Lymphocyte-activation gene 3+ (LAG3+) forkhead box protein 3− (FOXP3−) regulatory T cells induced by B cells alleviates joint inflammation in collagen-induced arthritis

Rheumatoid arthritis (RA) is an autoimmune disease in which dysregulated immune cells primarily target synovial joints. Despite recent advances in the treatment of RA, including the introduction of biologic therapies and employment of combination disease-modifying antirheumatic drug strategies, remission rates remain suboptimal. Previous studies have demonstrated that the adoptive transfer of induced regulatory T cells (iTregs) was effective in treating a murine model of collagen-induced arthritis (CIA). The objective of this study was to develop optimal potential iTreg-based therapy for CIA by adoptively transferring LAG3⁺ Treg-of-B cells. B-cell-induced Treg-of-B cells expressed LAG3 but not Foxp3 (designated LAG3⁺ Treg-of-B), and secreted IL-4, IL-10, and TGF-β. Furthermore, LAG3⁺ Treg-of-B cells suppressed the proliferation of CD4⁺CD25⁻ responder T cells through both LAG3 and IL-10 production. In the murine CIA model, adoptive transfer of LAG3⁺ Treg-of-B cells alleviated the joint severity as well as local and systemic inflammation. Treatment with LAG3⁺ Treg-of-B cells also promoted IL-10 production in lymphocytes isolated from the spleen and draining lymph nodes. Moreover, mice receiving LAG3⁺ Treg-of-B cell treatment showed significantly less pronounced osteolysis in the hind footpads, which correlated with the downregulation of tartrate-resistant acid phosphatase expression. In conclusion, we identified a novel subset of Tregs for CIA treatment. This insight may facilitate exploring novel regulatory T-cell-based therapies for human autoimmune diseases.     

Pathologic finding of increased expression of interleukin-17 in the synovial tissue of rheumatoid arthritis patients

Rheumatoid arthritis (RA) is a common autoimmune disease of chronic systemic inflammatory disorder that will affect multiple tissues and organs such as skin, heart or lungs; but it principally attacks the joints, producing a nonsuppurative inflammatory and proliferative synovitis that often progresses to major damaging of articular cartilage and joint ankylosis. Although the definite etiology is still unknown, recent studies suggest that T-helper cells (Th17) may play a pivotal role in the pathogenesis of RA. And interleukin-17 (IL-17), which is a cytokine of Th17 cells, may be a key factor in the occurrence of RA. The binding of IL-17 to specific receptor results in the expression of fibroblasts, endothelial and epithelial cells and also synthesis of several major factors such as tumor necrosis factor alpha (TNF-α), IL-1β that result in the structural damage of RA joints. Though some previous studies have shown that IL-17 exists in the synovium of RA, few has definite proof quantitatively by pathology about its existence in synovial membrane. This study comprised of 30 RA patients and 10 healthy control, pathologic study of the synovial membrane showed increased expression of IL-17 in the synovial tissue of RA patients, the intensity is compatible with clinical severity of disease as validated by DAS28 score and disease duration. Northern blot study also confirmed the increased expression of IL-17 in the synovial tissues. This study sheds further light that IL-17 may be a key factor in the pathogenesis of RA and a determinant of disease severity.     

Tolicizumab研究进展

类风湿关节炎（RA）、幼年特发性全身性关节炎（sJIA）、Castleman病等多种炎性疾病发生、进展过程中均有IL-6的过量表达,且发挥关键作用。Tolicizumab是第一个人源性的IL-6R抗体,通过特异性识别结合IL-6R而阻断IL-6生物学活性,发挥抑制上述疾病炎症反应的作用。大量研究结果证实。Tocilizumab不仅能明显缓解RA的临床症状,而且能防止关节继续被破坏。此外,Tocilizumab在治疗全身性和关节型sJIA、Castleman病等疾病也获得了巨大的成功。     

