Aca 2013 Abstract Export Plenary Session: Plenary Session 1: Novel aspect of therapeutics of autoimmune diseases
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- Free Communication: Free Communications 2- Pathogenetic and Immunoregulatory Mechanisms in Autoimmunity
- 078 POSTTRANSCRIPTIONAL GENE REGULATION OF CD4+ TH17 DIFFERENTIATION BY THE RNA-BINDING PROTEIN HUR
- 079 EXPRESSION OF INFLAMMASOME IS DIFFERENT IN MUSCLE OF DERMATOMYOSITIS AND POLYMYOSITIS PATIENTS
- 080 CD98HC REGULATES THE DEVELOPMENT OF EXPERIMENTAL COLITIS BY DIFFERENTIALLY CONTROLLING INDUCIBLE AND NATURALLY OCCURING REGULATORY T CELLS
- 082 TYROSINE KINASE BTK IS REQUIRED FOR NK CELL-MEDIATED AUTOIMMUNE HEPATITIS
- 083 NEURO-VASCULAR DISRUPTION IN THE CEREBRAL CORTEX DURING EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS AND THE EFFECT OF GLATIRAMER ACETATE
- 084 MYCOPLASMA SUPERANTIGEN AS A TRIGGER PROMOTES AUTOREACTIVE T CELL RESPONSES AND DISEASE SEVERITY IN AUTOIMMUNE DIABETIC MOUSE
- 085 UP-REGULATION OF MICRORNA-210 INDUCES IMMUNE DYSFUNCTION VIA TARGETING FOXP3 IN CD4+ T CELLS OF PSORIASIS VULGARIS
- 086 INTERLEUKIN 27 INHIBITS ATHEROSCLEROSIS VIA IMMUNOREGULATION OF MACROPHAGES IN MICE
- 087 B-LYMPHOCYTE SUBSETS IN CHRONIC FATIGUE SYNDROME
Methods and results: DV was expressed in the Pichia pastoris (P. pastoris) (P.rβ2-GPI DV) expression system, and used in binding assays. Enzyme-linked immunoassay (ELISA) demonstrated that P.rβ2-GPI DV interacted with oxLDL via 7-keocholestryl-9-carboxynonanoate (oxLig-1), a negatively charged lipid moiety from oxLDL. It was also shown that free ω-carboxyl residue of oxLig-1 was required for this interaction. Serologic tests also showed elevated levels of oxLDL/β2-GPI complexes in patients with APS. Moreover, P.rβ2-GPI DV bound oxLDL with a high affinity in APS sera, and that the addition of P.rβ2-GPI DV inhibited the formation of circulating oxLDL/β2-GPI complexes. Conclusions: Recombinant P.rβ2-GPI DV was capable of binding to oxLDL, and this binding could effectively inhibit the formation of oxLDL/β2-GPI complexes in the sera of the APS patients, suggesting that P.rβ2-GPI DV could potentially be a new candidate compound for autoimmune disease therapy. Free Communication: Free Communications 2- Pathogenetic and Immunoregulatory Mechanisms in Autoimmunity 077 ROLES OF TOLL-LIKE RECEPTORS IN GENE/ENVIRONMENT INTERACTIONS IN A MOUSE MODEL OF MULTIPLE SCLEROSIS S. Miranda-Hernandez1, S. Chowdhury1, M. Jordan1, A.G. Baxter1 1Comparative Genomics Centre, James Cook University, Townsville, Australia The most commonly used animal model for the study of Multiple Sclerosis (MS) is Experimental Autoimmune Encephalomyelitis (EAE). EAE causes a demyelinating paralysis resembling some forms of Multiple sclerosis (MS). Previously we found that C57BL/6.Tlr2-/- mice decrease the severity of EAE. In addition, transfer of EAE into C57BL/6.Tlr2-/- mice failed to transfer disease. This protection from EAE was associated with fewer infiltrating CD4+ T cells in the CNS, reduced prevalence of detectable circulating IL-6, and increased proportions of central (CD62L+) Foxp3+ Tregs. As these results indicate that TLR signalling affects CNS-autoimmunity, in this study we examined the role of TLRs in NOD mice, which are susceptible to Type 1 Diabetes (T1D) in order to dissect the interrelationships between organ-specific autoimmune diseases. NOD/Lt mice develop a mild form of relapsing/remitting EAE. In contrast to C57BL/6 mice, TLR2 deletion in NOD mice did not affect the severity of disease. We tested whether insulitis associated with T1D development compensated for the lack of TLR2 signalling in these mice. The course of EAE was analyzed in NOD.H2b and NOD.H2d mice, which do not develop insulitis or T1D. NOD.H2b mice showed increased severity of EAE compared to NOD/Lt mice, again in a relapsing/remitting pattern. In the absence of insulitis, EAE was ameliorated in NOD.H2b.Tlr2-/- mice and no relapses occurred after the initial peak of disease. These data suggest that both TLR2 signalling and immunological events associated with autoimmunity at a distant site can mediate relapses in CNS autoimmunity.
