抗RA33抗体、RF、CRP、ANA与ENA联合检测在类风湿性关节炎患者诊断、治疗及预后中的作用

目的：探讨血清抗RA33抗体、RF、CRP联合检测在类风湿性关节炎患者诊断、治疗及预后中的作用。方法：采用ELISA法对35例类风湿性关节炎患者和30例健康对照者进行抗RA33抗体的检测,间接免疫荧光法检测ANA、免疫印迹法检测ENA,免疫比浊法进行类风湿因子（RF）及CRP的检测。结果：类风湿性关节炎患者组RF水平为：[（104．51±153．88）KIU／L]与健康对照组[（10．89±2．78）KIU／L]比较,差异非常显著（p〈0．01）;类风湿性关节炎组CRP[（11．60±23．24）mg／L]与健康对照组[（2．57±2．18）mg／L]比较,差异显著（p〈0．05）;类风湿性关节炎组抗RA33抗体水平：[（17．81±35．11）U／mL]与健康对照组[（8．10±8．40）U／mL]比较,差异非常显著（p〈0．01）。RF、抗RA33抗体、CRP及ANA诊断类风湿性关节炎的灵敏度分别为：80．00％,34．29％,42．86％,62．86％;RF、抗RA33抗体、CRP及ANA诊断类风湿性关节炎的特异性分别为：93.33％,93.33％,90．00％,96．67％;ENA的检出率均较低。结论：类风湿性关节炎患者抗RA33抗体检出的灵敏度低,但特异性强。对类风湿性关节炎患者进行抗RA33抗体、ANA、RF、CRP、ENA联合检测,对于疾病的进展、病因分析、指导治疗和改善预后均具有重要意义。     

Can anti-cyclic citrullinated peptide antibody-negative RA be subdivided into clinical subphenotypes?

Introduction  

Studies investigating genetic risk factors for susceptibility to rheumatoid arthritis (RA) studied anti-citrullinated peptide antibody (CCP)-positive RA more frequently than anti-CCP-negative RA. One of the reasons for this is the perception that anti-CCP-negative RA may include patients that fulfilled criteria for RA but belong to a wide range of diagnoses. We aimed to evaluate the validity of this notion and explored whether clinical subphenotypes can be discerned within anti-CCP-negative RA.     

A monoclonal anti-idiotype specific for human polyclonal IgM rheumatoid factor.

One of the hallmarks of rheumatoid arthritis (RA) is the production of high titers of rheumatoid factor (RF) antibody directed against the Fc portion of IgG. Anti-Id that recognize the majority of monoclonal RF from patients with B cell dyscrasias are reactive with only 1 to 2% of these polyclonal RF from RA patients. We describe a new monoclonal anti-Id, 4C9, that recognizes a L chain determinant on polyclonal IgM RF from patients with RA but does not recognize a panel of monoclonal RF from patients with B cell malignancies. 4C9 reactivity is found in the serum of 34/43 RF-positive RA patients and in 12/12 RF-positive synovial fluids, but in only 1/14 RF-negative sera from RA patients and 1/22 sera containing monoclonal IgM RF. 4C9 reactivity is highly enriched in purified IgM RF from nine RA patients and represents a variable percentage of total IgM RF up to a maximum of 23%. Furthermore, 4C9 reactivity is enriched in the synovial fluid of three of five RA patients compared with serum, suggesting that 4C9-reactive IgM RF are synthesized within the joint. IgG RF from RA synovial fluids are not 4C9 reactive, indicating either that different genes are used to encode IgM and IgG RF in RA patients, or that IgG RF have somatically mutated away from idiotypic reactivity.     

Safety of rituximab in the treatment of B cell malignancies: implications for rheumatoid arthritis

The chimeric anti-CD20 monoclonal antibody rituximab has been used extensively in the treatment of B cell malignancies, and more recently it has emerged as a potential treatment for rheumatoid arthritis (RA), via selective B lymphocyte depletion. Experience in oncology shows that rituximab is well tolerated in a variety of settings, with mild-to-moderate infusion related reactions following the first infusion being the most common adverse event. Current data suggest that the safety profile of rituximab in patients with RA is similar to that in oncology, but that the adverse events are less frequent and less severe in patients with RA.     