Nanomedicines for chronic non-infectious arthritis: the clinician's perspective

Rheumatoid arthritis (RA) and osteoarthritis (OA) are prevalent chronic health conditions. However, despite recent advances in medical therapeutics, their treatment still represents an unmet medical need because of safety and efficacy concerns with currently prescribed drugs. Accordingly, there is an urgent need to develop and test new drugs for RA and OA that selectively target inflamed joints thereby mitigating damage to healthy tissues. Conceivably, biocompatible, biodegradable, disease-modifying antirheumatic nanomedicines (DMARNs) could represent a promising therapeutic approach for RA and OA. To this end, the unique physicochemical properties of drug-loaded nanocarriers coupled with pathophysiological characteristics of inflamed joints amplify bioavailability and bioactivity of DMARNs and promote their selective targeting to inflamed joints. This, in turn, minimizes the amount of drug required to control articular inflammation and circumvents collateral damage to healthy tissues. Thus, nanomedicine could provide selective control both in space and time of the inflammatory process in affected joints. However, bringing safe and efficacious DMARNs for RA and OA to the marketplace is challenging because regulatory agencies have no official definition of nanotechnology, and rules and definitions for nanomedicines are still being developed. Although existing toxicology tests may be adequate for most DMARNs, as new toxicity risks and adverse health effects derived from novel nanomaterials with intended use in humans are identified, additional toxicology tests would be required. Hence, we propose that detailed pre-clinical in vivo safety assessment of promising DMARNs leads for RA and OA, including risks to the general population, must be conducted before clinical trials begin.     

A cytokine-centric view of the pathogenesis and treatment of autoimmune arthritis

Cytokines are immune mediators that play an important role in the pathogenesis of rheumatoid arthritis (RA), an autoimmune disease that targets the synovial joints. The cytokine environment in the peripheral lymphoid tissues and the target organ (the joint) has a strong influence on the outcome of the initial events that trigger autoimmune inflammation. In susceptible individuals, these events drive inflammation and tissue damage in the joints. However, in resistant individuals, the inflammatory events are controlled effectively with minimal or no overt signs of arthritis. Animal models of human RA have permitted comprehensive investigations into the role of cytokines in the initiation, progression, and recovery phases of autoimmune arthritis. The discovery of interleukin-17 (IL-17) and its association with inflammation and autoimmune pathology has reshaped our viewpoint regarding the pathogenesis of arthritis, which previously was based on a simplistic T helper 1 (Th1)-Th2 paradigm. This review discusses the role of the newer cytokines, particularly those associated with the IL-17/IL-23 axis in arthritis. Also presented herein is the emerging information on IL-32, IL-33, and IL-35. Ongoing studies examining the role of the newer cytokines in the disease process would improve understanding of RA as well as the development of novel cytokine inhibitors that might be more efficacious than the currently available options.     

Therapeutic potential of IL-15 in rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic, destructive inflammatory autoimmune disease. Cytokine-mediated immunity has been found to play an important role in the pathogenesis of autoimmune diseases including RA. Recently, much attention has been paid on the role of IL-15, which is a member of the 4 α-helix bundle cytokine family. IL-15 was detected in serum and synovial fluid from RA patients and arthritis mice models. Moreover, administration of IL-15 leads to the development of severe inflammatory arthritis, suggesting that IL-15 may be therapeutically relevant in RA. Therefore, targeting IL-15 may be significantly important and valuable. In this article, we discuss the biological features and effects of IL-15 and summarize recent advances on the pathological roles of IL-15 in RA and treatment for RA.     

Joint cytokine quantification in two rodent arthritis models: kinetics of expression, correlation of mRNA and protein levels and response to prednisolone treatment