Interleukin 17 (IL-17A) is a proinflammatory cytokine produced by activated CD4+ Th17. Th17 cells are major contributors to autoimmune diseases, including multiple sclerosis. Although the transcriptional control of IL-17 and Th17 differentiation have been well studied, posttranscriptional gene regulation is unclear. The RNA-binding protein (RBP) HuR regulates the stability of many target mRNAs via binding the AU-rich elements (ARE) present in the 3Õ untranslated region (UTR), of many pro-inflammatory cytokines such as IL-4, IL-13 and TNF. Since IL-17 mRNA also possesses HuR binding motifs, we investigated whether HuR can regulate Il-17 expression. We demonstrated that HuR directly binds IL-17 mRNA 3Õ UTR by using RNA immunoprecipitation methods as well as biotin pull down assays. CD4+ T cells from HuRfl/fl KO mice have decreases in Th17 differentiation, IL-17 steady-state mRNA and protein expression compared to wild type Th17 cells. IL-17 mRNA stability was decreased by 3-fold in HuR KO mice. We used the experimental autoimmune encephalitis (EAE) model of neuroinflammation to investigate whether HuR plays a role in CD4+ Th17 differentiation and disease initiation. Mice with adoptive transfer of HuR KO CD4+ T cells had delayed onset, reduced severity of EAE symptoms, as well as reduced CNS inflammation and decreased IL-17 production by Th17-producing cells recovered from the CNS. Our results reveal HuR-controlled posttranscriptional regulatory mechanisms of Th17 differentiation and direct effects upon EAE initiation. These findings could provide novel therapeutic targets for treatment of IL-17- mediated autoimmune neuroinflammation and also potentially other diseases in which IL-17 plays a major role. 079 EXPRESSION OF INFLAMMASOME IS DIFFERENT IN MUSCLE OF DERMATOMYOSITIS AND POLYMYOSITIS PATIENTS Q.Z. Zhao1, Y.L. Zhang1, H. Zhou1, X.M. Shu1, X. Lu1, G.C. Wang1 1Rheumatology, China-Japan Friendship Hospital, Beijing, China Background and aims Activation of innate immune plays important role in the pathogenesis of autoimmune disease. Our study was to investigate the expression of inflammasomes in muscle of dermatomyositis(DM) and polymyositis(PM) patients. Methods Immunohistochemistry was performed to determine the expression of NALP (NACHT-LRR-PYD-containing protein)1, NALP3, ASC (apoptosis-associated speck-like protein containing a CARD) and caspase-1 in muscle of 10 newly diagnosed and untreated adult DM/PM patients (4 DM and 6 PM) and 5 healthy controls. Results In muscle of 4 DM patients, ASC and caspase-1 were expressed in the plasma of CD3+ T and CD20+ B cells, while in 2 of them NALP3 was also expressed. Only 1 PM patient in 6 expressed NALP3, ASC and caspase-1 in the plasma of CD3+ T and CD20+ B cells. NALP1 was not expressed in all of the 10 patients. No NALP1, NALP3, ASC and caspase-1 were expressed in the muscle of 5 healthy controls. The positive rate of ASC and caspase-1 in DM group was significantly higher than that in PM and control group (P<0.01 and P<0.05), while the positive rate of NALP3 in DM group had no difference with that in PM and control group. The level of creatine kinase, erythrocyte sedimentation rate, C-reactive protein had no difference between NALP3, ASC and caspase-1 positive group and negative group. Conclusions Innate immune plays different role in the pathogenesis of DM and PM. Inflammasome participates in the inflammation of DM but not in PM, and may be activated by NALP3. 