Quantitative histological analysis of antigen-induced arthritis in the rabbit.

Antigen-induced arthritis in the rabbit (AIAR) provides the closest experimental equivalent to human rheumatoid arthritis in terms of infiltration of synovial tissue by lymphoid cells. A method is described for quantitative histological analysis of AIAR. Measurements of total cell numbers, lymphocyte and polymorphonuclear leucocyte infiltration, and thickness of infiltrated synovium were obtained for ranges of antigen dosage and duration of arthritis. The method has been devised as part of a system for the analysis of joint swelling, synovial fluid biochemistry and cytology, cartilage proteoglycan chemistry and synovial histology on the same specimen.     

INFLAMMATORY ACTIVITIES OF SYNOVIAL FLUIDS FROM RHEUMATOID- AND OSTEO-ARTHRITIS TO GUINEA PIG SKIN

For initial study to examine the chemical pathogenesis of rheumatoid and non-rheumatoid arthritis, the nature and inflammatory activities of 12 synovial fluids from rheumatoid arthritis and 10 from osteoarthritis were studied. Both rheumatoid and non-rheumatoid synovial fluids were found to have high protein concentration, acid and neutral protease activities. When injected 0.1 ml intradermally to normal guinea pig skin, both synovial fluids induced a strong vascular permeability at 0 minute, mild one at 60 minutes, and mild inflammation which comprised histologically moderate leukocyte infiltration around the venules, swelling of venular endothelial cells, edema, and degeneration of collagen fibrils in the deep dermis of the guinea pig skin 4 hours following the injection. However, these parameters of inflammation induced by synovial fluids from both arthritis were of same degree in intensity. The identification of chemical mediators of inflammation in the synovial fluids of both arthritis requires further study.     

Complement C4-derived monocyte-directed chemotaxis-inhibitory factor. A molecular mechanism to cause polymorphonuclear leukocyte-predominant infiltration in rheumatoid arthritis synovial cavities

To reveal the mechanism of the lesser infiltration of monocytes in synovial cavities with rheumatoid arthritis despite the presence of chronic inflammation, the synovial fluid from 15 rheumatoid arthritis patients was analyzed with respect to leukocyte chemotaxis. The synovial fluid possessed strong chemotactic activity to polymorphonuclear leukocytes but rather suppressed one to monocytes. The synovial fluid contained two different inhibitory activities in monocyte chemotaxis. One, which also suppressed polymorphonuclear leukocyte chemotaxis, was identified as alpha 1 protease inhibitor. The other, with molecular weight of 8 kd, possessed the specificity to monocytes and shared the antigenicity with complement C4 but not with C3 or C5. A similar inhibitor was generated in normal human plasma when the classical pathway of the complement system was initiated with aggregated human IgG, while it was not when alternative pathway was initiated with zymosan. The small size factor in the synovial fluid, apparently derived from C4, seemed to be a cyto-directed factor that might block an early part of signal transduction system of monocytes in the chemotaxis. After removal of the small-size inhibitor, the synovial fluid exhibited chemotactic ability to monocytes. Therefore the apparent C4-derived factor might play a key role in the polymorphonuclear leukocyte-predominant infiltration in the synovial fluid of rheumatoid arthritis.     

Antibodies to the mycobacterial 65-kd heat-shock protein are reactive with synovial tissue of adjuvant arthritic rats and patients with rheumatoid arthritis and osteoarthritis.

The expression of 65-kd mycobacterial heat-shock protein (HSP)-related antigens in synovial membrane from rats and humans with arthritis was studied using three monoclonal antibodies and one polyclonal antiserum directed to antigens of mycobacteria. The antibodies labeled synovial tissue sections from both adjuvant arthritis (AA) rats and from patients with either rheumatoid arthritis (RA) or osteoarthritis (OA); especially the synovial lining cells appeared to be positive. The cytoplasmic staining patterns in rats and humans were essentially the same and were not related to the extent of inflammation ie, the size of lymphoid infiltration. In control tissues no cytoplasmic staining was observed. The results suggest a role for a 65-kd HSP or a cross-reactive molecule in the immunopathologic process of arthritic disease.     