Biomarker quantification in disease tissues from animal models of rheumatoid arthritis (RA) can help to provide insights into the mechanisms of action of novel therapeutic agents. In this study we validated the kinetics of IL-1beta, TNF-alpha and IL-6 mRNA and protein expression levels in joints from DBA/1OlaHsd murine collagen-induced arthritis (CIA) and Lewis rat Streptococcal cell wall (SCW)-induced arthritis by real-time polymerase chain reaction (PCR) TaqMan and Enzyme-linked immunosorbent assay (ELISA). Prednisolone was used as a reference to investigate any correlation between clinical response and cytokine levels at selected time-points. To our knowledge this is the first report showing a close pattern of expression between mRNA and protein for IL-1beta and IL-6, but not for TNF-alpha, in these two models of RA. The kinetics of expression for these biomarkers suggested that the optimal sampling time-points to study the effect of compounds on both inflammation and cytokine levels were day 4 postonset in CIA and day 3 after i.v challenge in SCW-induced arthritis. Prednisolone reduced joint swelling through a mechanism associated with a reduction in IL-1beta and IL-6 protein and mRNA expression levels. At the investigated time points, protein levels for TNF-alpha in arthritic joints were lower than the lower limit of detection of the ELISA, whereas mRNA levels for this cytokine were reliably detected. These observations suggest that RT-PCR TaqMan is a sensitive technique that can be successfully applied to the quantification of mRNA levels in rodent joints from experimental arthritis models providing insights into mechanisms of action of novel anti-inflammatory drugs.     

Complex pattern of Th1 and Th2 activation with a preferential increase of autoreactive Th1 cells in BALB/c mice with proteoglycan (aggrecan)-induced arthritis

The central role of CD4+ T cells and the balance between T helper (Th) subpopulations in the pathogenesis of autoimmune diseases have been extensively studied. Proteoglycan (aggrecan)-induced arthritis (PGIA) is a murine model for rheumatoid arthritis (RA), which is characterized by a Th1 dominance at the onset of the disease. In addition to CD4+ T cells, antigen-presenting B cells and autoantibodies seem to play an important role in the development and regulation of PGIA. To identify proteoglycan-specific CD4+ T cell subsets and Th1- and Th2-supported antibody isotypes during the progression of PGIA, spleen cells of proteoglycan-immunized BALB/c mice were harvested at different times of immunization, and at different stages of the disease, and their cytokine production and antigen-specific antibody isotype profiles were determined by enzyme-linked immunospot (ELISPOT) assays. Both Th1 and Th2 cytokine-producing cells, with the predominance of IL-4/IL-5-secreting cells, were detected during the prearthritic stage, and a shift toward a Th1 dominance was observed at the time of onset of arthritis. Tissue homogenates of acutely inflamed joints contained significantly higher levels of interferon-gamma than IL-4. The prearthritic period and both the acute and chronic phases of joint inflammation were characterized by IgG1 dominance in the sera and this correlated with the number of IgG1-secreting B cells in the spleen. However, the ratio of autoreactive IgG1/IgG2a-secreting cells decreased in arthritic animals. These results indicate the activation and possible regulatory roles of both Th1 and Th2 subsets in the autoimmune process, with the necessity of a relative increase of autoreactive Th1 cells for the induction of joint inflammation.     

Arthritis and pneumonitis produced by the same T cell clones from mice with spontaneous autoimmune arthritis

SKG mice, a newly established model of rheumatoid arthritis (RA), spontaneously develop autoimmune arthritis accompanying extra-articular manifestations, such as interstitial pneumonitis. To examine possible roles of T cells for mediating this systemic autoimmunity, we generated T cell clones from arthritic joints of SKG mice. Two distinct CD8(+) clones were established and both showed in vitro autoreactivity by killing syngeneic synovial cells and a variety of MHC-matched cell lines. Transfer of each clone to histocompatible athymic nude mice elicited joint swelling and histologically evident synovitis accompanying the destruction of adjacent cartilage and bone. Notably, the transfer also produced diffuse severe interstitial pneumonitis. Clone-specific TCR gene messages in the inflamed joints and lungs of the recipients gradually diminished, becoming hardly detectable in 6-11 months; yet, arthritis and pneumonitis continued to progress. Thus, the same CD8(+) T cell clones from arthritic lesions of SKG mice can elicit both synovitis and pneumonitis, which chronically progress and apparently become less T cell dependent in a later phase. The results provide clues to our understanding of how self-reactive T cells cause both articular and extra-articular lesions in RA as a systemic autoimmune disease.     

