Th17 and Th22 cells in psoriatic arthritis and psoriasis

Introduction

The aim of this study was to characterize interleukin 17 (IL-17) and interleukin 22 (IL-22) producing cells in peripheral blood (PB), skin, synovial fluid (SF) and synovial tissue (ST) in patients with psoriasis (Ps) and psoriatic arthritis (PsA).

Methods

Flow cytometry was used to enumerate cells making IL-22 and IL-17, in skin and/or SF and PB from 11 patients with Ps and 12 patients with PsA; skin and PB of 15 healthy controls and SF from rheumatoid arthritis (RA) patients were used as controls. Expression of the interleukin 23 receptor (IL-23R) and chemokine receptors CCR4 and CCR6 was examined. Secretion of IL-17 and IL-22 was measured by ELISA. ST was analysed by immunohistochemical staining of IL-17 and IL-22.

Results

Increased frequencies of IL-17+ and IL-22+ CD4+ T cells were seen in PB of patients with PsA and Ps. IL-17 secretion was significantly elevated in both PsA and Ps, whilst IL-22 secretion was higher in PsA compared to Ps and healthy controls. A higher proportion of the CD4+ cells making IL-17 or IL-22 expressed IL-23R and frequencies of IL-17+, CCR6+ and CCR4+ T cells were elevated in patients with Ps and those with PsA. In patients with PsA, CCR6+ and IL-23R + T cells numbers were elevated in SF compared to PB. Increased frequencies of IL-17+ and IL-22+ CD4+ T cells were demonstrated in Ps skin lesions. In contrast, whilst elevated frequencies of CD4+ IL-17+ cells were seen in PsA SF compared to PB, frequencies of CD4+ IL-22+ T cells were lower. Whereas IL-17 expression was equivalent in PsA, osteoarthritis (OA) and RA ST, IL-22 expression was higher in RA than either OA or PsA ST, in which IL-22 was strikingly absent.

Conclusions

Elevated frequencies of IL-17 and IL-22 producing CD4+ T cells were a feature of both Ps and PsA. However their differing distribution at disease sites, including lower frequencies of IL-22+ CD4+ T cells in SF compared to skin and PB, and lack of IL-22 expression in ST suggests that Th17 and Th22 cells have common, as well as divergent roles in the pathogenesis of Ps and PsA.     

Frequency of Th17 CD20+ cells in the peripheral blood of rheumatoid arthritis patients is higher compared to healthy subjects

Introduction  

Rheumatoid arthritis (RA) is considered a T cell driven autoimmune disease, therefore, the ability of B cell depleting biologics, e.g., anti-CD20 antibodies, to alleviate RA is unclear. This study examined the proportions of IL-17-secreting lymphocytes in the blood of healthy subjects and RA patients and determined if Th17 cells belong to a CD20+ subset of T cells.     

Surgery in the treatment of rheumatoid arthritis and ankylosing spondylitis

The pain, deformities and disabilities resulting from rheumatoid arthritis and ankylosing spondylitis must be treated by a team composed of physician, physical medicine expert, orthopaedic surgeon, and, in certain cases, deep X-ray therapist working simultaneously.The principle of "rest" in order to relieve pain has to be combined with methods designed to preserve and restore function. The multiple joint deformities in these cases may necessitate a long programme of reconstructive or functional treatment, which entails whole-hearted co-operation on the part of the patient in intensive post-operative exercise regime.PROCEDURES ADVOCATED FOR THE UPPER LIMB INCLUDE EXCISION OF THE ACROMION PROCESS TOGETHER WITH THE SUBACROMIAL BURSA TO ALLOW FREE MOVEMENT BETWEEN THE CENTRAL TENDON OF THE DELTOID AND THE TENDINOUS SHOULDER CUFF: arthrodesis of the shoulder in cases where there is more severe joint destruction: in certain cases of elbow-joint arthritis, excision of the radial head and sub-total synovectomy may preserve joint function and avoid or delay the necessity for arthroplasty which can be carried out in two ways: (a) similar to the formal joint excision, or (b) re-shaping the lower end of the humerus and upper end of the ulna lining these surfaces with fascia. The former method is preferable in cases of rheumatoid arthritis. To overcome wrist-joint deformity and restore pronation and supination excision of the lower end of the ulna together with radiocarpal fusion in position for optimum function is advocated. Finger and toe joints may be corrected by resection of the bone ends and capsulectomy.In the lower limbs bilateral involvement of the hip-joint is best treated by vitallium mould arthroplasty which may be carried out in four ways: (1) Routine arthroplasty; (2) Modified Whitman procedure; (3) Modified Colonna operation; and (4) The proximal shaft or intertrochanteric arthroplasty. It is essential in these operations to have knowledge of the operative technique, the use of special hip gouges and reamers, and detailed post-operative supervision.For dorsal kyphosis of the spine, spinal osteotomy at the lumbar level provides excellent correction but is an operation demanding care and skill in its execution.The author's remarks are based on experience gained when working with Dr. M. N. Smith-Petersen at the Massachusetts General Hospital, Boston, U.S.A.     

