , Blimkie Darren , Way Sing Sing , Hajjar Adeline M. , Arispe Kiea , Shaulov Angela , Wilson Christopher B.
Published: April 2014

Neonates suffer unduly from infections and also respond suboptimally to most commonly used vaccines. However, a CD8 T cell response can be elicited in neonates if the Ag is introduced into the cytoplasm of APCs. Listeria monocytogenes (Lm) targets the cytoplasm of APC and is a strong CD8 and CD4 Th1-promoting vaccine vehicle in adult mice. We hypothesized that an attenuated strain of Lm would be safe and induce long-lasting protective immunity, even in neonates. We found that neonatal mice immunized only once with the attenuated strain ΔactA-Lm developed robust primary and secondary CD8 and CD4 Th1 responses and were fully protected from lethal challenge with virulent wild-type Lm without the need for a booster immunization. Furthermore, ΔactA-Lm expressing a heterologous recombinant Ag induced a strong CD8 and Th1 memory response to that Ag. Based on these data, we propose that ΔactA-Lm or derivatives thereof might serve as a vaccine vehicle for neonatal immunization.