bluuebunny/crossref_metadata_2025 huggingface dataset:
Interaction of Mite Allergens Der P3 and Der P9 with Protease-Activated Receptor-2 Expressed by Lung Epithelial Cells
Authors: Sun Guo , Stacey Martin A. , Schmidt Matthias , Mori Luca , Mattoli Sabrina
Published: April 2014

The respiratory epithelium represents the first barrier encountered by airborne Ags. Two major dust mite Ags, Der p3 and Der p9, are serine proteases that may activate lung epithelial cells by interaction with the protease-activated receptor 2 (PAR-2). In this study both Der p3 and Der p9 cleaved the peptide corresponding to the N terminus of PAR-2 at the activation site. Both Ags sequentially stimulated phosphoinositide hydrolysis, transient cytosolic Ca2+ mobilization, and release of GM-CSF and eotaxin in human pulmonary epithelial cells. These responses were similar to those observed with trypsin and a specific PAR-2 agonist and were related to the serine protease activity of Der p3 and Der p9. Cell exposure to the Ags resulted in a refractory period, indicating that a PAR had been cleaved. Partial desensitization to Der p3 and Der p9 by the PAR-2 agonist suggested that PAR-2 was one target of the Ags. However, PAR-2 was not the only target, because the PAR-2 agonist caused less desensitization to Der p3 and Der p9 than did trypsin. A phospholipase C inhibitor prevented the cytokine-releasing effect of the PAR-2 agonist and abolished or reduced (>70%) the cytokine-releasing effects of Der p3 and Der p9. Our results suggest that Der p 3 and Der p9 may induce a nonallergic inflammatory response in the airways through the release of proinflammatory cytokines from the bronchial epithelium and that this effect is at least in part mediated by PAR-2.
Distinct Tissue Site-Specific Requirements of Mast Cells and Complement Components C3/C5a Receptor in IgG Immune Complex-Induced Injury of Skin and Lung
Authors: Baumann Ulrich , Chouchakova Nelli , Gewecke Britta , Köhl Jörg , Carroll Michael C. , Schmidt Reinhold E. , Gessner J. Engelbert
Published: April 2014

We induced the passive reverse Arthus reaction to IgG immune complexes (IC) at different tissue sites in mice lacking C3 treated or not with a C5aR-specific antagonist, or in mice lacking mast cells (KitW/KitW-v mice), and compared the inflammatory responses with those in the corresponding wild-type mice. We confirmed that IC inflammation of skin can be mediated largely by mast cells expressing C5aR and FcγRIII. In addition, we provided evidence for C3-independent C5aR triggering, which may explain why the cutaneous Arthus reaction develops normally in C3−/− mice. Furthermore, some, but not all, of the acute changes associated with the Arthus response in the lung were significantly more intense in normal mice than in C3−/− or KitW/KitW-v mice, indicating for C3- and mast cell-dependent and -independent components. Finally, we demonstrated that C3 contributed to the elicitation of neutrophils to alveoli, which corresponded to an increased synthesis of TNF-α, macrophage-inflammatory protein-2, and cytokine-induced neutrophil chemoattractant. While mast cells similarly influenced alveolar polymorphonuclear leukocyte influx, the levels of these cytokines remained largely unaffected in mast cell deficiency. Together, the phenotypes of C3−/− mice and KitW/KitW-v mice suggest that complement and mast cells have distinct tissue site-specific requirements acting by apparently distinct mechanisms in the initiation of IC inflammation.
H-2D End Confers Dominant Protection from IL-10-Mediated Acceleration of Autoimmune Diabetes in the Nonobese Diabetic Mouse
Authors: La Cava Antonio , Balasa Balaji , Good Augusta , van Gunst Kurt , Jung Nadja , Sarvetnick Nora
Published: April 2014

BALB/c mice that express IL-10 as a transgene in their pancreatic β cells (Ins-IL-10 mice) do not develop diabetes, even after crossing to nonobese diabetic (NOD) mice ((Ins-IL-10 × NOD)F1 mice). However, backcross of F1 mice to NOD mice (NOD.Ins-IL-10 mice) results in N2 and N3 generations that develop accelerated diabetes. In this study, we found that NOD.Ins-IL-10 mice that expressed BALB/c-derived MHC molecules (NOD.Ins-IL-10(H-2g7/d) mice) were protected from diabetes. This protection associated with peri-islet infiltration and preserved β cell function. Moreover, expression of I-Ad and I-Ed MHC class II molecules of BALB/c origin was not responsible for protection, but NOD.Ins-IL-10 mice that expressed BALB/c MHC class I Dd molecules (NOD.Ins-IL-10(H-2g7/d) mice) did not develop diabetes. To directly test the possibility of a protective role of H-2Dd in the development of accelerated diabetes, we generated transgenic mice expressing Dd under the control of the MHC class I promoter. We found that double transgenic NOD.Ins-IL-10-Dd mice developed accelerated diabetes in a fashion similar to NOD.Ins-IL-10 mice that were Dd negative. Microsatellite analysis of H-2Dd-linked loci confirmed association between BALB/c-derived alleles and protection of NOD.Ins-IL-10(H-2g7/d) mice. These results suggest a control of H-2Dd-linked gene(s) on IL-10-mediated acceleration of autoimmune diabetes and dominant protection of the Dd region in NOD.Ins-IL-10 mice.
Lipopolysaccharide-Activated B Cells Down-Regulate Th1 Immunity and Prevent Autoimmune Diabetes in Nonobese Diabetic Mice
Authors: Tian Jide , Zekzer Dan , Hanssen Lorraine , Lu Yuxin , Olcott Angelica , Kaufman Daniel L.
Published: April 2014