Association study of single nucleotide polymorphisms in pre-miRNA and rheumatoid arthritis in a Han Chinese population

The aim of this study was to perform an association study between two single nucleotide polymorphisms (SNPs) rs2910164 G>C and rs3746444 T>C in pre-miRNA (hsa-mir-146a and hsa-mir-499) and rheumatoid arthritis (RA) in the Han Chinese population. 208 Han Chinese patients with RA and 240 healthy controls were recruited in this study. The SNPs was genotyped by polymerase chain reaction-restriction fragment length polymorphism. Anti-cyclic citrullinated peptide (anti-CCP) antibody was measured by enzyme linked immunosorbent assay and rheumatoid factor (RF) was measured by rate nephelometry. The genotype frequencies between cases and controls were compared by χ(2) analysis. No significant association between the SNPs (rs2910164 and rs3746444) and RA was observed (P = 0.631 and 0.775, respectively), and the SNPs did not show any association with the RF-positive (P = 0.631 and 0.775, respectively). However, there was a significant difference on the level of anti-CCP antibody between different genotypes in rs3746444 (P = 0.007). The heterozygote CT had significantly higher level of anti-CCP antibody compared with homozygote CC and TT (P = 0.054 and 0.003, respectively). We first investigated the association between the SNPs (rs2910164 G>C and rs3746444 T>C) in the pre-miRNA (hsa-mir-146a and hsa-mir-499) and RA in a Han Chinese population. We did not find a significant association between the SNPs and the susceptibility to RA, while the SNP rs3746444 may affect anti-CCP antibody production.     

抗环瓜氨酸肽抗体联合类风湿因子在检测类风湿关节炎中的临床价值

目的：探讨抗环瓜氨酸肽抗体（抗CCP抗体）以及类风湿因子（RV）检测对类风湿关节炎（RA）诊断的意义。方法：采用酶联免疫吸附试验（ELISA）检测355份人血清的抗CCP抗体,同时采用使用贝克曼库尔特Image800双光镜免疫浊度分析仪定量监测类风湿因子（RF）,其中包括门诊及住院RA患者135例,非RA组170例,正常对照组来自本院的健康体检人员50例。结果：抗-CCP检测在RA组与非RA组（和正常对照组）之间的检测结果有统计学差异（P〈0．05）。RF检测在RA组与非RA组（和正常对照组）之间的检测结果有统计学差异（P〈0．05）。在135例RA病人中,抗CCP抗体的阻性率为70．4％,在非RA病人中的阳性率为3．5％,抗CCP抗体对RA的敏感性和特异性分别为70．4％、96．5％。RF的阳性率为63．7％,在非RA病人中的阳性率为14．1％,RF对RA的敏感性和特异性分别为63．7％、81．1％。联合应用抗CCP抗体与RF进行诊断,串联时敏感性为59．3％,特异性为97．1％。并联时敏感性为74．8％,特异性为85.3％。结论：抗CCP抗体和RF对RA具有较好的敏感性和很高的特异性,二者联合检测可提高对RA早期诊断的准确性。     

Denosumab

Denosumab is an anti-receptor activator of nuclear factor (NF)-kappaB (RANK) ligand human monoclonal antibody studied as a treatment for postmenopausal osteoporosis (PMO) and bone destruction due to rheumatoid arthritis (RA) or metastatic cancers. As of February 2009, the candidate was undergoing US Food and Drug Administration review, and might be approved by October 2009. Late phase clinical trials demonstrated that denosumab possesses a similar safety profile to bisphosphonates and that it can be either equally or more effective than bisphosphonates at preventing bone loss due to PMO, RA or cancer treatment and metastases.Key Words: monoclonal antibody, RANKL, bone loss, osteoporosis, breast cancer, rheumatoid arthritis     

A possible role for TSLP in inflammatory arthritis

Thymic stromal lymphopoietin (TSLP) is an IL-7-like cytokine that triggers dendritic cell-mediated Th2-type inflammatory responses and is considered as a master switch for allergic inflammation. In this study, we found increased levels of TSLP and, also TNF-alpha as previously reported, in synovial fluid specimens derived from patients with rheumatoid arthritis (RA) when compared with those from patients with osteoarthritis (OA). In addition, TNF-alpha up-regulated TSLP expression in RA- and OA-derived synovial fibroblasts, which was inhibited by IFN-gamma. Furthermore, anti-TSLP neutralizing antibody ameliorated a TNF-alpha-dependent experimental arthritis induced by anti-type II collagen antibody in mice. Collectively, these results suggest that TSLP, as a downstream molecule of TNF-alpha, may be involved in the pathophysiology of inflammatory arthritis. TSLP might thus play a role not only in allergic diseases but also in inflammatory arthritis such as RA.     

