CXCL16 EXPRESSION IN SYSTEMIC LUPUS ERYTHEMATOSUS AND THE INFLUENCING FACTORS ABOUT LUPUS ACTIVITY

Objective: To investigate the relationship between the change of serum CXC chemokine ligand 16(CXCL16)with lupus activity in SLE patients and lupus mice.

Methods: Through clinical observation and animal experiment, 20 SLE patients,20 normal subjects were observed. Height, weight, waist circumference, age, gender, blood glucose, C-peptide , levels of CXCL16 , ceatinine and C-reactive protein(CRP) were obtained.

In Fas-/-lupus mice,CXCL16 -/-C57BL/6J mice?normol C57BL/6J mice (20 in each group), We Compare with the control group and model group, observate them 4 week, collecte and determine including serum CXCL16, levels of antibodies to double-strand DNA, oxidized phospholipids, C-reactive protein, as well as IL-6 and TNF-α. To analyze the clinical datas, studentÕs unpaired t-test was used for the comparison between 2 groups. One-way ANOVA assay and multiple stepwise regression were used for multiple groups.

Results: Serum level of CXCL16 is significantly increased in SLE patients compared with the healthy control subjects (P<0.05). Serum levels of CXCL16 of Fas-/-lupus mice are higher than the control group(P<0.05). Serum levels of CXCL16 of model group are higher than the control group(P<0.05). Multiple stepwise regression analysis indicated that levels of CXCL16 are positive correlation with lupus activity index such as CRP and antibodies to ds-DNA.

Conclusions: Both of clinical observation and animal experiment indicate Serum CXCL16 levels are positive associated with activity levels of SLE. But how to elucidate the pathophysiologcial mechanism of CXCL16 in SLE requires further research.

Systemic Lupus Erythematosus is a chronic autoimmune disease with complex etiology. It is characterized by the presence of anti-nuclear autoantibodies (ANA). These form self-reactive immune complexes which deposit in tissues where they initiate inflammation and the destruction of organs, such as the kidney. Recent data from multiple murine models has demonstrated that Toll-like receptor (TLR)-7, is involved in both the breakdown in central tolerance and the progression to severe disease. Previous work from multiple groups has demonstrated that the Yaa murine lupus susceptibility locus drives the development of glomerulonephritis (GN) via an increase in expression of TLR7.

Vitamin D is a steroid hormone that exhibits a number of regulatory effects on growth, proliferation, apoptosis and functions of various cell types of the human immune system on top of its well-known actions on calcium homeostasis and bone metabolism. The immunomodulatory actions of vitamin D on the immune system are highly relevant to the pathophysiology of systemic lupus erythematosus (SLE). Low vitamin D state is common in patients with SLE because of a number of actors that include avoidance of sunshine, sun protection, renal dysfunction and the use of medications such as glucocorticoids, anti-convulsants. anti-malarials and the calcineurin inhibitors, which either alter the metabolism of vitamin D or down-regulate the functions of the vitamin D receptors. Hypovitaminosis D in SLE correlates with disease activity scores, and is associated with low bone mineral density, fatigue and increased cardiovascular risk. This talk updates the recent evidence on the interplay between vitamin D status and SLE regarding disease onset, activity state and disease-related complications. Recommendations for vitamin D supplementation in SLE patients will also be discussed.

Dysregulated activation of autoreactive B cells is critical for the development of autoimmune diseases. Autoantibodies produced by such B cells can induce a variety of autoimmune diseases, including HashimotoÕs thyroiditis, systemic lupus erythematosus (SLE) and others. The physiological mechanisms responsible for the maintenance of B cell tolerance by T cells with regulatory activity remain to be investigated. Recently, we identified early growth response gene 2 (Egr2)-controlled CD4⁺CD25^-Foxp3^- regulatory T cells (Treg) that express lymphocyte activation gene 3 (LAG3). LAG3⁺ Treg effectively suppress autoantibody production and autoimmunity. Adoptive transfer of LAG3⁺ Treg, but not CD4⁺CD25⁺ Foxp3+ Treg, significantly suppressed the progression of nephritis and autoantibody production in MRL/lpr lupus-prone mice. LAG3⁺ Treg strongly suppressed antibody production and the development of germinal center B cells in a programmed death 1 (PD-1)- and Egr2-dependent manner. Interestingly, Fas/FasL co-stimulation was required for the full-expression of Egr2 in LAG3⁺ Treg. These findings indicate an unexpected role of Fas/FasL interaction in the pathogeneses of autoimmune diseases. From these results, we believe that a novel regulatory T cell subset (LAG3⁺ Treg) is one of essential elements in the regulation of the immune responses involved in the maintenance of self-tolerance.