B cells can serve dual roles in modulating T cell immunity through their potent capacity to present Ag and induce regulatory tolerance. Although B cells are necessary components for the initiation of spontaneous T cell autoimmunity to β cell Ags in nonobese diabetic (NOD) mice, the role of activated B cells in the autoimmune process is poorly understood. In this study, we show that LPS-activated B cells, but not control B cells, express Fas ligand and secrete TGF-β. Coincubation of diabetogenic T cells with activated B cells in vitro leads to the apoptosis of both T and B lymphocytes. Transfusion of activated B cells, but not control B cells, into prediabetic NOD mice inhibited spontaneous Th1 autoimmunity, but did not promote Th2 responses to β cell autoantigens. Furthermore, this treatment induced mononuclear cell apoptosis predominantly in the spleen and temporarily impaired the activity of APCs. Cotransfer of activated B cells with diabetogenic splenic T cells prevented the adoptive transfer of type I diabetes mellitus (T1DM) to NOD/scid mice. Importantly, whereas 90% of NOD mice treated with control B cells developed T1DM within 27 wk, <20% of the NOD mice treated with activated B cells became hyperglycemic up to 1 year of age. Our data suggest that activated B cells can down-regulate pathogenic Th1 immunity through triggering the apoptosis of Th1 cells and/or inhibition of APC activity by the secretion of TGF-β. These findings provide new insights into T-B cell interactions and may aid in the design of new therapies for human T1DM.
CD4+ T Cells Engrafted with a Recombinant Immunoreceptor Efficiently Lyse Target Cells in a MHC Antigen- and Fas-Independent Fashion
Authors: Hombach Andreas , Heuser Claudia , Marquardt Thomas , Wieczarkowiecz Anja , Groneck Volker , Pohl Christoph , Abken Hinrich
Published: April 2014

T cells engrafted by a recombinant immunoreceptor with predefined Ag specificity can efficiently lyse Ag-positive target cells in a MHC Ag-independent manner. It is yet unresolved how receptor-grafted CD4+ T cells contribute to MHC Ag-independent target cell lysis. To address this issue, we grafted isolated CD8+ and CD4+ T cells from the peripheral blood with recombinant anti-carcinoembryonic Ag and anti-CD30 receptors, respectively. Cytotoxicity analyses revealed that grafted CD4+ T cells exert cytolysis of Ag-positive target cells with an efficiency similar to that of grafted CD8+ T cells. Lysis by receptor-grafted CD4+ T cells is Ag specific and is inhibited by blocking the target Ag or the Ag binding site of the recombinant receptor. Both Fas-sensitive and Fas-resistant target cells are lysed with equal efficiency, and lysis of Fas-sensitive target cells is not blocked by an anti-Fas ligand Ab, indicating that cytolysis by receptor-grafted CD4+ T cells is independent of the Fas pathway. We conclude that cytolysis by CD4+ T cells equipped with a recombinant immunoreceptor is MHC Ag and Fas independent and likely to be mediated by perforin present in receptor-grafted CD4+ T cells.
Costimulation Blockade, Busulfan, and Bone Marrow Promote Titratable Macrochimerism, Induce Transplantation Tolerance, and Correct Genetic Hemoglobinopathies with Minimal Myelosuppression
Authors: Adams Andrew B. , Durham Megan M. , Kean Leslie , Shirasugi Nozomu , Ha Jongwon , Williams Matthew A. , Rees Phyllis A. , Cheung Michael C. , Mittelstaedt Stephen , Bingaman Adam W. , Archer David R. , Pearson Thomas C. , Waller Edmund K. , Larsen Christian P.
Published: April 2014

Mixed hemopoietic chimerism has the potential to correct genetic hemological diseases (sickle cell anemia, thalassemia) and eliminate chronic immunosuppressive therapy following organ transplantation. To date, most strategies require either recipient conditioning (γ-irradiation, depletion of the peripheral immune system) or administration of “mega” doses of bone marrow to facilitate reliable engraftment. Although encouraging, many issues remain that may restrict or prevent clinical application of such strategies. We describe an alternative, nonirradiation based strategy using a single dose of busulfan, costimulation blockade, and T cell-depleted donor bone marrow, which promotes titratable macrochimerism and a reshaping of the T cell repertoire. Chimeras exhibit robust donor-specific tolerance, evidenced by acceptance of fully allogeneic skin grafts and failure to generate donor-specific proliferative responses in an in vivo graft-versus-host disease model of alloreactivity. In this model, donor cell infusion and costimulation blockade without busulfan were insufficient for tolerance induction as donor-specific IFN-γ-producing T cells re-emerged and skin grafts were rejected at ∼100 days. When applied to a murine β-thalassemia model, this approach allows for the normalization of hemologic parameters and replacement of the diseased red cell compartment. Such a protocol may allow for clinical application of mixed chimerism strategies in patients with end-stage organ disease or hemoglobinopathies.
Defective CD8+ T Cell Peripheral Tolerance in Nonobese Diabetic Mice
Authors: Kreuwel Huub T. C. , Biggs Judith A. , Pilip Ingrid M. , Pamer Eric G. , Lo David , Sherman Linda A.
Published: April 2014