Mannan Binding Lectin (MBL) genotypes coding for high MBL serum levels are associated with rheumatoid factor negative rheumatoid arthritis in never smokers

Introduction  

Previous studies have provided inconsistent results on whether variants in the MBL2 gene, coding for the complement-activating mannan-binding lectin (MBL) protein, associate with rheumatoid arthritis (RA). We re-evaluated this in context of the main environmental and genetic risk factors (smoking, HLA-DRB1 'shared epitope' (SE), PTPN22*620W), which predispose to rheumatoid factor (RF) and/or anti-citrullinated-protein antibody (ACPA)-positive RA.     

Targeting nanomedicines in the treatment of rheumatoid arthritis: focus on certolizumab pegol

Anti-TNFalpha therapy has revolutionized the treatment of rheumatoid arthritis (RA) and other inflammatory diseases. These drugs are powerful and expensive. A new anti-TNFalpha agent, a nanomolecule comprising a humanized Fab' antibody fragment against TNFalpha with a polyethylene glycol tail, is shortly to complete phase III trials in RA. In this review we will discuss the construct of this new molecule, data from trials so far, and its potential place in the market place.     

Type II collagen antibody response is enriched in the synovial fluid of rheumatoid joints and directed to the same major epitopes as in collagen induced arthritis in primates and mice

Introduction

Antibodies towards type II collagen (CII) are detected in patients with rheumatoid arthritis (RA) and in non-human primates and rodents with collagen induced arthritis (CIA). We have previously shown that antibodies specific for several CII-epitopes are pathogenic using monoclonal antibodies from arthritic mice, although the role of different anti-CII epitopes has not been investigated in detail in other species. We therefore performed an inter-species comparative study of the autoantibody response to CII in patients with RA versus monkeys and mice with CIA.

Methods

Analysis of the full epitope repertoire along the disease course of CIA was performed using a library of CII triple-helical peptides. The antibody responses to the major CII epitopes were analyzed in sera and synovial fluid from RA patients, and in sera from rhesus monkeys (Macaca mulatta), common marmosets (Callithrix jacchus) and mice.

Results

Many CII epitopes including the major C1, U1, and J1 were associated with established CIA and arginine residues played an important role in the anti-CII antibody interactions. The major epitopes were also recognized in RA patients, both in sera and even more pronounced in synovial fluid: 77% of the patients had antibodies to the U1 epitope. The anti-CII immune response was not restricted to the anti-citrulline protein antibodies (ACPA) positive RA group.

Conclusion

CII conformational dependent antibody responses are common in RA and are likely to originate from rheumatoid joints but did not show a correlation with ACPA response. Importantly, the fine specificity of the anti-CII response is similar with CIA in monkeys and rodents where the recognized epitopes are conserved and have a major pathogenic role. Thus, anti-CII antibodies may both contribute to, as well as be the consequence of, local joint inflammation.     

Characterization of a human synovial cell antigen: VCAM-1 and inflammatory arthritis

The contribution of synovial cells to the pathogenesis of rheumatoid arthritis (RA) is only partly understood. Monoclonal antibody (mAb) 1D5 is one of very few mAb ever raised against RA synovial cells in order to study the biology of these cells. Studies on the expression pattern and structural features of the 1D5 Ag suggest that 1D5 recognizes human vascular cell adhesion molecule-1 (VCAM-1), which is an intercellular adhesion molecule. Vascular cell adhesion molecule-1 may be involved in a number of crucial intercellular interactions in RA.     

Identification of citrullinated alpha-enolase as a candidate autoantigen in rheumatoid arthritis

Antibodies against citrullinated proteins are highly specific for rheumatoid arthritis (RA), but little is understood about their citrullinated target antigens. We have detected a candidate citrullinated protein by immunoblotting lysates of monocytic and granulocytic HL-60 cells treated with peptidylarginine deiminase. In an initial screen of serum samples from four patients with RA and one control, a protein of molecular mass 47 kDa from monocytic HL-60s reacted with sera from the patients, but not with the serum from the control. Only the citrullinated form of the protein was recognised. The antigen was identified by tandem mass spectrometry as alpha-enolase, and the positions of nine citrulline residues in the sequence were determined. Serum samples from 52 patients with RA and 40 healthy controls were tested for presence of antibodies against citrullinated and non-citrullinated alpha-enolase by immunoblotting of the purified antigens. Twenty-four sera from patients with RA (46%) reacted with citrullinated alpha-enolase, of which seven (13%) also recognised the non-citrullinated protein. Six samples from the controls (15%) reacted with both forms. Alpha-enolase was detected in the RA joint, where it co-localised with citrullinated proteins. The presence of antibody together with expression of antigen within the joint implicates citrullinated alpha-enolase as a candidate autoantigen that could drive the chronic inflammatory response in RA.     