Inflammatory diseases of the bones and joints

The inflammatory diseases of the bones and joints encompass infections and the consequences of immunologically mediated local and systemic disease. Infections involve bones (osteomyelitis) and joints (septic arthritis) separately as well as together and result in necrosis with inflammatory features determined by the duration of the infection. In many cases, the infecting organism, whether bacterial, fungal or mycobacterial, is present within the infected site, but occasionally is no longer identifiable locally despite the persistence of infection-related phenomena. Granulomatous infections in bones and joints require distinction from Sarcoidosis. The diagnosis of the immunologically mediated inflammatory diseases, such as RA, depends as much on the clinical features as on the histologic ones, with a few findings that might point to one or the other in ambiguous cases. Any discussion of inflammatory arthropathies should at least mention Osteoarthritis, if for no other reason than to compare it with the traditionally regarded inflammatory diseases. However, there has been increasing interest on the potential role that synovial inflammation may play in the pathogenesis of this vary common arthritis. Ultimately, the diagnosis of the inflammatory diseases of the bones and joints requires the synthesis of information from many sources: clinical, serological, microbiological, radiographic, and pathological.     

Functions of interleukin‐34 and its emerging association with rheumatoid arthritis

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic, synovial inflammation affecting multiple joints, finally leading to extra‐articular lesions for which limited effective treatment options are currently available. Interleukin‐34 (IL‐34), recently discovered as the second colony‐stimulating factor‐1 receptor (CSF‐1R) ligand, is a newly discovered cytokine. Accumulating evidence has disclosed crucial roles of IL‐34 in the proliferation and differentiation of mononuclear phagocyte lineage cells, osteoclastogenesis and inflammation. Recently, IL‐34 was detected at high levels in patients with active RA and in experimental models of inflammatory arthritis. Blockade of functional IL‐34 with a specific monoclonal antibody can reduce the severity of inflammatory arthritis, suggesting that targeting IL‐34 or its receptors may constitute a novel therapeutic strategy for autoimmune diseases such as RA. Here, we have comprehensively discussed the structure and biological functions of IL‐34, and reviewed recent advances in our understanding of the emerging role of IL‐34 in the development of RA as well as its potential utility as a therapeutic target.     

Alleviation of rheumatoid arthritis by cell-transducible methotrexate upon transcutaneous delivery

Rheumatoid arthritis (RA) is a systemic autoimmune disease that is initiated and maintained by various inflammatory/immune cells and their cytokines, leading to cartilage degradation and bone erosion. Despite its potent therapeutic efficacy on RA, the oral administration of methotrexate (MTX) provokes serious adverse systemic complications, thus necessitating the local application of MTX. Here, we show that transcutaneous MTX (TC-MTX) can efficiently penetrate joint skin ex vivo and in vivo, and that TC-MTX can significantly improve the various inflammatory symptoms associated with RA. Further, TC-MTX preserved the joint-structures in mice with collagen-induced arthritis (CIA), which was also confirmed by three-dimensional micro-computed tomography scan. TC-MTX markedly decreased the secretion of inflammatory cytokines both in the serum and in inflamed joints of CIA mice. Further, its therapeutic potential is comparable to that of etanercept, a biological agent that block tumor necrosis factor (TNF)-α. Importantly, the systemic cytotoxicity of TC-MTX was not detected. Thus, TC-MTX can be a new therapeutic modality for RA patients without systemic complications.     

Functional niche of inflamed synovium for Th17-cell expansion and activation in rheumatoid arthritis: Implication to clinical therapeutics

Th17 cells selectively produce the signature cytokines such as IL-17, IL-21 and IL-22, and play a critical role for the chronic inflammatory response and subsequent tissue damage in synovial joints of patients with rheumatoid arthritis (RA). The preliminary clinical study indicates that IL-17 neutralizing therapy can ameliorate inflammatory cascades within peripheral synovial joints in the major population of patients with active RA. Multiple cellular and molecular modulations for the Th17-cell-polarized responses could exist, however, in the inflamed synovium, possibly resulting in a functional niche for the generation and activation of pathogenic Th17 cells. This might establish a vicious cycle culminating in the striking marginal erosions of cartilage and bone in the RA joints, and at least partially abrogate the potential therapeutic benefits related to IL-17 antagonizing or Th17-cell depleting therapy. This article is aimed to discuss the cellular and molecular pathways critically involved in the expansion and activation of pathogenic Th17 cells in RA synovium, with emphasis on the potential therapeutic implications for targeting these pathways to the present and future RA clinics.     