Nonobese diabetic (NOD) mice develop spontaneous autoimmune diabetes that involves participation of both CD4+ and CD8+ T cells. Previous studies have demonstrated spontaneous reactivity to self-Ags within the CD4+ T cell compartment in this strain. Whether CD8+ T cells in NOD mice achieve and maintain tolerance to self-Ags has not previously been evaluated. To investigate this issue, we have assessed the extent of tolerance to a model pancreatic Ag, the hemagglutinin (HA) molecule of influenza virus, that is transgenically expressed by pancreatic islet β cells in InsHA mice. Previous studies have demonstrated that BALB/c and B10.D2 mice that express this transgene exhibit tolerance of HA and retain only low-avidity CD8+ T cells specific for the dominant peptide epitope of HA. In this study, we present data that demonstrate a deficiency in peripheral tolerance within the CD8+ T cell repertoire of NOD-InsHA mice. CD8+ T cells can be obtained from NOD-InsHA mice that exhibit high avidity for HA, as measured by tetramer (KdHA) binding and dose titration analysis. Significantly, these autoreactive CD8+ T cells can cause diabetes very rapidly upon adoptive transfer into NOD-InsHA recipient mice. The data presented demonstrate a retention in the repertoire of CD8+ T cells with high avidity for islet Ags that could contribute to autoimmune diabetes in NOD mice.
Cutting Edge: Differential Requirements for Stat4 in Expression of Glycosyltransferases Responsible for Selectin Ligand Formation in Th1 Cells
Authors: White Samuel J. , Underhill Gregory H. , Kaplan Mark H. , Kansas Geoffrey S.
Published: April 2014

A role for Stat4 in IL-12-induced up-regulation of selectin ligands on Th1 cells was explored. Th1 cells generated from Stat4−/− mice exhibited no IL-12-inducible P-selectin ligands, no up-regulation of core 2 β1,6-glucosaminyltransferase I (C2GlcNAcT-I), and low levels of the Th1 transcription factor T-bet. In contrast, Stat4−/− Th1 cells exhibited only a partial defect in expression of IL-12-inducible E-selectin ligands and expressed equivalently high levels of α1,3-fucosyltransferase VII (FucT-VII) as wild-type Th1 cells. FucT-VII expression was induced by T cell activation, and was enhanced by IL-12 independently of Stat4, whereas C2GlcNAcT-I up-regulation was mediated exclusively by IL-12, acting through Stat4. These data show that FucT-VII and C2GlcNAcT-I are controlled through distinct pathways and imply the existence of at least one other IL-12-inducible glycosyltransferase required for E-selectin and possibly P-selectin ligand formation in Th1 cells.
Cutting Edge: Integration of Human T Lymphocyte Cytoskeleton by the Cytolinker Plectin
Authors: Brown Martin J. , Hallam John A. , Liu Yin , Yamada Kenneth M. , Shaw Stephen
Published: April 2014

Chemokine-induced polarization of lymphocytes involves the rapid collapse of vimentin intermediate filaments (IFs) into an aggregate within the uropod. Little is known about the interactions of lymphocyte vimentin with other cytoskeletal elements. We demonstrate that human peripheral blood T lymphocytes express plectin, an IF-binding, cytoskeletal cross-linking protein. Plectin associates with a complex of structural proteins including vimentin, actin, fodrin, moesin, and lamin B in resting peripheral blood T lymphocytes. During chemokine-induced polarization, plectin redistributes to the uropod associated with vimentin and fodrin; their spatial distribution indicates that this vimentin-plectin-fodrin complex provides a continuous linkage from the nucleus (lamin B) to the cortical cytoskeleton. Overexpression of the plectin IF-binding domain in the T cell line Jurkat induces the perinuclear aggregation of vimentin IFs. Plectin is therefore likely to serve as an important organizer of the lymphocyte cytoskeleton and may regulate changes of lymphocyte cytoarchitecture during polarization and extravasation.
The Peyer’s Patch Microenvironment Suppresses T Cell Responses to Chemokines and Other Stimuli
Authors: Kellermann Sirid-Aimée , McEvoy Leslie M.
Published: April 2014

Immunosurveillance of mucosal sites presents immune cells with challenges not encountered in the periphery. T cells in the gut must distinguish enteric pathogens from innocuous non-self Ag derived from food or commensal bacteria. The mechanisms that regulate T cells in the gut remain incompletely understood. We assessed the effect of the Peyer’s patch microenvironment on T cell responses to chemokines. Chemokines are believed to play an important role during T cell priming by facilitating T cell migration into and within lymphoid tissues as well as T cell encounter and interaction with APCs. We found a profound suppression of chemokine-stimulated T cell chemotaxis and actin polymerization in Peyer’s patch relative to lymph node. Chemokine hyporesponsiveness is imposed upon T cells within hours of their entry into Peyer’s patches and is reversed following their removal. Suppression was not restricted to chemokine stimulation, as T cell responses to Con A and PMA were also suppressed. The global nature of this defect is further underscored by an impairment in calcium mobilization. Evidence indicates that a soluble factor contributes to this hyporesponsiveness, and comparison of Peyer’s patches and lymph nodes revealed striking differences in their chemokine and cytokine constitution, indicating a marked Th2 bias in the Peyer’s patches. The role of the Th2 microenvironment in mediating suppression is suggested by the ability of Nippostrongylus brasiliensis to elicit hyporesponsiveness in lymph node T cells. The suppressive milieu encountered by T cells in Peyer’s patches may be critical for discouraging undesired immune responses and promoting tolerance.
Enforced Expression of GATA-3 in Transgenic Mice Inhibits Th1 Differentiation and Induces the Formation of a T1/ST2-Expressing Th2-Committed T Cell Compartment In Vivo
Authors: Nawijn Martijn C. , Dingjan Gemma M. , Ferreira Rita , Lambrecht Bart N. , Karis Alar , Grosveld Frank , Savelkoul Huub , Hendriks Rudolf W.
Published: April 2014