Interleukin-1 production by mononuclear cells from rheumatoid synovial effusions

The polypeptide interleukin-1 (IL-1) is a cytokine that may mediate inflammation and connective tissue damage in rheumatoid arthritis (RA). We examined cytokine production by normal blood and by rheumatoid synovial mononuclear cells with sensitive (picomolar) assays. The assays were immunolabeling and immunoblotting with rabbit anti-IL-1 beta sera, and proliferation of the murine D10 cell line to IL-1. Little or no cytokine was detected in rheumatoid joint fluid or in exudate mononuclear cells from patients with acute rheumatoid flares. The mononuclear cells could be induced to make IL-1 upon stimulation with lipopolysaccharide (LPS). The responsive cells were monocytes, since all could be double-labeled with anti-IL-1 and the monocyte-specific CD14 antibody. More than 80% of the synovial fluid monocytes made IL-1 beta after 24 hr in 2 ng/ml LPS. Other agents failed to induce IL-1 from enriched populations of monocytes including interferon gamma (IFN-gamma), poly (I/C), phorbol myristate acetate (PMA), concanavalin A (Con A), phytohemagglutinin (PHA), and anti-CD3 antibodies. Relatively high levels of dendritic cells (DC) were present in RA effusions, but these did not produce IL-1 in response to any of the above stimuli. Blood dendritic cells also did not make IL-1, whereas blood monocytes responded comparably to synovial exudate cells. The data indicate that rheumatoid exudate monocytes make very little IL-1 during acute flares of arthritis and that this cytokine is primarily a macrophage rather than a dendritic cell product.     

High synovial expression of the inhibitory FcgammaRIIb in rheumatoid arthritis

Activating Fc gamma receptors (FcgammaRs) have been identified as having important roles in the inflammatory joint reaction in rheumatoid arthritis (RA) and murine models of arthritis. However, the role of the inhibitory FcgammaRIIb in the regulation of the synovial inflammation in RA is less known. Here we have investigated synovial tissue from RA patients using a novel monoclonal antibody (GB3) specific for the FcgammaRIIb isoform. FcgammaRIIb was abundantly expressed in synovia of RA patients, in sharp contrast to the absence or weak staining of FcgammaRIIb in synovial biopsies from healthy volunteers. In addition, the expression of FcgammaRI, FcgammaRII and FcgammaRIII was analyzed in synovia obtained from early and late stages of RA. Compared with healthy synovia, which expressed FcgammaRII, FcgammaRIII but not FcgammaRI, all activating FcgammaRs were expressed and significantly up-regulated in RA, regardless of disease duration. Macrophages were one of the major cell types in the RA synovium expressing FcgammaRIIb and the activating FcgammaRs. Anti-inflammatory treatment with glucocorticoids reduced FcgammaR expression in arthritic joints, particularly that of FcgammaRI. This study demonstrates for the first time that RA patients do not fail to up-regulate FcgammaRIIb upon synovial inflammation, but suggests that the balance between expression of the inhibitory FcgammaRIIb and activating FcgammaRs may be in favour of the latter throughout the disease course. Anti-inflammatory drugs that target activating FcgammaRs may represent valuable therapeutics in this disease.     

Adalimumab clinical efficacy is associated with rheumatoid factor and anti-cyclic citrullinated peptide antibody titer reduction: a one-year prospective study

Studies on autoantibody production in patients treated with tumor necrosis factor-alpha (TNF-alpha) inhibitors reported contradictory results. We investigated in a prospective study the efficacy of a treatment with human monoclonal anti-TNF-alpha antibody (adalimumab) in patients with rheumatoid arthritis (RA) and we evaluated the relationship between treatment efficacy and the incidence and titers of disease-associated and non-organ-specific autoantibodies. Fifty-seven patients with RA not responsive to methotrexate and treated with adalimumab were enrolled. Antinuclear, anti-double-stranded(ds)DNA, anti-extractable nuclear antigens, anti-cardiolipin (aCL), anti-beta2 glycoprotein I (anti-beta2GPI) autoantibodies, rheumatoid factor (RF) and anti-cyclic citrullinated peptide (anti-CCP) autoantibodies were investigated at baseline and after 6 and 12 months of follow-up. Comparable parameters were evaluated in a further 55 patients treated with methotrexate only. Treatment with adalimumab induced a significant decrease in RF and anti-CCP serum levels, and the decrease in antibody titers correlated with the clinical response to the therapy. A significant induction of antinuclear autoantibodies (ANA) and IgG/IgM anti-dsDNA autoantibodies were also found in 28% and 14.6% patients, respectively, whereas aCL and anti-beta2GPI autoantibodies were not detected in significant quantities. No association between ANA, anti-dsDNA, aCL and anti-beta2GPI autoantibodies and clinical manifestations was found. Clinical efficacy of adalimumab is associated with the decrease in RF and anti-CCP serum levels that was detected after 24 weeks and remained stable until the 48th week of treatment. Antinuclear and anti-dsDNA autoantibodies, but not anti-phospholipid autoantibodies, can be induced by adalimumab but to a lower extent than in studies with other anti-TNF blocking agents.     