080 CD98HC REGULATES THE DEVELOPMENT OF EXPERIMENTAL COLITIS BY DIFFERENTIALLY CONTROLLING INDUCIBLE AND NATURALLY OCCURING REGULATORY T CELLS Z. Bhuyan1, H. Arimochi1, K. Yasutomo1 1Immunology and Parasitology, Univ of Tokushima Grad Sch of Health Biosciences, Tokushima, Japan CD4+ T cell activation is controlled by signaling through the T cell receptor in addition to various co-receptors, and is also affected by their interactions with effector and regulatory T cells in the microenvironment. Inflammatory bowel diseases (IBD) are caused by the persistent activation and expansion of auto-aggressive CD4+ T cells that attack intestinal epithelial cells. However, the molecular basis for the persistent activation of CD4+ T cells in IBD remains unclear. In this study, we investigated how the CD98 heavy chain (CD98hc, Slc3a2) affected the development of colitis in an experimental animal model. Transferring CD98hc-deficient CD4+CD25- T cells into Rag2-/- mice skewed CD4+ T cells toward Foxp3+ inducible regulatory T cells (iTregs), rather than their differentiation into Th1 or Th17 cells, and did not induce colitis. By comparison, CD98hc-deficient naturally occurring regulatory T cells (nTregs) had a decreased capability to suppress colitis induced by CD4+CD25- T cells, although CD98hc-deficient mice did not have a defect in the development of nTregs. Blocking CD98hc with an anti-CD98 blocking antibody prevented the development of colitis. Our results suggest that CD98hc has distinct roles in iTregs and nTregs in vivo during the progression of colitis and also suggest that CD98hc on CD4+ T cells contributes to immune dysregulation in autoimmune colitis. 081 C-FLIP OVEREXPRESSION IS ASSOCIATED WITH PARANEOPLASTIC MYASTHENIA GRAVIS (MG) AND LATE-ONSET MG (LOMG) D. Belharazem1, D. Belharazem1, A. Grass1, B. Schalke2, A. Marx1 1Pathology, University Hospital Mannheim, Mannheim, Germany 2Neurology, University Hospital Regensburg, Mannheim, Germany Tumors of the thymus especially lymphoepithelial thymomas are 30%associated with the autoimmune Myasthenia gravis (MG). The auto reactive T lymphocytes generated in thymus tumors stimulate antibodies production against several muscle antigens, thereby acquiring a heterogeneous neuromuscular junction disease characterized by muscular weakness. Antiapoptotic proteins overexpression is one of the tumor-promoting mechanism. We ask whether the antiapoptotic molecule cFlip may be operative in thymoma and thymoma associated MG and in LOMG. We investigated 87 thymomas (10A, 20AB, 20B2. 15B3 and 22 thymus carcinomas (TC)) without MG, 21 thymomas associated MG(8AB, 8B2 and 5B3) and 13 thymic hyperplasia from late-onset MG (LOMG) for the expression of cFlip using real time RT-PCR and western blotting for cryo-preserved material and immunohistochemistry (IHC) for formalin-fixed, paraffin-embedded tissues. cFlip expression was also performed in peripheral blood lymphocytes from LOMG, thymomas associated and not associated MG and compared to lymphocytes from donors. An overexpression was detected in all thymomas tissues without MG and in thymus carcinomas (p<0.0001****). In paraneoplastic MG cFlip mRNA and proteins Level were higher expressed (p<0.0001****).The non paraneoplastic MG cases showed also a significant cFlip mRNA transcription compared to normal thymi. In addition cFlip was highly detectable in paraneoplastic and non paraneoplastic MG Lymphocytes. The anti-apoptotic protein cFlip is high expressed on the transcriptional and proteomic level in thymomas especially when they are associated with MG. This suggests that cFlip might play a role in the pathogenesis of paraneoplastic MG deserves future studies for its regulation as a new therapeutic target. 