Liwei Lu, Department of Pathology, The University of Hong Kong, Hong Kong

TNF cytokines play a crucial role in immune regulation by modulating lymphocyte proliferation and apoptosis. Our previous studies have shown that B cell activating factor (BAFF), a member of TNF family cytokines produced by myeloid lineage cells such as macrophages and dendritic cells, was critically involved in promoting B cell survival and autoantibody production in mice with collagen-induced arthritis (CIA). Recently, we identified a novel function of BAFF in the induction of IL-10-producing regulatory B cells. BAFF-induced IL-10-producing B cells showed a distinct CD1d^hiCD5⁺ phenotype mainly derived from marginal-zone B cells, which possessed a potent function in inhibiting T cell activation and cytokine production. BAFF was found to activate the transcription factor AP-1 for binding to IL-10 promoter. In collagen-immunized mice, adoptive transfer of BAFF-induced IL-10-producing B cells markedly reduced the disease severity and joint damage of autoimmune arthritis via suppression of Th17 cell response. Taken together, our findings have provided further insight in understanding the roles of BAFF and regulatory B cells in autoimmune pathogenesis, which may facilitate the development of new therapeutic strategies for targeting autoimmune diseases.

Tumour Necrosis Factor (TNF) is a key mediator of inflammatory diseases such as rheumatoid arthritis and plays a central role in dendritic cell (DC) biology. Our aim was to dissect the individual contributions of the two TNF receptors (TNFR1 and TNFR2) in regulating the survival and function of human inflammatory monocyte-derived (moDC) and steady-state myeloid DC. A DNA binding assay was used to demonstrate that in moDC TNFR1, but not TNFR2, activates the classical p65 NFκB pathway whereas both TNFR1 and TNFR2 activate the alternative p100/p52 NFκB pathway, highlighting differences in signalling downstream of the receptors. In moDC, TNFR1-selective, but not TNFR2-selective stimulation resulted in increased expression of DC maturation markers CD83 and CD86, and enhanced T cell stimulatory capacity. Furthermore, moDC survival was prolonged by selective stimulation of either TNFR1 or TNFR2 as shown by reduced intracellular levels of active caspase-3, indicating that innate signals can promote DC survival in the absence of DC maturation. Accordingly, the p65 NFκB pathway was involved in the pro-survival effect of TNFR1 whereas the Bcl-2/Bcl-xL pathway was essential to survival mediated by both TNFR. In contrast, in myeloid DC, maturation was mainly mediated through TNFR1, whereas TNFR2 was superior in protecting DC from cell death. An antagonistic TNFR1-specific antibody was used to confirm that cell death protection via TNFR2 was independent of TNFR1-mediated signalling. Understanding the immunoregulatory properties of signalling through these two TNF receptors is important for the design of more targeted anti-TNF therapy.

Natural killer (NK) cells are multifunctional lymphocytes with divergent phenotypic properties in different organs. Recent studies has uncovered that liver NK cells possess memory-like properties in contact hypersensitivity (CHS) models, which challenges the traditional definitions of innate and adptive immunity. However, the phenotype and origin of these ÒmemoryÓ NK cells cannot be distinguished from other NK cell subpopulations. Here, we defined the transcriptional, phenotypic and functional features of liver NK cell subsets and their roles in mediating CHS. Liver NK cells can be divided into two distinct subsets, CD49a⁺DX5^- and CD49a^-DX5⁺. Substantial transcriptional and phenotypic differences existed between liver CD49a⁺DX5^- NK cells and other NK cell subsets; the former possessed memory potential and conferred hapten-specific CHS responses upon hapten challenge. Importantly, CD49a⁺DX5^- NK cells were liver-resident, present in the liver sinusoid blood but not the afferent and efferent blood of the liver, whereas CD49a^-DX5⁺ NK cells are highly migratory and incapable of mediating CHS responses. Moreover, CD49a⁺DX5^- subset appeared to originate from hepatic hematopoietic progenitor/stem cells but not from the bone marrow, and maintained their phenotypes at steady state. Our findings of liver-resident NK cells shed new light on understanding how NK cells contribute to skin-contact hepersensitivity.