Presence of HLA-DR antigen on synovial type A and B cells: an immunoelectron microscopic study in rheumatoid arthritis, osteoarthritis and normal traumatic joints

The HLA-DR antigen was investigated in synovial epithelia of rheumatoid arthritis, osteoarthritis and normal traumatic joints, using monoclonal anti-human HLA-DR antibody and horseradish peroxidase-conjugated antibody. The HLA-DR staining was observed in an electron microscope. HLA-DR antigen was observed to be present on the surface of both macrophage-like (type A) and fibroblast-like (type B) cells in synovial epithelia in all rheumatoid arthritis, osteoarthritis and normal joints. Since type A and B cells are ultrastructurally considered to be of synovial origin, the findings suggest that the expression of HLA-DR antigen on the surface is one of the common attributes of type A and B cells in synovial epithelia, even before cellular infiltration of chronic inflammation in rheumatoid arthritis. These cells may play an important role in the initiation of rheumatoid inflammation, since HLA-DR antigen is considered equivalent to murine Ia antigen.     

Morphological changes in the joint tissues and synovial fluid in rheumatoid arthritis and arthrosis deformans]

A simultaneous morphological investigation of bioptic material of the synovial sheath, articular cartilage, and synovial fluid from patients with various forms of rheumatoid arthritis (100 patients with a typical articular form, 10 with benign evolution, and 20 with lesions of visceral organs) was carried out. It was noted that in the usual articular form of rheumatoid arthritis the most common morphological component in the synovial sheath was lympho-plasmo-cellular infiltration. In the "benign" form of rheumatoid arthritis immunomorphological shifts were manifested but slightly, in the articular-visceral form in the immunocompetent cells there were observed karyopycnosis and plasmorrhexis. Cells and the main matter of the cartilage apparently underwent an enzymatic lysis, the intensity of which correlated with the degree of phagocytosis of the synovial fluid. In the deforming osteo-arthrosis (150 observations) in the synovial sheath there were usually noted drastic sclerosis and atrophy of organ-specific structures, impairmement of the production of the synovial fluid, and dystrophic falling into fibers of the articular cartilage with intensive proliferation of the cartilage cells. It is probable that distructiion of the cartilage in arthrosis depends upon an impairment of the function of the synovial sheath to produce synovial fluid.     

Tyrosine Phosphorylated Proteins in Synovial Cells of Rheumatoid Arthritis

Two of the key events in the pathogenesis of rheumatoid arthritis are the synovial cell proliferation and lymphocyte infiltration into the synovium. The resulting synovitis is longlasting and leads to destructive arthritis, which is a hallmark of rheumatoid arthritis. Accumulating evidence suggests that one of the key biochemical events in the altered cell function of RA is phosphorylation of the tyrosine residues of proteins.

In this paper we review the cellular components participating in the chronic inflammation of RA joints. We present the results of analyzing tyrosine phosphorylated proteins of synovial cells from RA patients and discuss a possible pathogenic role of non-receptor tyrosine kinase in RA.     

Tyrosine phosphorylated proteins in synovial cells of rheumatoid arthritis

Two of the key events in the pathogenesis of rheumatoid arthritis are the synovial cell proliferation and lymphocyte infiltration into the synovium. The resulting synovitis is longlasting and leads to destructive arthritis, which is a hallmark of rheumatoid arthritis. Accumulating evidence suggests that one of the key biochemical events in the altered cell function of RA is phosphorylation of the tyrosine residues of proteins. In this paper we review the cellular components participating in the chronic inflammation of RA joints. We present the results of analyzing tyrosine phosphorylated proteins of synovial cells from RA patients and discuss a possible pathogenic role of non-receptor tyrosine kinase in RA.     

Differential expression and response to anti-TNFalpha treatment of infiltrating versus resident tissue macrophage subsets in autoimmune arthritis

Synovial macrophages play a pivotal role in the pathogenesis of chronic autoimmune arthritis by contributing to local inflammation and tissue damage and are therefore a primary target for therapeutic intervention. The aim of the present study was to investigate in more detail the relative contribution of different synovial macrophage subsets with potentially different inflammatory or anti-inflammatory functions by analysing the two most frequent forms of human autoimmune arthritis, spondyloarthropathy (SpA) and rheumatoid arthritis (RA). Both infiltrating macrophages from peripheral blood expressing myeloid-related proteins (MRP) 8 and 14, and resident tissue macrophages expressing CD163 were abundant in inflamed synovium. Whereas the global number of synovial macrophages was similar in both diseases, infiltrating macrophages were increased in the RA lining layer in contrast with resident tissue macrophages, which were more frequently observed in SpA. Soluble MRP8/MRP14 complexes, which were secreted locally in the joint during the infiltration process, were increased in the serum of arthritis patients and, in contrast with soluble CD163 shed from resident tissue macrophages, correlated well with global inflammatory parameters. Treatment in vivo with anti-TNFalpha had a rapid and pronounced effect on the infiltration of MRP-positive macrophages into tissues, as evidenced by histopathological analysis and serum MRP8/MRP14 levels. Taken together, these data support an important role for infiltrating versus resident tissue macrophages in human autoimmune synovitis and indicate that macrophage products such as soluble MRP8/MRP14 complexes are valuable biomarkers for the experimental and clinical monitoring of specific disease mechanisms in vivo.     