Effectiveness of adalimumab in treating patients with ankylosing spondylitis associated with enthesitis and peripheral arthritis

Introduction

The purpose of this study was to investigate the effectiveness of adalimumab in enthesitis and peripheral arthritis in patients with ankylosing spondylitis (AS).

Methods

Adults with active AS (Bath ankylosing spondylitis disease activity index [BASDAI] ≥ 4) received adalimumab 40 mg every other week with standard antirheumatic therapies in a 12-week, open-label study. Effectiveness in enthesitis was assessed using the Maastricht ankylosing spondylitis enthesitis score (MASES, 0-13) and by examining the plantar fascia in patients with enthesitis (≥ 1 inflamed enthesis) at baseline; effectiveness in peripheral arthritis was evaluated using tender and swollen joint counts (TJC, 0-46; SJC, 0-44) in patients with peripheral arthritis (≥ 1 swollen joint) at baseline. Overall effectiveness measures included Assessment of SpondyloArthritis International Society 20% response (ASAS20).

Results

Of 1,250 patients enrolled, 686 had enthesitis and 281 had peripheral arthritis. In 667 patients with MASES ≥ 1 at baseline, the median MASES was reduced from 5 at baseline to 1 at week 12. At week 12, inflammation of the plantar fascia ceased in 122 of 173 patients with inflammation at baseline. The median TJC in 281 patients with SJC ≥ 1 at baseline was reduced from 5 at baseline to 1 at week 12; the median SJC improved from 2 to 0. ASAS20 responses were achieved by 70.5% of 457 patients with no enthesitis and no arthritis; 71.0% of 512 patients with only enthesitis; 68.0% of 107 patients with only arthritis; and 66.7% of 174 patients with both.

Conclusions

Treatment with adalimumab improved enthesitis and peripheral arthritis in patients with active AS.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00478660","term_id":"NCT00478660"}}NCT00478660.     

Increased native chemiluminescence in granulocytes isolated from synovial fluid and peripheral blood of patients with rheumatoid arthritis

Polymorphonuclear leukocytes (PMNs) isolated from peripheral blood and synovial fluid of patients with rheumatoid arthritis and from peripheral blood of volunteers were stimulated with 12-Phorbol-13-myristate acetate (PMA). No significant differences in luminol-amplified chemiluminescence were found between different patients and control groups. However, two distinct patterns of native chemiluminescence were observed. Type I showed no, or only a small, increase in native chemiluminescence with integral counts over 30 min less than 3 × 10⁵ cpm, and the majority of samples from volunteers were of this type. Type II was characterized by a burst of native chemiluminescence starting 8 to 15 min after cell stimulation. It was found in most PMN samples from patients with rheumatoid arthritis. Integral counts over 30 min were always higher than 10⁶ cpm and as high as 10⁸ cpm in some cases. A strong inhibition of the Type II native chemiluminescence was caused by desferal, catalase, thiourea, and glutathione. However, the luminol-amplified chemiluminescence remained unchanged or was only slightly decreased under the same experimental conditions. Sodium azide strongly inhibited both kinds of luminscence. Hydroxyl radicals, formed in a Fenton reaction, may be important intermediates in the Type II native chemiluminescence.     