The vasculitides are a set of related disorders characterized by blood vessel inflammation leading to tissue or organ injury. Specific vasculitic disorders are defined depending on the vessels affected (size, type, location). Many vasculitides are caused by immune complexes and the activation of the complement system can be involved in the pathogenesis. In particular, complement involvement was demonstrated in small vessel vasculitides, such as ANCA-associated vasculitides, cryoglobulinemia, Henoch-Sch*nlein purpura and IgA nephropathy. Evidences of complement activation come from the detection of complement components in affected tissues and from the observation of reduced plasma levels of complement intact proteins. However, the consumption of complement factors is mainly evident in urticarial vasculitis (UV), a relatively rare clinico-pathologic entity characterized by recurrent episodes of urticaria. In contrast to other forms of urticaria, the typical wheals of UV persist for more than 24 hours and hyperpigmentation residues after resolution. Among subjects affected, hypocomplementemic patients show more severe multi-organ involvement than normo-complementemic patients. Anti-C1q autoantibodies have been described in these patients. Circulating antigen-antibody complexes deposit in vessel and cause complement activation through the classical pathway. Main systemic manifestations include constitutional symptoms (fever, fatigue), arthralgia and arthritis, pleuritis and obstructive lung disease, glomerulonephritis and interstitial nephritis, pericarditis and cardiac valve disease, neuropathy, gastrointestinal symptoms (abdominal pain, nausea, vomiting, diarrhea) and ocular disorders (episcleritis, uveitis, conjunctivitis). Skin biopsy is mandatory for diagnosis and shows histopathologic features of leukocytoclastic vasculitis. Treatment is oriented by disease severity and include antihistamines, steroids, dapsone, hydroxychloroquine and immunosuppressant drugs.

Autoimmune hepatitis (AIH) is a chronic inflammation of the liver caused by an abnormal autoimmune reaction against hepatocytes. The pathogenesis of AIH involves a loss of tolerance to hepatic self antigens in a susceptible host. The diagnosis is based on histological abnormalities, and characteristic clinical and biochemical findings, which include abnormal levels of serum globulins, and the presence of one or more characteristic autoantibodies such as ANA, SMA, and anti-SLA. Interface hepatitis with abundant plasma cells in the infiltrate is characteristic histological features of AIH. The simplified criteria have high sensitivity and specificity for diagnosis of AIH. The revised original criteria have the complementary role to avoid the false negative diagnosis in atypical AIH patients. Immunosuppressive treatment can attenuate hepatic inflammation, reverse fibrosis, and eventually improve the patient's prognosis and life quality. Corticosteroids, either alone or in combination with azathioprine, are the standard treatment of choice for AIH. Most patients with AIH show a good response to immunosuppressive treatment, although patients with late-stage or severe disease are less likely to achieve remission.The simultaneous or successive coexistence of AIH with primary biliary cirrhosis (PBC) in the same patient has been described as PBC-AIH overlap syndrome. we found that there is a group of PBC-AIH patients who undergo a complete response to corticosteroids and that this variant may be identified using the Paris criteria, especially when lowering the IgG threshold to 1.3 xULN, in conjunction with the simplified IAIHG scoring system.

Lupus nephritis (LN) occurs in 40-75% of systemic lupus erythmatosus (SLE) patients. Various autoantibodies have been identified in SLE. Anti-ribosomal P protein (anti-P) antibodies are one of the marker antibodies in SLE. Association of these antibodies with psychiatric or neurological symptoms has been reported. However their involvement in LN is not very well known. We analyzed the presence of anti-P in LN patients.