异种脐血干细胞移植胶原性关节炎小鼠Bcl-2和Bax的表达*☆

背景：类风湿性关节炎的发生发展与滑膜细胞及淋巴细胞的增殖与凋亡不平衡密切相关,其滑膜细胞存在细胞凋亡过程的异常。 目的：观察异种、异基因双份脐血干细胞移植对Ⅱ型胶原性关节炎小鼠Bcl-2和Bax表达的影响。 方法：弗氏完全佐剂+Ⅱ型胶原诱导C57BL/6(H-2b)小鼠,建立Ⅱ型胶原性关节炎小鼠模型。小鼠二次免疫接种后第2天,模型组、正常对照组尾静脉注射生理盐水,细胞移植组将脐血造血干细胞注入小鼠尾静脉内(其中单份剂量2×106/50 g;双份剂量：每份1×106/50 g,共2×106/50 g)。甲氨喋呤阳性对照组小鼠灌胃甲氨蝶呤,每次0.017 5 g/kg,每5天1次,共6次。 结果与结论：移植后第42天全部处死动物取膝及肘以下关节组织,病理组织学显示正常对照组关节面光滑,滑膜层未见炎性细胞浸润,软骨细胞形态正常;模型组滑膜组织高度增生,大量的炎性细胞浸润,软骨面破坏;甲氨蝶呤阳性对照组、单份脐血造血干细胞移植组滑膜组织可见轻度增生,少量炎性细胞浸润,双份脐血造血干细胞移植组关节软骨表面光滑,未见破坏,有极少量炎性细胞浸润。免疫组织化学检测显示,双份脐血造血干细胞移植组Bax、Bcl-2的表达低于单份脐血造血干细胞移植治疗组(P < 0.05);与正常对照组比较,差异无显著性意义(P > 0.05)。结果说明双份脐血干细胞在一定数量及作用时间内可诱导Ⅱ型胶原性关节炎小鼠滑膜细胞凋亡,对滑膜组织损伤起保护作用。     

New Drug Targets in Rheumatoid Arthritis

Rheumatoid arthritis is a chronic inflammatory disease where the synovial tissue is characterized by heavy infiltration of leukocytes. Chemokines and chemokine receptors play an important role in cell migration and positioning of leukocytes within the inflamed rheumatoid synovium. There is now much focus on the specific contribution and role of each chemokine and chemokine receptor in the chronic inflammatory process in the synovial tissue. Recent evidence indicates that interference with the chemokines released from the inflamed synovial cells or the chemokine receptors expressed on the cells infiltrating the synovial tissue may lead to discovery of new therapeutics for this disease.     

The neurogenic contribution to synovial leucocyte infiltration and other outcome measures in a guinea pig model of arthritis

A neurogenic contribution to joint inflammation has been demonstrated in rat adjuvant arthritis, however as inflammatory mechanisms vary between species it is unclear whether these observations can be applied more generally. The aim of this study was to assess the neurogenic contribution to cellular infiltration and other outcome measures in a guinea pig model of arthritis. Compared to arthritic controls, animals pre-treated with capsaicin at doses sufficient to reduce sensory activity exhibited a significant attenuation of both mechanical and thermal hyperalgesia. Measures of inflammation, including swelling and radiological scores were also improved. Furthermore, capsaicin selectively reduced synovial T cell infiltration whereas no difference was seen with respect to synovial macrophages. These observations confirm a neurogenic component in guinea pig arthritis and indicate a selective sensory influence on T cell activity within the chronically inflamed joint. As T cells are strongly implicated in the pathogenesis of rheumatic disease, such an influence may serve to explain some of the clinical features observed in these disorders.     