Cyclosporin A inhibits the production of IL-17 by memory Th17 cells from healthy individuals and patients with rheumatoid arthritis

Recent evidence from several studies indicated that IL-17-producing Th17 cells can represent the key effector cells in the induction and development of autoimmune disorders. Cyclosporine A (CsA) is a commonly used immunosuppressant to treat lots of autoimmune diseases including rheumatoid arthritis (RA). Here, we demonstrated that PBMCs and purified CD4⁺ T cells from healthy individuals and patients with RA could be induced to produce large amounts of IL-17 after stimulation with anti-CD3 plus anti-CD28 mAbs. Phenotypic analysis indicated that the majority of IL-17-producing cells were Th17 cells with memory phenotype. The addition of CsA into cell cultures significantly inhibited the IL-17 production by Th17 cells at protein and at mRNA levels. Compared to the PBMCs from normal individuals, PBMCs from the patients with RA produced higher levels of IL-17 that was also significantly inhibited by CsA both at protein and at mRNA levels. The mechanism might be the effect of CsA on the T cells activation because the expression of CD69 and CD25 molecules on T cells was markedly reduced in the presence of CsA. Taken together, these results demonstrated that CsA suppressed the IL-17 production and inhibited the Th17 cells differentiation from both healthy individuals and patients with RA.     

Increased number of Tc17 and correlation with Th17 cells in patients with immune thrombocytopenia

Background

IL-17-secreting CD8+ T cells (Tc17 subset) have recently been defined as a subpopulation of effector T cells implicated in the pathogenesis of autoimmune diseases. The role of Tc17 and correlation with Th17 cells in the pathophysiology of immune thrombocytopenia (ITP) remain unsettled.

Design and Methods

We studied 47 ITP patients (20 newly-diagnosed and 27 with complete response) and 34 healthy controls. IL-17-producing CD3+CD8+ cells (Tc17) and IL-17-producing CD3+CD8− cells (Th17) were evaluated by flow cytometry and expressed as a percentage of the total number of CD3+ cells. Specific anti-platelet glycoprotein (GP) GPIIb/IIIa and/or GPIb/IX autoantibodies were measured by modified monoclonal antibody specific immobilization of platelet antigens. Peripheral blood mononuclear cells of ITP patients were isolated, incubated in the presence of 0, 0.25, 0.5, or 1 µmol/L of dexamethasone for 72 h, and collected to detect Tc17 and Th17 cells by flow cytometric analysis.

Results

IL-17 was expressed on CD3+CD8− and CD3+CD8+ T cells. The percentages of Tc17 and Th17 cells in newly-diagnosed patients were significantly elevated compared to controls, and Tc17 was decreased after clinical treatment. The Th17∶Tc17 ratio was significantly lower in newly-diagnosed patients compared with controls, and was increased in patients who had complete response. There was a significantly positive correlation between Tc17 and Th17 cells in the control group, but not in the ITP patients. A positive correlation existed between Tc17 and the CD8∶CD4 ratio, as well as CD8+ cells in patients with ITP. The frequencies of Tc17 were marginally higher in autoantibody-negative patients than autoantibody-positive patients. Moreover, both Tc17 and Th17 cell percentages decreased as the concentration of dexamethasone in the culture media increased in ITP patients.

Conclusions

Tc17 and the Th17 subset are involved in the immunopathology of ITP. Blocking the abnormally increased number of Tc17 may be a reasonable therapeutic strategy for ITP.     

隐球菌性脑膜炎患者外周血中Th9和Th17细胞水平变化及意义

目的：比较初发隐球菌性脑膜炎患者治疗前后与健康对照人群外周血 CD4＋ T 细胞中 Th9和 Th17细胞的比值,探讨 Th9和 Th17细胞在隐球菌性脑膜炎发病机制中的作用。方法选取初发未经治疗隐球菌性脑膜炎患者及健康对照各12例,抽取隐球菌性脑膜炎患者治疗前和治疗后3周及健康对照的外周血,分离外周血单核细胞,应用流式细胞仪检测技术对3组病例外周血 CD4＋ T 细胞中 Th9和 Th17的比值进行比较。结果与健康对照相比,隐球菌性脑膜炎患者治疗前Th17表达下调,差异有统计学意义;在治疗好转患者中,治疗后 Th17表达显著上调,与治疗前及健康对照相比差异均有统计学意义。Th9在治疗前与健康对照相比无差异,在治疗后隐球菌性脑膜炎患者中表达上调。结论 Th17免疫途径是隐球菌性脑膜炎患者抵御隐球菌感染的重要免疫机制,隐球菌性脑膜炎发病及治疗拮抗可能与 Th17缺乏有关。     