082 TYROSINE KINASE BTK IS REQUIRED FOR NK CELL-MEDIATED AUTOIMMUNE HEPATITIS Y. Bao1, J. Zheng2, C. Han3 1Translational Medicine Center, Shanghai Changzheng Hospital Second Military Medical University, Shanghai, China 2Department of Paediatrics and Adolescent Medicine, Li Ka Shing Faculty of Medicine the University of Hong Kong, Hong Kong, China 3National Key Laboratory of Medical Immunology & Institute of Immunology, Second Military Medical University, Shanghai, China BrutonÕs tyrosine kinase (Btk) is not only critical for B cell development and differentiation, but also involved in the regulation of Toll-like receptor (TLR)-triggered innate response of macrophages. However, whether Btk is involved in the regulation of NK cell innate function remains unknown. Through FACS and immunoblot, we show that Btk expression is upregulated during maturation and activation of mouse NK cells. Murine Btk-/- NK cells have decreased innate immune responses to TLR3 ligand, with reduced expressions of IFN-γ, perforin and Granzyme-B, and decreased cytotoxic activity. Furthermore, Btk is found to promote TLR3-triggered NK cell activation by activating NF-κB pathway. Poly I:C-induced, NK cell-mediated acute hepatitis is observed to be attenuated in Btk-/- mice or the mice with in vivo administration of Btk inhibitor. Accordingly, liver damage is aggravated in Btk-/- mice once adoptive transfer of Btk+/+ NK cells, further indicating that Btk-mediated NK cell activation contributes to TLR3-triggered acute liver injury. Importantly, reduced TLR3-triggered activation of human NK cells is observed in Btk-deficient patients with X-linked agammaglobulinemia (XLA), as evidenced by the reduced IFN-γ, CD69, CD107a expression and cytotoxic activity. These results indicate that Btk is required to activation of NK cells, thus providing insight into the physiological significance of Btk in the regulation of immune cell functions and innate inflammatory response. 083 NEURO-VASCULAR DISRUPTION IN THE CEREBRAL CORTEX DURING EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS AND THE EFFECT OF GLATIRAMER ACETATE R. Aharoni1, R. Eilam2, V. Kalchenko2, Y. Kuznetsov2, M. Sela1, R. Arnon1 1Immunology, The Weizmann Institute of Science, Rehovot, Israel 2Veterinary Resorces, The Weizmann Institute of Science, Rehovot, Israel Blood brain barrier (BBB) dysfunction is a key pathological event in multiple sclerosis (MS) and its model experimental autoimmune encephalomyelitis (EAE). The cerebral cortex is an important target for the disease process, but its pathophysiology is not yet understood. In this study we analyzed the function of the BBB and the communication between the blood vessels, the astrocytes and the nerves in the cortex of MOG-induced EAE inflicted mice. The consequences of treatment by glatiramer acetate (GA, Copaxone) on these processes were also investigated. We developed a system in which permeabilization of sodium fluorescein into the CNS indicated BBB functionality. Extensive sodium fluorescein leakage was observed in EAE-induced mice, but not in na•ve controls. Notably, BBB disruption was evident even before the development of clinical manifestations. In mice treated by GA either at disease induction (prevention) or after disease exacerbation (therapeutic treatment), sodium fluorescein leakage was significantly lower. Immunohistochemical analysis of blood vessels, astrocytes and neurons revealed loss of astrocytes processes towards the blood vessels as well as the neurons in the cortex of EAE-inflicted mice. GA treatment significantly reduced these manifestations resulting in normal appearing neurovascular formation. These findings support the central role of BBB disruption and vascular pathology in the disease, suggesting that neurovascular changes lead to abstracted crosstalk between the blood vessels and the neurons. Additionally, we herein demonstrate a new therapeutic consequence of GA treatment. Specifically, GA treatment prevents the leakage of BBB and reduces the neurovascular damage. 