Lymph node biopsy analysis reveals an altered immunoregulatory balance already during the at‐risk phase of autoantibody positive rheumatoid arthritis

The balance between proinflammatory and regulatory CD4⁺ T cells is tightly controlled in lymphoid organs. In autoimmune diseases this balance is altered in the periphery and target tissue of patients. However, not much is known about the balance initiated in lymphoid organs during the development of disease. Since systemic autoimmunity is present years before the clinical manifestations of rheumatoid arthritis (RA), it is possible to study the immunoregulatory balance during the earliest (preclinical) phases of disease. Here, we report for the first time the frequency and phenotype of proinflammatory and regulatory CD4⁺ T cells in lymph node biopsies obtained from autoantibody positive individuals at risk for developing RA, patients with established disease and healthy controls. The frequency of proinflammatory LN Th1 cells was increased in RA patients compared with HCs, while the frequency of regulatory T cells was lower in LN biopsies of RA‐risk individuals. Upon in vitro stimulation LN CD4⁺ T cells produced lower levels of proinflammatory cytokines, IFN‐γ and IL‐17A, in both RA‐risk individuals and early RA patients. This study shows that already during the earliest phases of systemic autoimmunity the immunoregulatory balance between proinflammatory and regulatory CD4⁺ T cells is altered in LN tissue.     

Novel multimeric IL-1 receptor antagonist for the treatment of rheumatoid arthritis

Protein therapeutics targeting inflammatory mediators have shown great promise for the treatment of autoimmunities such as rheumatoid arthritis (RA). However, a significant challenge in this area has been their low in vivo stability and consequently their severely compromised therapeutic efficacy. One such therapeutic molecule IL-1 receptor antagonist (IL-1ra), used in the treatment of rheumatoid arthritis, has displayed only modest efficacy in human clinical trials owing to its short biological half-life. Herein, we report a novel approach to conglomerate individual protein entities into a drug depot by incorporation of an amyloidogenic motif Lys-Phe-Phe-Glu (KFFE) thereby dramatically improving their systemic persistence and in turn their therapeutic efficacy in a mice model of autoimmune arthritis.     

Immunoglobulin heavy chain variable region gene utilization by B cell hybridomas derived from rheumatoid synovial tissue.

Rheumatoid arthritis (RA) is a chronic inflammatory disease that primarily affects synovial joints. Activated B lymphocytes and plasma cells are present in the synovial tissue and are thought to contribute to the immunopathology of the rheumatoid joint. To investigate rheumatoid synovial B lymphocytes, we have generated B cell hybridomas from synovial tissue of an RA patient. Here we describe the immunoglobulin VH gene repertoire of eight IgM- and 10 IgG-secreting synovial-derived hybridomas. The VH4 gene family is highly represented (38.5%) in this panel of hybridomas compared with the frequency of VH4 gene expression in circulating B lymphocytes reported previously (19-22%) and with the VH4 gene frequency we observed in a panel of hybridomas derived in the same manner from the spleen and tonsil of normal individuals (19%). The increased frequency of VH4 gene expression was not due to the expansion of a single B cell clone in vivo as none of these hybridomas was clonally related. Two synovial-derived hybridomas secreted autoantibodies; one (VH3+) secreted an IgM-rheumatoid factor (RF) and the other (VH4+) secreted IgM with polyreactive binding to cytoskeletal proteins and cardiolipin. The antibodies secreted by the remaining synovial-derived hybridomas were not reactive with the autoantigens tested. The VH gene usage in a proportion (5/17) of synovial-derived hybridomas that expressed CD5 antigen provided preliminary evidence that CD5+ B cells in RA synovium have a similar increase of VH4 gene expression reported for CD5+ B cells from normal individuals and patients with chronic lymphocytic leukaemia.     