Decreased expression of microRNA‐21 correlates with the imbalance of Th17 and Treg cells in patients with rheumatoid arthritis

The imbalance of Th17/Treg cell populations has been suggested to be involved in the regulation of rheumatoid arthritis (RA) pathogenesis; however, the mechanism behind this phenomenon remains unclear. Recent studies have shown how microRNAs (miRNAs) are important regulators of immune responses and are involved in the development of a variety of inflammatory diseases, including RA. In this study, we demonstrated that the frequencies of CD3⁺CD4⁺IL‐17⁺Th17 cells were significantly higher, and CD4⁺CD25⁺FOXP3⁺ Treg cells significantly lower in peripheral blood mononuclear cells from RA patients. Detection of cytokines from RA patients revealed an elevated panel of pro‐inflammatory cytokines, including IL‐17, IL‐6, IL‐1β, TNF‐α and IL‐22, which carry the inflammatory signature of RA and are crucial in the differentiation and maintenance of pathogenic Th17 cells and dysfunction of Treg cells. However, the level of miR‐21 was significantly lower in RA patients, accompanied by the increase in STAT3 expression and activation, and decrease in STAT5/pSTAT5 protein and Foxp3 mRNA levels. Furthermore, lipopolysaccharide stimulation up‐regulated miR‐21 expression from healthy controls, but down‐regulated miR‐21 expression from RA patients. Therefore, we speculate that miR‐21 may be part of a negative feedback loop in the normal setting. However, miR‐21 levels decrease significantly in RA patients, suggesting that this feedback loop is dysregulated and may contribute to the imbalance of Th17 and Treg cells. MiR‐21 may thus serve as a novel regulator in T‐cell differentiation and homoeostasis, and provides a new therapeutic target for the treatment of RA.     

Enhanced activity of NLRP3 inflammasome in peripheral blood cells of patients with active rheumatoid arthritis

IntroductionInterleukin-1β (IL-1β) is a major inflammatory cytokine, produced predominantly by innate immune cells through NLRP3-inflammasome activation. Both intrinsic and extrinsic danger signals may activate NLRP3. Genetic variations in NLRP3-inflammasome components have been reported to influence rheumatoid arthritis (RA) susceptibility and severity. We sought to assess the activity of NLRP3-inflammasome in patients with active RA compared to healthy individuals.MethodIntracellular protein expression of NLRP3, ASC, pro- and active caspase-1, pro- and active IL-1β was assessed by immunoblotting both at baseline and upon inflammasome activation. NLRP3 function (IL-1β secretion) was assessed upon priming of TLR2 (Pam(3)CysSK(4), TLR3 (poly(I:C)) or TLR4 (LPS) and ATP sequential treatment. We used caspase inhibitors (casp-1, 3/7 and 8) to assess their contribution to IL-1β maturation. All experiments were performed in whole blood cells.ResultsActive RA patients (n = 11) expressed higher basal intracellular levels of NLRP3 (p < 0.008), ASC (p < 0.003), active caspase-1 (p < 0.02) and pro-IL-1β (p < 0.001). Upon priming with TLR4 (LPS) and ATP, RA-derived cell extracts (n = 7) displayed increased expression of NLRP3 (p < 0.01) and active caspase-1 (p < 0.001). Secreted IL-1β in culture supernatants from whole blood cells activated with TLR4 (LPS) or TLR3 agonist (poly(I:C)) plus ATP was higher in RA patients (n = 20) versus controls (n = 18) (p < 0.02 for both). Caspase-1 inhibition significantly reduced IL-1β secretion induced by all stimuli, whereas caspase-8 inhibition affected only TLR4 and TLR3 cell priming.ConclusionPatients with active RA have increased expression of NLRP3 and NLRP3-mediated IL-1β secretion in whole blood cells upon stimulation via TLR3 and TLR4 but not TLR2. In these patients, IL-1β secretion seems to be predominately driven by caspase-1 and caspase-8. Targeting NLRP3 or downstream caspases may be of benefit in suppressing IL-1β production in RA.     