084 MYCOPLASMA SUPERANTIGEN AS A TRIGGER PROMOTES AUTOREACTIVE T CELL RESPONSES AND DISEASE SEVERITY IN AUTOIMMUNE DIABETIC MOUSE H. Mu1 1Internal Medicine, University of Utah School of Medicine, Salt Lake City, USA Background and aims: Infection has been extensively documented as a serious problem causing high mortality and morbidity in diabetic populations. Due to the complex nature of immunological mechanisms, little is known regarding how an infection affects the outcomes of diabetes. Our laboratory has previously established a mouse model of septic shock by infecting mice with the Mycoplasma arthriditis. M. arthritidis produces a potent superantigen (SAg) MAM, which has been shown to be at least partially responsible for the lethal toxicity induced in certain susceptible mouse strains. Methods: In the present study, we have investigated how diabetic immune cells respond to the M. arthritidis and MAM SAg. By using two well-defined animal models of diabetes, STZ and NOD mice. Results: We found that STZ mice infected with M. arthritidis developped severe lethal toxicity and the immune cells predominantly produces pro-inflammatory cytokines TNF?, IL-6, IL-1β, chemokines CCR2/4 and Tbx21, a Th1-related transcription factor; while MAM-activated T cells from NOD mice display a mixture of Th1 and Th17 profiles which are associated with the elevated levels of ICOS molecule and a unique transcription factor LSF (tcfcp2). Since ERK, a signaling protein of MAPK family, is reported to be responsible for the phosphorylation of LSF in T cell activation, thus we propose a novel ERK-LSF-ICOS signaling cascade that is required for the Th1/Th17 development in NOD mice in response to MAM. Conclusions: Our findings suggest that the inhibition of these molecules might be potentially therapeutic in the alleviation of mortality and morbidity in diabetic individuals with severe infections. This work was supported by a grant from the Nora Eccles Treadwell Foundation 085 UP-REGULATION OF MICRORNA-210 INDUCES IMMUNE DYSFUNCTION VIA TARGETING FOXP3 IN CD4+ T CELLS OF PSORIASIS VULGARIS L.T. Wang1, Y.W. Su1, G.P. Liang1, H.Y. Zhai1, P. Zhang1, X.J. Deng1, M. Zhao1, Q.J. Lu1 1Department of Dermatology, Second Xiangya Hospital Central South University, Changsha, China Background and aims Psoriasis vulgaris (PV) has been known as one kind of autoimmune skin diseases. The mechanism of aberrant immunological regulation in PV remains unclear. Here, we investigated the role and mechanism of microRNA-210 (miR-210) in CD4+ T cells of PV. Methods. 18 PV patients and 18 healthy individuals were recruited and CD4+ T cells were isolated by magnetic beads. The transcription levels of miR-210, FOXP3 and cytokines were detected by real-time PCR. Luciferase reporter gene assay was used to identify the target gene of miR-210. CD4+ T cells were transfected by nucleofector device in combination with the human T-cell nucleofector kit. FOXP3 protein level was detected by western blot. The levels of cytokines in the supernatant of cells culture were measured using enzyme-linked immunosorbent assay. Results The expression of miR-210 is up-regulated significantly in the CD4+ T cells of PV patients compared with healthy controls, which are inversely correlated with the mRNA levels of FOXP3. Luciferase reporter gene assay showed that FOXP3 is a target gene of miR-210. Transfection of miR-210 mimic leads to the reduced FOXP3, IL-10 and TGF-β expression and the increased IFN-γ expression