B-cell depletion for rheumatic diseases: where are we?

Immune system dysfunction is common to rheumatic disorders, with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) being classic examples. Altered development and function of B cells may play a prominent role. B-cell abnormalities also occur in other rheumatic diseases, eg, Sjogren's syndrome, Behcet's disease, antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis, and dermatomyositis. Hence, B-cell depletion has been investigated as a therapeutic option. Clinical trials in RA and SLE have shown that rituximab, an anti-CD20 monoclonal antibody, can profoundly reduce disease activity and is generally well tolerated. Reports of rituximab treatment for ANCA-associated vasculitis and dermatomyositis are also promising. These encouraging results validate the strategy of B-cell depletion in various rheumatic diseases. B-cell depletion with rituximab is under study in larger clinical trials for the purposes of regulatory approval to define more closely its place in RA and SLE treatment paradigms, and smaller clinical trials are ongoing or planned in associated inflammatory diseases.     

一种基于量子点检测抗CCP抗体的免疫荧光层析法

抗环瓜氨酸多肽 (cyclic citrullinated peptide, CCP) 抗体是类风湿关节炎 (rheumatoid arthritis, RA) 早期诊断的重要生物标志物. 为了实现对RA的早期诊断,本研究建立了一种基于CdTe量子点标记技术检测抗CCP抗体的免疫荧光层析法. 将CCP多肽与小牛血清白蛋白 (bovine serum albumin, BSA) 连接,再将CCP-BSA和 羊抗鼠IgG分别在硝酸纤维素膜 (nitrocellulose membrane, NC膜) 上划线,作为检测线 (test line, T线) 和质控线 (control line, C线). 制备量子点并在量子点上标记鼠抗人IgG,喷在玻璃纤维上并烘干,最后组装大卡、切割并封装制成检测试纸条. 应用该试纸条检测了RA患者及健康人血清临床样本200份,以酶联免疫吸附 测定法 (enzyme-linked immunosorbent assay, ELISA) 为对照,计算免疫荧光层析法的检测灵敏度和特异性. 结果显示,建立的量子点免疫荧光层析试纸条检测抗 CCP抗体的灵敏度为97.5 %,特异性为95.8%. 该方法操作简单、快速,可实现床旁检测 (point-of-care testing, POCT),能应用于RA的早期诊断.     

Interleukin-6 Receptor Blockade Selectively Reduces IL-21 Production by CD4 T Cells and IgG4 Autoantibodies in Rheumatoid Arthritis

Interleukin-6 (IL-6) levels are known to be increased in patients with rheumatoid arthritis (RA). Tocilizumab, a monoclonal antibody to the IL-6 receptor (IL-6R), reduces disease activity in RA, although its mechanisms of action remain unclear. Since IL-6 regulates cytokine production by CD4 T cells during activation, we investigated whether treatment with tocilizumab altered the phenotype and cytokine production by CD4 T cells in patients with rheumatoid arthritis. We show here that tocilizumab treatment does not change the production of cytokines by naïve CD4 T cells. However, tocilizumab treatment causes a selective decrease of IL-21 production by memory/activated CD4 T cells. Since IL-21 is known to promote plasma cell differentiation, we examined the effect of tocilizumab on the production of autoantibodies. We show that there is a decrease in the levels of IgG4 anti-CCP antibodies, but there is no effect on IgG1 anti-CCP antibodies. In addition, we show that IL-21 is a powerful inducer of IgG4 production by B cells. Thus, IL-6 contributes to the presence of IgG4-specific anti-CCP autoantibodies in RA patients, likely through its effect on IL-21 production by CD4 T cells, and IL-6R blockade down-regulates this pathway.