Serum keratan sulphate levels rise in rheumatoid arthritis patients, but fall in ankylosing spondylitis patients compared with normal controls

Serum levels of keratan sulphate (KS) were found to be significantly elevated in patients with destructive and predominantly seronegative rheumatoid arthritis (RA) compared with a control population. Levels in RA did not correlate with clinical or laboratory indices of joint activity or damage. Conversely levels were depressed in ankylosing spondylitis (AS) compared with controls.     

Proteomic analysis of peripheral blood mononuclear cells: selective protein processing observed in patients with rheumatoid arthritis

In a comparative proteome analysis of peripheral blood mononuclear cells (PBMCs), we analyzed 130 two-dimensional gels obtained from 33 healthy control individuals and 32 patients diagnosed with rheumatoid arthritis (RA). We found 16 protein spots that are deregulated in patients with RA and, using peptide mass fingerprinting and Western blot analyses, identified these spots as belonging to 9 distinct proteins. A hierarchical clustering procedure organizes the study subjects into two main clusters based on the expression of these 16 protein spots, one that contains mostly healthy control individuals and the other mostly RA patients. The majority of the proteins differentially expressed in RA patients when compared with healthy controls can be detected as protein fragments in PBMCs obtained from RA patients. This set of deregulated proteins includes several factors that have been shown to be autoantigens in autoimmune diseases.     

Upregulated miR-146a expression in peripheral blood mononuclear cells from rheumatoid arthritis patients

Introduction

MicroRNAs are small noncoding RNA molecules that negatively regulate gene expression via degradation or translational repression of their targeted mRNAs. It is known that aberrant microRNA expression can play important roles in cancer, but the role of microRNAs in autoimmune diseases is only beginning to emerge. In this study, the expression of selected microRNAs is examined in rheumatoid arthritis.

Methods

Total RNA was isolated from peripheral blood mononuclear cells obtained from patients with rheumatoid arthritis, and healthy and disease control individuals, and the expression of miR-146a, miR-155, miR-132, miR-16, and microRNA let-7a was analyzed using quantitative real-time PCR.

Results

Rheumatoid arthritis peripheral blood mononuclear cells exhibited between 1.8-fold and 2.6-fold increases in miR-146a, miR-155, miR-132, and miR-16 expression, whereas let-7a expression was not significantly different compared with healthy control individuals. In addition, two targets of miR-146a, namely tumor necrosis factor receptor-associated factor 6 (TRAF6) and IL-1 receptor-associated kinase 1 (IRAK-1), were similarly expressed between rheumatoid arthritis patients and control individuals, despite increased expression of miR-146a in patients with rheumatoid arthritis. Repression of TRAF6 and/or IRAK-1 in THP-1 cells resulted in up to an 86% reduction in tumor necrosis factor-α production, implicating that normal miR-146a function is critical for the regulation of tumor necrosis factor-α production.

Conclusions

Recent studies have shown that synovial tissue and synovial fibroblasts from patients with rheumatoid arthritis exhibit increased expression of certain microRNAs. Our data thus demonstrate that microRNA expression in rheumatoid arthritis peripheral blood mononuclear cells mimics that of synovial tissue/fibroblasts. The increased microRNA expression in rheumatoid arthritis patients is potentially useful as a marker for disease diagnosis, progression, or treatment efficacy, but this will require confirmation using a large and well defined cohort. Our data also suggest a possible mechanism contributing to rheumatoid arthritis pathogenesis, whereby miR-146a expression is increased but unable to properly function, leading to prolonged tumor necrosis factor-α production in patients with rheumatoid arthritis.     

Decreased expression and activity of G-protein-coupled receptor kinases in peripheral blood mononuclear cells of patients with rheumatoid arthritis.