in normal CD4+ T cells. In contrast, transfection of miR-210 inhibitor increases FOXP3, IL-10 and TGF-β expression and reduces IFN-γ expression in CD4+ T cells of PV patients. Conclusions Our data demonstrates that up-regulation of miR-210 leads to immune dysfunction by targeting FOXP3 in CD4+ T cells of PV paitents, which suggests the important role of miR-210 in the pathogenesis of PV. 086 INTERLEUKIN 27 INHIBITS ATHEROSCLEROSIS VIA IMMUNOREGULATION OF MACROPHAGES IN MICE H. Yoshida1, T. Hirase2, H. Hara1, K. Node3 1Department of Biomolecular Sciences Division of Immunology, Saga University Faculty of Medicine, Saga, Japan 2Department of Bioscience, National Cardiovascular Center Research Institute, Suita, Japan 3Department of Cardiovascular and Renal Medicine, Saga University Faculty of Medicine, Saga, Japan Chronic inflammation in arterial wall that is driven by immune cells and cytokines plays pivotal roles in the development of atherosclerosis. Interleukin 27 (IL-27) is a member of IL-12 family cytokines that is consisted of IL-27p28 and EBI-3 and has anti-inflammatory properties that regulate T cell polarization and cytokine production. In this study, the role of IL-27/IL-27 receptor signaling in the pathogenesis of atherosclerosis. IL-27-deficient (Ldlr-/-Ebi-3-/-) and IL-27 receptor-deficient (Ldlr-/-WSX-1-/-) Ldlr-/- mice were generated and fed with a high-cholesterol diet (high fat diet; HFD) to induce atherosclerosis. Roles of bone marrow-derived cells in vivo and macrophages in vitro were studied using bone marrow reconstitution by transplantation and cultured peritoneal macrophages, respectively. We demonstrate that mice lacking IL-27 or IL-27 receptor are more susceptible to atherosclerosis compared to wild type due to enhanced accumulation and activation of macrophages in arterial walls (See figure below.) 
The number of circulating pro-inflammatory Ly6Chi monocytes showed no significant difference between wild type mice and mice lacking IL-27 or IL-27 receptor. Administration of IL-27 suppressed the development of atherosclerosis in vivo and macrophage activation in vitro that was indicated by increased uptake of modified low density lipoprotein and augmented production of proinflammatory cytokines. These findings define a novel inhibitory role for IL-27 in atherosclerosis that regulates macrophage activation in mice. 087 B-LYMPHOCYTE SUBSETS IN CHRONIC FATIGUE SYNDROME K. Fuller1, E.W. Brenu1, T.K. Huth1, S.L. Hardcastle1, S. Johnston1, D.R. Staines2, S. Marshall-Gradisnik1 1National Center for Neuroimmunology and Emerging Diseases, Griffith University, Southport, Australia 2Gold Coast Public Health Unit, Queensland Health, Robina, Australia Background and Aims: Chronic Fatigue Syndrome (CFS) is a multi-factorial disease with a significant immune component. To date, limited studies have focused on B-lymphocytes and their implications in CFS. However, it has recently been shown that B-lymphocyte depletion therapy via the administration of Rituximab may be beneficial to some CFS patients, thus suggesting a potential role of B-lymphocytes in CFS. Hence, this study aims to examine the levels of specific B-lymphocyte phenotypes in CFS patients in comparison to non-fatigued control participants. Methods: Heparinised peripheral blood samples were collected from 38 CFS patients fulfilling the 1994 Centers for Disease Control and Prevention criteria for CFS, and 29 non-fatigued control participants. B-lymphocyte phenotypes were characterised using specific antibody combinations and analysed using flow cytometry protocols. Download 1.13 Mb. Share with your friends:
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