The transcription factor GATA-3 is essential for early T cell development and differentiation of naive CD4+ T cells into Th2 effector cells. To study the function of GATA-3 during T cell-mediated immune responses in vivo, we investigated CD2-GATA3-transgenic mice in which GATA-3 expression is driven by the CD2 locus control region. Both in the CD4+ and the CD8+ T cell population the proportion of cells exhibiting a CD44highCD45RBlowCD62Llow Ag-experienced phenotype was increased. In CD2-GATA3-transgenic mice, large fractions of peripheral CD4+ T cells expressed the IL-1 receptor family member T1/ST2, indicative of advanced Th2 commitment. Upon in vitro T cell stimulation, the ability to produce IL-2 and IFN-γ was decreased. Moreover, CD4+ T cells manifested rapid secretion of the Th2 cytokines IL-4, IL-5, and IL-10, reminiscent of Th2 memory cells. In contrast to wild-type CD4+ cells, which lost GATA-3 expression when cultured under Th1-polarizing conditions, CD2-GATA3-transgenic CD4+ cells maintained expression of GATA-3 protein. Under Th1 conditions, cellular proliferation of CD2-GATA3-transgenic CD4+ cells was severely hampered, IFN-γ production was decreased and Th2 cytokine production was increased. Enforced GATA-3 expression inhibited Th1-mediated in vivo responses, such as Ag-specific IgG2a production or a delayed-type hypersensitivity response to keyhole limpet hemocyanin. Collectively, these observations indicate that enforced GATA-3 expression selectively inhibits Th1 differentiation and induces Th2 differentiation. The increased functional capacity to secrete Th2 cytokines, along with the increased expression of surface markers for Ag-experienced Th2-committed cells, would argue for a role of GATA-3 in Th2 memory formation.
IL-12 Decreases Activation-Induced Cell Death in Human Naive Th Cells Costimulated by Intercellular Adhesion Molecule-1. I. IL-12 Alters Caspase Processing and Inhibits Enzyme Function
Authors: Palmer Ellen M. , Farrokh-Siar Lili , Maguire van Seventer Jean , van Seventer Gijs A.
Published: April 2014

Th cells can receive costimulatory signals through the LFA-1/ICAM-1 accessory pathway that are sufficient to induce early Th cell proliferation, but not subsequent cell expansion and maintenance of cell viability. To investigate the regulatory role for IL-12 in ICAM-1-mediated costimulation, human naive Th cells were stimulated with coimmobilized anti-CD3 mAb and ICAM-1 Ig in the presence or absence of IL-12. The ICAM-1-mediated costimulatory signals in this model resulted in early Th cell proliferation followed by cell death that was partially mediated by Fas and involved loss of mitochondrial membrane potential, processing of procaspase-9 and -3, and activation of caspase-3. Addition of IL-12 prevented activation-induced cell death and promoted late proliferation. ICAM-1 + IL-12-costimulated Th cells were resistant to Fas-mediated cell death through a mechanism that did not appear to involve a decrease in either Fas or Fas ligand expression. IL-12 did not inhibit the loss of mitochondrial membrane potential induced by ICAM-1-mediated costimulation, and this finding was consistent with the inability of IL-12 to increase expression of the antiapoptotic Bcl-2 family members, Bcl-2 and Bcl-xL. Interestingly, IL-12 promoted an altered processing of procaspase-9 and -3 and a decrease in the percentage of cells displaying caspase-3 catalytic function. In conclusion, we now describe a novel regulatory function for IL-12 in preventing Th cell death and, as a result, in greatly increasing Th cell viability and expansion. Together, our findings indicate that IL-12 may perform this regulatory role by preventing Fas-mediated activation-induced cell death through inhibition of caspase-3 enzyme activity.
The Binding of Thymus Leukemia (TL) Antigen Tetramers to Normal Intestinal Intraepithelial Lymphocytes and Thymocytes
Authors: Tsujimura Kunio , Obata Yuichi , Matsudaira Yasue , Ozeki Satoshi , Yoshikawa Kazuhiro , Saga Shinsuke , Takahashi Toshitada
Published: April 2014

Thymus leukemia (TL) Ags belong to the family of nonclassical MHC class I Ags and can be recognized by both TCRαβ and TCRγδ CTL with TL, but not H-2 restriction. We previously reported that the CTL epitope is TAP independent, but the antigenic molecule(s) presented by TL has yet to be determined. In the present study, TL tetramers were prepared with T3b-TL and murine β2-microglobulin, not including antigenic peptides, and binding specificity was studied. CTL clones against TL Ags were stained with the T3b-TL tetramer, and the binding shown to be CD3 and CD8 dependent. Normal lymphocytes from various origins were also studied. Surprisingly, most CD8+ intraepithelial lymphocytes derived from the small intestines (iIEL), as well as CD8+ and CD4+CD8+ thymocytes, were stained, while only very minor populations of CD8+ cells derived from other peripheral lymphoid tissues, such as spleen and lymph nodes, were positive. The binding of T3b-TL tetramers to CD8+ iIEL and thymocytes was CD8 dependent, but CD3 independent, in contrast to that to TL-restricted CTL. These results altogether showed that TL-restricted CTL can be monitored by CD3-dependent binding of T3b-TL tetramers. In addition, CD3-independent T3b-TL tetramer binding to iIEL and thymocytes may imply that TL expressed on intestinal epithelium and cortical thymocytes has a physiological function interacting with these tetramer+CD8+ T lymphocytes.
Wnt Signaling Regulates Hemopoiesis Through Stromal Cells
Authors: Yamane Toshiyuki , Kunisada Takahiro , Tsukamoto Hirotake , Yamazaki Hidetoshi , Niwa Hitoshi , Takada Shinji , Hayashi Shin-Ichi
Published: April 2014

Hemopoietic cells develop in a complex milieu that is made up of diverse components, including stromal cells. Wnt genes, which are known to regulate the fate of the cells in a variety of tissues, are expressed in hemopoietic organs. However, their roles in hemopoiesis are not well characterized. In this study, we examined the roles of Wnt proteins in hemopoiesis using conditioned medium containing Wnt-3a. This conditioned medium dramatically reduced the production of B lineage cells and myeloid lineage cells, except for macrophages in the long-term bone marrow cultures grown on stromal cells, although the sensitivity to the conditioned medium differed, depending on the hemopoietic lineage. In contrast, the same conditioned medium did not affect the generation of B lineage or myeloid lineage cells in stromal cell-free conditions. These results suggested that Wnt proteins exert their effects through stromal cells. Indeed, these effects were mimicked by the expression of a stabilized form of β-catenin in stromal cells. In this study, we demonstrated that Wnt signaling regulates hemopoiesis through stromal cells with selectivity and different degrees of the effect, depending on the hemopoietic lineage in the hemopoietic microenvironment.
Regulation of TGF-β Response During T Cell Activation Is Modulated by IL-10
Authors: Cottrez Françoise , Groux Hervé
Published: April 2014