It is a descriptive and cross sectional study in which sera of 133 ANA positive SLE patients were investigated for the presence of anti-P antibodies (from January 2010 to September 2012). Other autoantibodies tested were anti-dsDNA, anti-Sm, anti-SS-A, anti-SS-B, anti-histones, anti-RNP and anti-PCNA antibodies. Clinical features and histopathological classification of LN were also analyzed in these patients.

Data was analyzed using SPSS software version.12

Results

Anti-P antibodies were found in 15(11.5%) patients along with one or more other autoantibodies. Isolated anti-P antibodiesÕ positivity was not found. Nine (60%) patients were positive for anti-dsDNA and eight (53%) for anti-SS-A antibodies. Anti-P was mostly associated with class IV and class V lupus nephritis. The most common symptom was pedal edema followed by arthralgia and oral ulcers. Three patients had history of seizures. Psychosis was not found in any of these patients.

Presence of anti-P antibodies may add to the severity in SLE patients with LN. Therefore careful monitoring of these patients may help in better patient management. We did not find an association of anti-P antibodies with psychiatric involvement in our LN patients.

Among all approaches proposed for MS therapy, an approach that neutralizes only the pathogenic T cells reacting against myelin, while leaving the innocent immune cells intact, is the ultimate goal in the immune-specific therapy for MS. However, the multiplicity of primary target antigens, along side the dynamic nature of autoimmunity in MS, whereby the specificity of anti-myelin pathogenic autoreactivities may shift or expand in the same patient with disease progression, impose major difficulties in devising immune-specific therapy to MS. Among possible reasons for therapeutic failure of clinical trial with MBP-APL, is the complexity of anti-myelin autoreactivities. To overcome this multiplicity and complexity of pathogenic autoreactivities in MS, we have put forward the concept of concomitant multi-antigen targeting as, a conceivably more effective approach to immunotherapy of MS.

We investigated such Òmulti-epitope-targetingÓ approach in EAE associated with multiple pathogenic autoimmunities (ÒcomplexÓ EAE) in (C57Bl/6JxSJL/J)F1 mice via a multi-APL agent designated mMEP-APL, a protein product of a synthetic gene designed to encode only characterized APLs of the defined encephalitogenic epitopes of five myelin encephalitogenic proteins (MBP, MOG, PLP, MOBP and OSP).

mMEP-APL was found highly effective in suppressing and treatment of classical and ÒcomplexÓ EAE. We show that systemic administration of mMEP-APL was associated with downregulation of Th1/Th17 cytokine secretion and upregulation of TGF-b secretion together with induction of regulatory CD4⁺FoxP3⁺ cells.

Our data emphasize that a ÔÔmulti-epitope-targetingÕÕ strategy is required for effective immune-specific therapy of organ-specific autoimmune diseases associated with complex and dynamic pathogenic autoimmunity, such as MS.
152 ESTABLISHMENT OF EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS IN CYNOMOLGUS MONKEYS, A MODEL FOR MULTIPLE SCLEROSIS
J. Ni¹, J. Liu¹, J. Deng¹, G. Cui¹, L. Le¹, S. Wang¹, H. Zhang<sup>1


1</sup>Pharmacology, Prisys Biotechnologies, Shanghai, China

he experimental autoimmune encephalomyelitis (EAE) model in rodents is widely used, however, the immune system of rodents is quite different from humans, plus the genetic homogeneity and microbiological cleanness of the inbred strains make the model system significantly apart from human multiple sclerosis (MS). To more closely model MS, we established the EAE model in Cynomolgus monkeys. Brain white matter, spinal cord and MOG peptides were tested for induction of EAE in monkeys. The typical disease related neurological symptoms, such as anorexia, weight loss, ataxia, neck spasticity, and blindness, were observed two weeks after the immunization. A set of disease scoring criteria was adopted to semi-quantitatively measure the disease severity. MRI examinations discovered the multiple hyper-intense areas in the brain of the EAE monkeys at disease active stages, similar to those found in MS patients. The findings of gross autopsy indicated multiple hemorrhagic lesions in the white matter of the affected monkey brains. Both disease incidence and severity of the EAE were antigen-dose dependent. Interestingly, the disease progression induced by brain white matter and spinal cord presented mixed types of the EAE episodes, while the typical relapsing-remitting disease, representing the major type of MS in patients, was induced by MOG peptide.