类风关滑膜浸润T细胞特性与致病机制研究

为研究类风湿关节炎时关节滑膜浸润性T细胞生物学特性与致病机制 ,对 10例RA患者滑膜液中淋巴细胞的免疫表型、细胞因子分泌格局与趋化因子受体表达进行了分析。用双色荧光标记法分别测定滑膜液中和外周血淋巴细胞表型与趋化因子受体表达。用ELISA方法检测滑膜液与外周血中IFN γ、IL 10、IL 4与IL 12的含量。结果是滑膜液中的CD4 + T淋巴细胞为 4 0 0 %± 11% ,CD8+ T细胞为 34 0 %± 6 % ,CD4 + 与CD8+ T细胞的比值为 1 2 ,显著低于外周血中CD4 /CD8的比值。滑膜液中CD3和CD2 5双阳性的活化T细胞占 16 %± 6 0 %。趋化因子受体CCR5表达较低 ,与外周血无明显差异。但CX CR3表达水平较高 ,为 16 %± 4 0 % ,远远高于外周血 (仅为 0 5 %± 0 3% )。IFN γ在滑膜液中含量很高 ,达 (36 6 7± 4 3 2 )pg/ml,而外周血中含量仅为 (2 0 1± 3 2 )pg/ml。IL 4含量未能测得 (<15pg/ml ) ,与外周血相似。IL 12含量为 (4 19 9±89 2 )pg/ml,远高于外周血中的含量 (6 5 32± 34 2 )pg/ml。IL 10含量为 (187 7± 34 5 )pg/ml,高于外周血中的含量 (85±12 7)pg/ml。在所测细胞因子中 ,关节滑膜液中IFN γ和IL 12的含量与外周血相比具有显著的统计学差异。表明RA关节滑膜液中有相当数量的T细胞浸润。这些T细胞     

Arthritis induced by continuous infusion of hr-interleukin-1 alpha into the rabbit knee-joint

Continuous infusion of 200 ng/day hrIL-1 alpha for 14 days into knee-joints of rabbits leads to a severe arthritis of low aggressivity. This arthritis shows simultaneously characteristics of acute (serous and fibrinous exudation, polymorph infiltration, etc.) as well as chronic (synovial cell proliferation and fibrosis, pannus formation, cartilage and bone erosion, etc.) inflammation. The arthritis was associated with a distinct loss of metachromasia of the articular cartilage. These results indicate that IL-1 might play an important role in the induction and maintenance of arthritis.     

Experimental arthritis induced by continuous infusion of IL-8 into rabbit knee joints.

To determine the roles of IL-8 in inflammatory synovitis, examination was made of the results of continuously injecting human recombinant IL-8 into the knee joints of New Zealand while rabbits. Recombinant human IL-8 was infused continuously into the joint cavity at 75 ng/h for 14 days by a polypropylene catheter connected to a mini-osmotic pump implanted in each rabbit. Infiltration of inflammatory cells into joint cavity and histopathological changes in synovial tissue were examined at 7 and 14 days following the start of infusion. The continuous infusion of IL-8 for 14 days led to severe arthritis characterized by apparent erythema and joint pain, the accumulation of leucocytes, infiltration of mononuclear cells in synovial tissue, and marked hypervascularization in the synovial lining layer. IL-8 may be a factor which can contribute to the inflammatory process of chronic arthritis by mediating leucocyte recruitment and hypervascularization in inflamed joints.     

The morphogenesis of chronic synovitis in rheumatoid arthritis]

The synovial sheath obtained in synovectomy in 35 patients with rheumatic- and rheumatic-visceral forms of rheumatoid arthritis was studied histochemically and immunomorphologically. At early stages of exacerbation of the pathological process in the synovial tissue there were revealed predominantly catabolic processes: an increased permeability of vessels; mucoid oedema; fibrinoid changes in the subintimal layer. Further development of the disease was characterized by predominance of anabolic processes with proliferation of synoviocytes, subintimal histiocytes, productive vasculites, massive lymphoid-plasmocytic infiltration, diffuse, or in the form of lymphoid follicles. Using the immunofluorescent technique the authors revealed luminescence of the rheumatoid factor and gamma=globulin in plasmatic cells, extracellularly, and more rarely in macrophages. Pronounced immunological changes in the synovial sheath in the active course of rheumatoid arthritis were accompanied by a high level of metabolic processes and an intensive phagocytic reaction in synoviocytes and subintimal histiocytes. In observations with a low activity of rheumatoid arthritis the synovial tissue was characterized by low levels of enzymes of oxidative metabolism and hydrolysis, emptying of the capillary bed, processes of sclerosis, hyalinosis, amyloidosis.     

Latent Epstein-Barr virus (EBV) infection and cytomegalovirus (CMV) infection in synovial tissue of autoimmune chronic arthritis determined by RNA- and DNA-in situ hybridization.