TGF-β1 is an important pleiotropic cytokine that has been described to have both stimulatory and inhibitory effects on cell growth and differentiation. For several cell types, the effect of TGF-β1 was found to correlate with the differentiation stage of the cells and the presence of other cytokines. In this report, we address the influence of TGF-β1 on CD4+ T cell activation by evaluating the effect of TGF-β1 on the proliferative and cytokine responses of purified resting and activated human or mouse CD4+ T cells. TGF-β1 inhibits proliferation and cytokine secretion on resting CD4+ T cells but has no inhibitory effect on activated T cells. Moreover, TGF-β1 unresponsiveness of activated T cells was correlated with a down-regulation in the expression of the TGF-β receptor type II. Interestingly, IL-10 addition enhances TGF-β receptor type II expression and restores TGF-β responsiveness on activated T cells. These results indicated that TGF-β responsiveness is sequentially regulated on T cells by the modulation of the of TGF-β receptor type II chain expression. Moreover, we have identified a novel regulatory role of IL-10 on TGF-β-dependent T cell growth that can explain the control of T cell activation on chronic vs acute inflammatory sites.
A Role for IL-12 Receptor Expression and Signal Transduction in Host Defense in Leprosy
Authors: Kim Jenny , Uyemura Koichi , Van Dyke Melissa K. , Legaspi Annaliza J. , Rea Thomas H. , Shuai Ke , Modlin Robert L.
Published: April 2014

The generation of cell-mediated immunity against intracellular infection involves the production of IL-12, a critical cytokine required for the development of Th1 responses. The biologic activities of IL-12 are mediated through a specific, high affinity IL-12R composed of an IL-12Rβ1/IL-12Rβ2 heterodimer, with the IL-12Rβ2 chain involved in signaling via Stat4. We investigated IL-12R expression and function in human infectious disease, using the clinical/immunologic spectrum of leprosy as a model. T cells from tuberculoid patients, the resistant form of leprosy, are responsive to IL-12; however, T cells from lepromatous patients, the susceptible form of leprosy, do not respond to IL-12. We found that the IL-12Rβ2 was more highly expressed in tuberculoid lesions compared with lepromatous lesions. In contrast, IL-12Rβ1 expression was similar in both tuberculoid and lepromatous lesions. The expression of IL-12Rβ2 on T cells was up-regulated by Mycobacterium leprae in tuberculoid but not in lepromatous patients. Furthermore, IL-12 induced Stat4 phosphorylation and DNA binding in M. leprae-activated T cells from tuberculoid but not from lepromatous patients. Interestingly, IL-12Rβ2 in lepromatous patients could be up-regulated by stimulation with M. tuberculosis. These data suggest that Th response to M. leprae determines IL-12Rβ2 expression and function in host defense in leprosy.
Engagement of FcεRI on Human Monocytes Induces the Production of IL-10 and Prevents Their Differentiation in Dendritic Cells
Authors: Novak Natalija , Bieber Thomas , Katoh Norito
Published: April 2014

The local cytokine environment and the presence of stimulatory signals determine whether circulating monocytes will finally acquire characteristics of dendritic cells (DCs) or macrophages. Because FcεRI expressed on professional APCs, e.g., monocytes and DCs, has been suggested to play a key role in the pathophysiology of atopic diseases, we evaluated the effect of receptor ligation on the generation of monocyte-derived DCs (MoDCs). Aggregation of FcεRI at the initiation of the IL-4-GM-CSF-driven differentiation resulted in the emergence of macrophage-like cells with a strong expression of the mannose receptor and a low level of CD1a and the DC-specific markers CD83 and the actin-bundling protein (p55). These cells sustained the ability to take up FITC-labeled Escherichia coli by phagocytosis and were significantly less efficient in stimulating purified allogeneic T cells. In addition, receptor ligation of FcεRI at the beginning of the culture prevented the generation of MoDCs, mainly due to a dramatic increase in the IL-10 production. These results suggest that FcεRI aggregation prevents the generation of CD1a+ MoDCs and imply a novel pivotal function of this receptor in modulating the differentiation of monocytes.
Peripheral T Cells Become Sensitive to Glucocorticoid- and Stress-Induced Apoptosis in Transgenic Mice Overexpressing SRG3
Authors: Han Sunmi , Choi Heonsik , Ko Myung-gon , Choi Young I. , Sohn Dong H. , Kim Joong K. , Shin Dongho , Chung Heekyoung , Lee Han W. , Kim Jae-B. , Park Sang D. , Seong Rho H.
Published: April 2014