Beta2-Adrenergic and chemokine receptor antagonists delay the onset and reduce the severity of joint injury in rheumatoid arthritis. beta2-Adrenergic and chemokine receptors belong to the G-protein-coupled receptor family whose responsiveness is turned off by the G-protein-coupled receptor kinase family (GRK-1 to 6). GRKs phosphorylate receptors in an agonist-dependent manner resulting in receptor/G-protein uncoupling via subsequent binding of arrestin proteins. We assessed the activity of GRKs in lymphocytes of rheumatoid arthritis (RA) patients by rhodopsin phosphorylation. We found a significant decrease in GRK activity in RA subjects that is mirrored by a decrease in GRK-2 protein expression. Moreover, GRK-6 protein expression is reduced in RA patients whereas GRK-5 protein levels were unchanged. In search of an underlying mechanism, we demonstrated that proinflammatory cytokines induce a decrease in GRK-2 protein levels in leukocytes from healthy donors. Since proinflammatory cytokines are abundantly expressed in RA, it may provide an explanation for the decrease in GRK-2 expression and activity in patients. No changes in beta2-adrenergic receptor number and Kd were detected. However, RA patients showed a significantly increased cAMP production and inhibition of TNF-alpha production by beta2-adrenergic stimulation, suggesting that reduced GRK activity is associated with increased sensitivity to beta2-adrenergic activation.     

类风湿性关节炎患者外周血单个核细胞 IL-10 基因启动子甲基化状态研究

白细胞介素-10（interleukin-10,IL-10）是在类风湿性关节炎中发挥重要免疫调节作用的细胞因子,其基因失活与已分化的Th1和Th2细胞染色质结构重塑有关。为了探讨IL-10基因启动子甲基化及基因失活在类风湿性关节炎(Rheumatoid Arthritis,RA)发病和进展中的作用,采用逆转录聚合酶链反应(RT-PCR)、酶联免疫吸附实验(enzyme linked immunosorbent assay,ELISA)及甲基化特异性聚合酶链反应(MSP), 分别检测34例类风湿性关节炎患者和30例健康人外周血单个核细胞 IL-10 mRNA、蛋白表达水平及基因启动子甲基化状态。结果显示,病例组IL-10 mRNA及蛋白表达均低于健康对照组,无统计学差异（P>0.05）。病例组甲基化率(85.29%)高于健康对照组(43.33%), 具有统计学差异(x² =12.439,P=0.000)。IL-10基因启动子甲基化状态与其mRNA表达呈显著负相关(r=-0.579, P=0.001), 与所累关节数显著相关,但与血沉（ESR）、C反应蛋白（CRP）、类风湿因子(RF)、年龄均无相关性(P>0.05)。IL-10 mRNA表达与年龄、所累关节数、ESR、CRP及RF均无相关性(P>0.05)。上述结果提示,启动子甲基化可能是IL-10基因失活的重要机制,IL-10基因异常高甲基化状态可能参与了RA的发生发展。     

Th17 cells expressing KIR3DL2+ and responsive to HLA-B27 homodimers are increased in ankylosing spondylitis

CD4 Th cells producing the proinflammatory cytokine IL-17 (Th17) have been implicated in a number of inflammatory arthritides including the spondyloarthritides. Th17 development is promoted by IL-23. Ankylosing spondylitis, the most common spondyloarthritis (SpA), is genetically associated with both HLA-B27 (B27) and IL-23R polymorphisms; however, the link remains unexplained. We have previously shown that B27 can form H chain dimers (termed B27(2)), which, unlike classical HLA-B27, bind the killer-cell Ig-like receptor KIR3DL2. In this article, we show that B27(2)-expressing APCs stimulate the survival, proliferation, and IL-17 production of KIR3DL2(+) CD4 T cells. KIR3DL2(+) CD4 T cells are expanded and enriched for IL-17 production in the blood and synovial fluid of patients with SpA. Despite KIR3DL2(+) cells comprising a mean of just 15% of CD4 T in the peripheral blood of SpA patients, this subset accounted for 70% of the observed increase in Th17 numbers in SpA patients compared with control subjects. TCR-stimulated peripheral blood KIR3DL2(+) CD4 T cell lines from SpA patients secreted 4-fold more IL-17 than KIR3DL2(+) lines from controls or KIR3DL2(-) CD4 T cells. Strikingly, KIR3DL2(+) CD4 T cells account for the majority of peripheral blood CD4 T cell IL-23R expression and produce more IL-17 in the presence of IL-23. Our findings link HLA-B27 with IL-17 production and suggest new therapeutic strategies in ankylosing spondylitis/SpA.