In rheumatoid arthritis (RA) viral triggers, especially Epstein-Barr virus (EBV) and cytomegalovirus (CMV), have been suggested. By PCR analysis DNA of several viruses among which EBV, CMV, and parvovirus B19 (B19) has been detected in RA synovial fluid and synovial tissue. In 63 synovial tissues of 29 rheumatoid arthritis (RA), 6 psoriatic arthritis (PsA), 26 reactive arthritis/synovitis (rA/S), and two normal synovial cases, we recently could demonstrate a high percentage of replicative B19 infection within the synovial tissue, being significantly more frequent in autoimmune arthritis. To further investigate the influence of synovial virus infections in rheumatoid arthritis, we now analyzed the same sample of synovial tissues for CMV and EBV infections by DNA-in situ hybridization (CMV), EBER1/2-RNA-in situ hybridization (EBV), and immunohistochemistry. A significant latent EBV infection of synovial lining cells, synovial fibroblasts, and/or infiltrating lymphocytes was identified in 5/29 (17.2 %) RA, 1/6 (16.7%) PsA, and to a much lower degree in 1/26 (3.8%) rA/S specimens. CMV-DNA was detected in 31% of RA, 3/6 (50%) of PsA, and 11.5% of rA/S. Immunohistochemical analysis of CMV early antigen revealed replicative CMV activity in 20.7% of RA and 2/6 (33.3%) of PsA specimens but not in reactive arthritis synovia. Comparative analysis of the EBV-, CMV-, and published B19-data demonstrated that relevant synovial virus infections in general and furthermore double or multiple infections are far more common in autoimmune arthritis than in rA/S. A triple virus infection was found solely in RA in 10.3% of cases. The evidence of increased synovial persistence of EBV, CMV, or B19 either alone or even more as coinciding infections may further reinforce the notion of a primary role of these viruses in autoimmune arthritis.     

Arthritis in DBA/1 Mice Induced with Passively Transferred Type II Collagen Immune Serum

Arthritis was induced in DBA/1 mice by passive transfer of syngeneic anti-type II collagen (CII) serum concentrate. After transfer of serum containing 0.2 or 0.5 mg anti-CII auto-antibodies the first clinical signs of arthritis appeared 48 h after injection. Severe clinical arthritis was detected 96 h after injection. Immunohistochemical analyses of joints 48 h after serum injection revealed synovial foci in intercarpal and metacarpophalangeal joints of macrophage-like cells, expressing C3bi-receptors and major histocompatibility complex class II molecules, and infiltration of few CD4+ lymphocytes. Later (96 h after injection), the inflamed synovia were dominated by C3bi-receptor+ polymorphonuclear cells. In contrast to conventionally induced collagen arthritis (CIA), the inflammatory infiltrates, filling joint spaces and synovial tissue, were extensively dominated by polymorphonuclear cells, whereas macrophage-like cells expressing class II molecules and a few T cells were seen only in the periphery of the developing pannus. The anti-CII serum induced arthritis may be used as a model for studies of humoral mediated mechanisms operating in conventionally induced CIA as well as in rheumatoid arthritis.     

Anti-inflammatory Effect of Piperine in Adjuvant-Induced Arthritic Rats—a Biochemical Approach

The present study was undertaken to investigate the anti-inflammatory effect of piperine against adjuvant-induced arthritis in rats, an experimental model for rheumatoid arthritis and compared it with that of the non-steroidal anti-inflammatory drug indomethacin. Administration of heat-killed Mycobacterium tuberculosis (0. 1 ml) intradermally into the right hind paw of rats resulted in increased paw volume, lysosomal enzymes, glycoproteins and tissue marker enzymes and decreased body weight. However, these changes were reverted to near normal levels upon piperine (30 mg/kg body weight, i.p.) treatment. Histopathological analysis of joints also revealed that synovial hyperplasia and mononuclear infiltration observed in arthritic rats were alleviated by piperine. Thus, the present study clearly indicated that piperine possesses promising anti-inflammatory effect against adjuvant-induced arthritis by suppressing inflammation and cartilage destruction.     

塞来昔布对类风湿关节炎大鼠肿瘤坏死因子-α信号通路及滑膜细胞的影响

目的:探讨塞来昔布对类风湿关节炎(RA)大鼠肿瘤坏死因子-α(TNF-α)信号通路及滑膜细胞的影响.方法:将大鼠分为正常对照组,建立RA模型(RA组),建立RA模型后灌胃塞来昔布(实验组),RA组及实验组大鼠建立RA大鼠模型,检测大鼠关节炎症指数后,通过多种方法检测病理、滑膜细胞凋亡、TNF-α蛋白水平及阳性表达.结果...