Immature double-positive thymocytes are sensitive to glucocorticoid (GC)-induced apoptosis, whereas mature single-positive T cells are relatively resistant. Thymocytes seem to acquire resistance to GCs during differentiation into mature single-positive thymocytes. However, detailed knowledge concerning what determines the sensitivity of thymocytes to GCs and how GC sensitivity is regulated in thymocytes during development is lacking. We have previously reported that the murine SRG3 gene (for SWI3-related gene) is required for GC-induced apoptosis in a thymoma cell line. Herein, we provide results suggesting that the expression level of SRG3 protein determines the GC sensitivity of T cells in mice. SRG3 associates with the GC receptor in the thymus, but rarely in the periphery. Transgenic overexpression of the SRG3 protein in peripheral T cells induces the formation of the complex and renders the cells sensitive to GC-induced apoptosis. Our results also show that blocking the formation of the SRG3-GC receptor complex with a dominant negative mutant form of SRG3 decreases GC sensitivity in thymoma cells. In addition, mice overexpressing the SRG3 protein appear to be much more susceptible to stress-induced deletion of peripheral T cells than normal mice, which may result in an immunosuppressive state in an animal.
Monomeric Class I Molecules Mediate TCR/CD3ε/CD8 Interaction on the Surface of T Cells
Authors: Block Matthew S. , Johnson Aaron J. , Mendez-Fernandez Yanice , Pease Larry R.
Published: April 2014

Both CD8 and the TCR bind to MHC class I molecules during physiologic T cell activation. It has been shown that for optimal T cell activation to occur, CD8 must be able to bind the same class I molecule that is bound by the TCR. However, no direct evidence for the class I-dependent association of CD8 and the TCR has been demonstrated. Using fluorescence resonance energy transfer, we show directly that a single class I molecule causes TCR/CD8 interaction by serving as a docking molecule for both CD8 and the TCR. Furthermore, we show that CD3ε is brought into close proximity with CD8 upon TCR/CD8 association. These interactions are not dependent on the phosphorylation events characteristic of T cell activation. Thus, MHC class I molecules, by binding to both CD8 and the TCR, mediate the reorganization of T cell membrane components to promote cellular activation.
Signal Transduction by Human-Restricted FcγRIIa Involves Three Distinct Cytoplasmic Kinase Families Leading to Phagocytosis
Authors: Cooney Damon S. , Phee Hyewon , Jacob Anand , Coggeshall K. Mark
Published: April 2014

Recent experiments indicate an important role for Src family and Syk protein tyrosine kinases and phosphatidylinositol 3-kinase in the signal transduction process initiated by mouse receptors for IgG and leading to phagocytosis. Considerably less is known regarding signal transduction by the human-restricted IgG receptor, FcγRIIa. Furthermore, the relationship among the Src family, Syk, and phosphatidylinositol 3-kinase in phagocytosis is not understood. Here, we show that FcγRIIa is phosphorylated by an Src family member, which results in recruitment and concomitant activation of the distal enzymes Syk and phosphatidylinositol 3-kinase. Using a FcγRI-p85 receptor chimera cotransfected with kinase-inactive mutants of Syk or application of a pharmacological inhibitor of Syk, we show that Syk acts in parallel with phosphatidylinositol 3-kinase. Our results indicate that FcγRIIa-initiated monocyte or neutrophil phagocytosis proceeds from the clustered IgG receptor to Src to phosphatidylinositol 3-kinase and Syk.
Chromatin Remodeling at the Ig Loci Prior to V(D)J Recombination
Authors: Maës Jérôme , O’Neill Laura P. , Cavelier Patricia , Turner Bryan M. , Rougeon François , Goodhardt Michele
Published: April 2014

Rearrangement of Ig H and L chain genes is highly regulated and takes place sequentially during B cell development. Several lines of evidence indicate that chromatin may modulate accessibility of the Ig loci for V(D)J recombination. In this study, we show that remodeling of V and J segment chromatin occurs before V(D)J recombination at the endogenous H and κ L chain loci. In recombination-activating gene-deficient pro-B cells, there is a reorganization of nucleosomal structure over the H chain JH cluster and increased DNase I sensitivity of VH and JH segments. The pro-B/pre-B cell transition is marked by a decrease in the DNase I sensitivity of VH segments and a reciprocal increase in the nuclease sensitivity of Vκ and Jκ segments. In contrast, JH segments remain DNase I sensitive, and their nucleosomal organization is maintained in μ+ recombination-activating gene-deficient pre-B cells. These results indicate that initiation of rearrangement is associated with changes in the chromatin structure of both V and J segments, whereas stopping recombination involves changes in only V segment chromatin. We further find an increase in histone H4 acetylation at both the H and κ L chain loci at the pro-B cell stage. Although histone H4 acetylation appears to be an early change associated with B cell commitment, acetylation alone is not sufficient to promote subsequent modifications in Ig chromatin.
Regulation of MHC Class II Antigen Presentation by Sorting of Recycling HLA-DM/DO and Class II within the Multivesicular Body
Authors: van Lith Marcel , van Ham Marieke , Griekspoor Alexander , Tjin Esther , Verwoerd Desiree , Calafat Jero , Janssen Hans , Reits Eric , Pastoors Liesbeth , Neefjes Jacques
Published: April 2014

MHC class II molecules bind antigenic peptides in the late endosomal/lysosomal MHC class II compartments (MIIC) before cell surface presentation. The class II modulatory molecules HLA-DM and HLA-DO mainly localize to the MIICs. Here we show that DM/DO complexes continuously recycle between the plasma membrane and the lysosomal MIICs. Like DMβ and the class II-associated invariant chain, the DOβ cytoplasmic tail contains potential lysosomal targeting signals. The DOβ signals, however, are not essential for internalization of the DM/DO complex from the plasma membrane or targeting to the MIICs. Instead, the DOβ tail determines the distribution of both DM/DO and class II within the multivesicular MIIC by preferentially localizing them to the limiting membrane and, in lesser amounts, to the internal membranes. This distribution augments the efficiency of class II antigenic peptide loading by affecting the efficacy of lateral interaction between DM/DO and class II molecules. Sorting of DM/DO and class II molecules to specific localizations within the MIIC represents a novel way of regulating MHC class II Ag presentation.
Toll-Like Receptor 2-Dependent Inhibition of Macrophage Class II MHC Expression and Antigen Processing by 19-kDa Lipoprotein ofMycobacterium tuberculosis
Authors: Noss Erika H. , Pai Rish K. , Sellati Timothy J. , Radolf Justin D. , Belisle John , Golenbock Douglas T. , Boom W. Henry , Harding Clifford V.
Published: April 2014