The genetic pathogenesis,diagnosis and therapeutic insight of rheumatoid arthritis

Rheumatoid arthritis (RA) is a systemic autoimmune disease that causes chronic inflammation of the joints. RA is a heterogeneous disorder caused by an abnormal autoimmune response triggered by the complex interactions of genetic and environmental factors that contribute to RA etiology. However, its underlying pathogenic mechanisms are yet to be fully understood. In this review, I provide an overview of the pathogenesis, diagnosis and therapeutic insight in the clinical management of RA in light of the recent updates to classification criteria and recent discoveries of genetic loci associated with susceptibility for RA.     

类风湿关节炎患者血清和关节液细胞因子水平的变化及临床意义

目的探讨幼年类风湿关节炎(JRA)及类风湿关节炎(RA)患者IL-6、IL-8、sIL-2R和TNF-α等细胞因子(CK)水平的变化,及其与风湿活动的传统指标血沉(ESR)和C-反应蛋白(CRP)的相关性。方法采用夹心ELISA法,对30例JRA和34例RA患者的血清中,4例JRA、7例RA、6例骨性关节炎(OA)和9例半月板损伤(MT)患者的关节液中IL-6、IL-8、sIL-2R和TNF-α的水平进行检测。结果①30例JRA、34例RA患者血清IL-6和sIL-2R的水平与对照组相差非常显著(P〈0.01);30例JRA患者血清IL-8水平与对照组比较相差显著(P〈0.05)。②JRA全身型、少关节型患者血清IL-8、sIL-2R的水平和JRA多关节型患者血清IL-6的水平与对照组相差非常显著(P〈0.01)。③4例JRA及7例RA患者关节液sIL-2R的水平和RA患者关节液的IL-6水平与对照组相差显著(P〈0.05)。④JRA患者血清IL-6和sIL-2R的水平与ESR和CRP的变化呈明显的相关关系(r值分别为0.532和0.621)。结论①IL-6、sIL-2R的水平与JRA、RA病的活动性有关,是类风湿活动性的主要指标。②sIL-2R不仅参与JRA和RA的全身病理损伤,而且是引起关节局部损伤的主要CK,IL-6也参与JRA关节局部的病理损伤,在RA关节局部损伤似乎更为重要。③IL-8主要参与JRA的全身病理损伤,对关节局部病理损伤似乎并不重要。     

Antigen-specific T cells transduced with interleukin-10 ameliorate experimentally induced arthritis without impairing the systemic immune response to the antigen]

For the treatment of rheumatoid arthritis, efficient drug delivery methods to the inflamed joints need to be developed. Since T cells expressing an appropriate autoantigen-specific receptor can migrate to inflamed lesions, it has been reasoned that they can be employed to deliver therapeutic agents. In order to examine the ability and efficiency of such T cells as a vehicle, we employed an experimentally induced model of arthritis. Splenic T cells from DO 11.10 T cell receptor transgenic mice specific for OVA were transduced with murine IL-10. Adoptive transfer of the IL-10-transduced DO 11.10 splenocytes ameliorated OVA-induced arthritis, in spite of the presence of around 95% non-transduced cells. Using GFP as a marker for selection, the number of transferred cells needed to ameliorate the disease was able to be reduced to 10(4). Preferential accumulation of the transferred T cells was observed in the inflamed joint, and the improvement in the disease was not accompanied by impairment of the systemic immune response to the antigen, suggesting that the transferred T cells exert their antiinflammatory task locally, mainly in the joints where the antigen exists. In addition, IL-10-transduced DO 11.10 T cells ameriolated mBSA-induced arthritis when the arthritic joint was co-injected with OVA in addition to mBSA. These results suggest that T cells specific for a joint specific antigen would be useful as a therapeutic vehicle in rheumatoid arthritis for which the arthritic autoantigen is still unknown.