Mycobacterium tuberculosis (MTB) induces vigorous immune responses, yet persists inside macrophages, evading host immunity. MTB bacilli or lysate was found to inhibit macrophage expression of class II MHC (MHC-II) molecules and MHC-II Ag processing. This report characterizes and identifies a specific component of MTB that mediates these inhibitory effects. The inhibitor was extracted from MTB lysate with Triton X-114, isolated by gel electroelution, and identified with Abs to be MTB 19-kDa lipoprotein. Electroelution- or immunoaffinity-purified MTB 19-kDa lipoprotein inhibited MHC-II expression and processing of both soluble Ags and Ag 85B from intact MTB bacilli. Inhibition of MHC-II Ag processing by either MTB bacilli or purified MTB 19-kDa lipoprotein was dependent on Toll-like receptor (TLR) 2 and independent of TLR 4. Synthetic analogs of lipopeptides from Treponema pallidum also inhibited Ag processing. Despite the ability of MTB 19-kDa lipoprotein to activate microbicidal and innate immune functions early in infection, TLR 2-dependent inhibition of MHC-II expression and Ag processing by MTB 19-kDa lipoprotein during later phases of macrophage infection may prevent presentation of MTB Ags and decrease recognition by T cells. This mechanism may allow intracellular MTB to evade immune surveillance and maintain chronic infection.
Negative Selection of T Cells byHelicobacter pylorias a Model for Bacterial Strain Selection by Immune Evasion
Authors: Wang Jide , Brooks Edward G. , Bamford Kathleen B. , Denning Timothy L. , Pappo Jacques , Ernst Peter B.
Published: April 2014

The majority of humans infected with Helicobacter pylori maintain a lifelong infection with strains bearing the cag pathogenicity island (PAI). H. pylori inhibits T cell responses and evades immunity so the mechanism by which infection impairs responsiveness was investigated. H. pylori caused apoptotic T cell death, whereas Campylobacter jejuni did not. The induction of apoptosis by H. pylori was blocked by an anti-Fas Ab (ZB4) or a caspase 8 inhibitor. In addition, a T cell line with the Fas rendered nonfunctional by a frame shift mutation was resistant to H. pylori-induced death. H. pylori strains bearing the cag PAI preferentially induced the expression of Fas ligand (FasL) on T cells and T cell death, whereas isogenic mutants lacking these genes did not. Inhibiting protein synthesis blocked FasL expression and apoptosis of T cells. Preventing the cleavage of FasL with a metalloproteinase inhibitor increased H. pylori-mediated killing. Thus, H. pylori induced apoptosis in Fas-bearing T cells through the induction of FasL expression. Moreover, this effect was linked to bacterial products encoded by the cag PAI, suggesting that persistent infection with this strain may be favored through the negative selection of T cells encountering specific H. pylori Ags.
Modulation of Macrophage Cytokine Production by ES-62, a Secreted Product of the Filarial NematodeAcanthocheilonema viteae
Authors: Goodridge Helen S. , Wilson Emma H. , Harnett William , Campbell Carol C. , Harnett Margaret M. , Liew Foo Y.
Published: April 2014

Parasite survival and host health may depend on the ability of the parasite to modulate the host immune response by the release of immunomodulatory molecules. Excretory-secretory (ES)-62, one such well-defined molecule, is a major secreted protein of the rodent filarial nematode Acanthocheilonema viteae, and has homologues in human filarial nematodes. Previously we have shown that ES-62 is exclusively associated with a Th2 Ab response in mice. Here we provide a rationale for this polarized immune response by showing that the parasite molecule suppresses the IFN-γ/LPS-induced production, by macrophages, of bioactive IL-12 (p70), a key cytokine in the development of Th1 responses. This suppression of the induction of a component of the host immune response extends to the production of the proinflammatory cytokines IL-6 and TNF-α, but not NO. The molecular mechanism underlying these findings awaits elucidation but, intriguingly, the initial response of macrophages to ES-62 is to demonstrate a low and transient release of these cytokines before becoming refractory to further release induced by IFN-γ/LPS. The relevance of our observations is underscored by the finding that macrophages recovered from mice exposed to “physiological” levels of ES-62 by the novel approach of continuous release from implanted osmotic pumps in vivo were similarly refractory to release of IL-12, TNF-α, IL-6, but not NO, ex vivo. Therefore, our results suggest that exposure to ES-62 renders macrophages subsequently unable to produce Th1/proinflammatory cytokines. This likely contributes to the generation of immune responses with an anti-inflammatory Th2 phenotype, a well-documented feature of filarial nematode infection.
Inhibition of Endotoxin-Induced Macrophage Chemokine Production by Vasoactive Intestinal Peptide and Pituitary Adenylate Cyclase-Activating Polypeptide In Vitro and In Vivo
Authors: Delgado Mario , Ganea Doina
Published: April 2014

Inflammatory chemokines recruit various populations of immune cells that initiate and maintain the inflammatory response against foreign Ags. Although such a response is necessary for the elimination of the Ag, the inflammation has to be eventually resolved in a healthy organism. Neuropeptides such as vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP), released after antigenic stimulation, contribute to the termination of an inflammatory response primarily by inhibiting the production of proinflammatory cytokines. Here we investigated the effects of VIP and PACAP on chemokine production. We report that VIP and PACAP inhibit the expression of the macrophage-derived CXC chemokines macrophage inflammatory protein-2 and KC (IL-8), and of the CC chemokines MIP-1α, MIP-1β, monocyte chemoattractant protein 1, and RANTES in vivo and in vitro. The inhibition of chemokine gene expression correlates with an inhibitory effect of VIP/PACAP on NF-κB binding and transactivating activity. The VIP/PACAP inhibition of both chemokine production and of NF-κB binding and transactivating activity is mediated through the specific VIP receptor VPAC1, and involves both cAMP-dependent and -independent intracellular pathways. In an in vivo model of acute peritonitis, the inhibition of chemokine production by VIP/PACAP leads to a significant reduction in the recruitment of polymorphonuclear cells, macrophages, and lymphocytes into the peritoneal cavity. These findings support the proposed role of VIP and PACAP as key endogenous anti-inflammatory agents and describe a novel mechanism, i.e., the inhibition of the production of macrophage-derived chemokines.
Cooperation of Toll-Like Receptor 2 and 6 for Cellular Activation by Soluble Tuberculosis Factor andBorrelia burgdorferiOuter Surface Protein A Lipoprotein: Role of Toll-Interacting Protein and IL-1 Receptor Signaling Molecules in Toll-Like Receptor 2 Signaling
Authors: Bulut Yonca , Faure Emmanuelle , Thomas Lisa , Equils Ozlem , Arditi Moshe
Published: April 2014

Toll-like receptor 2 (TLR2) and TLR4 play important roles in innate immune responses to various microbial agents. We have previously shown that human dermal endothelial cells (HMEC) express TLR4, but very little TLR2, and respond to LPS, but not to Mycobacterium tuberculosis 19-kDa lipoprotein, unless transfected with TLR2. Here we report that HMEC are unresponsive to several additional biologically relevant TLR2 ligands, including, phenol-soluble modulin (PSM), a complex of three small secreted polypeptides from the skin commensal Staphylococcus epidermidis, soluble tuberculosis factor (STF), and Borrelia burgdorferi outer surface protein A lipoprotein (OspA-L). Expression of TLR2 renders HMEC responsive to all these ligands. We further characterized the signaling pathway in response to STF, OspA-L, and PSM in TLR2-transfected HMEC. The TLR2 signaling pathway for NF-κB trans-activation shares the IL-1R signaling molecules. Dominant negative constructs of TLR2 or TLR6 inhibit the responses of STF and OspA-L as well as PSM in TLR2-transfected HMEC, supporting the concept of functional cooperation between TLR2 and TLR6 for all these TLR2 ligands. Moreover, we show that Toll-interacting protein (Tollip) coimmunoprecipitates with TLR2 and TLR4 using HEK 293 cells, and overexpression of Tollip inhibits NF-κB activation in response to TLR2 and TLR4 signaling. Collectively, these findings suggest that there is functional interaction between TLR2 and TLR6 in the cellular response to STF and OspA-L in addition to S. epidermidis (PSM) Ags, and that engagement of TLR2 triggers a signaling cascade, which shares the IL-1R signaling molecules, similar to the TLR4-LPS signaling cascade. Our data also suggest that Tollip may be an important constituent of both the TLR2 and TLR4 signaling pathways.
Human Eosinophils and Human High Affinity IgE Receptor Transgenic Mouse Eosinophils Express Low Levels of High Affinity IgE Receptor, but Release IL-10 upon Receptor Activation
Authors: Kayaba Hiroyuki , Dombrowicz David , Woerly Gaetane , Papin Jean-Paul , Loiseau Sylvie , Capron Monique
Published: April 2014

FcεRI expressed by human eosinophils is involved in IgE-mediated cytotoxicity reactions toward the parasite Schistosoma mansoni in vitro. However, because receptor expression is low on these cells, its functional role is still controversial. In this study, we have measured surface and intracellular expression of FcεRI by blood eosinophils from hypereosinophilic patients and normal donors. The number of unoccupied receptors corresponded to ∼4,500 Ab binding sites per cell, whereas 50,000 Ab binding sites per cell were detected intracellularly. Eosinophils from patients displayed significantly more unoccupied receptors than cells from normal donors. This number correlated to both serum IgE concentrations and to membrane-bound IgE. The lack of FcεRI expression by mouse eosinophils has hampered further studies. To overcome this fact and experimentally confirm our findings on human eosinophils, we engineered IL-5 × hFcεRIα double-transgenic mice, whose bone marrow, blood, spleen, and peritoneal eosinophils expressed FcεRI levels similar to levels of human eosinophils, after 4 days culture with IgE in the presence of IL-5. Both human and mouse eosinophils were able to secrete IL-10 upon FcεRI engagement. Thus, comparative analysis of cells from patients and from a relevant animal model allowed us to clearly demonstrate that FcεRI-mediated eosinophil activation leads to IL-10 secretion. Through FcεRI expression, these cells are able to contribute to both the regulation of the immune response and to its effector mechanisms.
Cutting Edge: Control of CD8+ T Cell Activation by CD4+CD25+ Immunoregulatory Cells
Authors: Piccirillo Ciriaco A. , Shevach Ethan M.
Published: April 2014

CD4+CD25+ regulatory T cells inhibit organ-specific autoimmune diseases induced by CD4+CD25− T cells and are potent suppressors of CD4+CD25− T cell activation in vitro. We demonstrate that CD4+CD25+ T cells also suppress both proliferation and IFN-γ production by CD8+ T cells induced either by polyclonal or Ag-specific stimuli. CD4+CD25+ T cells inhibit the activation of CD8+ responders by inhibiting both IL-2 production and up-regulation of IL-2Rα-chain (CD25) expression. Suppression is mediated via a T-T interaction as activated CD4+CD25+ T cells suppress the responses of TCR-transgenic CD8+ T cells stimulated with soluble peptide-MHC class I tetramers in the complete absence of APC. These results broaden the immunoregulatory role played by CD4+CD25+ T cells in the prevention of autoimmune diseases, but also raise the possibility that they may hinder the induction of effector CD8+ T cells to tumor or foreign Ags.