Removal of hyaline articular cartilage reduces lymphocyte infiltration and activation in rheumatoid synovial membrane.

OBJECTIVE: To analyze the effect of removal of hyaline articular cartilage on synovial membrane pathology in chronic arthritis. METHODS: Synovial membrane samples were obtained from patients with rheumatoid arthritis or ankylosing spondylitis in association with total hip arthroplasty, either primary or revision surgery. Synovial membrane histopathology was assessed by immunochemical staining and morphometry. RESULTS: CD68 positive macrophages were common in revision synovial membranes. In contrast, T lymphocytes were much more common in primary rheumatoid synovial membranes (p < 0.001). Many T lymphocytes in primary synovial membrane were HLA-D/DR positive (p < 0.001) and interleukin 2 receptor (IL-2R) positive (p < 0.001) and contained interferon-gamma(IFN-gamma; p < 0.001) and tumor necrosis factor-beta (TNF-beta; p < 0.001). In contrast, revision synovial membranes from patients with chronic arthritis contained only a few HLA-D/DR positive T cells and practically no IL-2R, IFN-gamma, or TNF-beta positive activated T lymphocytes. CONCLUSION: The components of hyaline articular cartilage may be the source of autoantigen responsible for perpetuation of chronic arthritides.     

Elevated levels of collagenase and prostaglandin e2 from synovium associated with chronic lyme arthritis

A patient with chronic Lyme arthritis and roentgenographic evidence of bony erosion underwent a synovectomy; proliferative synovium (pannus), containing aggreagates of small lymphocytes, was found adherent to eroded cartilage and bone. During 8 days in tissue culture, the synovial cells produced large amounts of collagenase and prostaglandin E2, but only low levels of both neurtral and acid proteinases. Sixty-seven percent of the lymphocytes from the synovium were T cells; 19% were B cells. Attempts to identify agent/antigen in the synovial cells were unsuccessful. Thus, the synovium of this patient, whose disease appears to be tick-transmitted, resembles that of rheumatoid arthritis. This finding further supports the hypothesis that many possible agents, including infectious ones, trigger a common pathway in synovium, which leads to joint destruction.     

Expression of CD44 on rheumatoid synovial fluid lymphocytes.

OBJECTIVES--To investigate the involvement of the adhesion molecule CD44 in the homing of lymphocytes to synovial tissue, by examining the density of expression and molecular mass of CD44 on rheumatoid synovial fluid lymphocytes. METHODS--Twenty patients with rheumatoid arthritis were studied. Peripheral blood and synovial fluid lymphocytes were isolated by Ficoll-Hypaque sedimentation. CD44 expression was analysed by two colour flow cytometry of CD3 positive T lymphocytes with calculation of mean fluorescence intensity. Expression of activation markers M21C5, M2B3, interleukin (IL)-2 receptor and transferrin receptor was quantitated. In addition, CD44 molecular mass was examined by Western blot in six patients. RESULTS--CD44 expression was markedly increased on synovial fluid T lymphocytes of rheumatoid patients relative to peripheral blood lymphocytes from the same individuals. CD44 molecular mass on peripheral blood mononuclear cells was 88 kDa, but that on synovial fluid lymphocytes was only 83 kDa. CD44 expression correlated significantly with expression of activation markers M21C5, M2B3, and the IL-2 receptor. CONCLUSIONS--Alterations in density of expression or of the molecular mass of CD44 could contribute to local tissue injury, either directly by facilitating adhesion, or indirectly through effects on other adhesion molecules.     

Quantitative analysis of immunohistologic features of very early rheumatoid synovitis in disease modifying antirheumatic drug- and corticosteroid-naïve patients

OBJECTIVE: To describe the immunohistochemical features of very early rheumatoid synovitis in disease modifying antirheumatic drug- and corticosteroid-na?ve patients. METHODS: Eight patients presenting with oligoarthritis or polyarthritis, who later met American Rheumatism Association criteria for rheumatoid arthritis (RA), underwent needle synovial biopsies of a knee joint within the first 6 weeks after onset of disease symptoms. Using antibodies to CD3, CD8, L26, CD68, and von Willebrand factor, a detailed quantitative immunohistochemical analysis was done. RESULTS: CD3+ T lymphocytes, CD8+ T lymphocytes, L26+ B lymphocytes, and CD68+ macrophages were seen in 8/8 (100%), 7/8 (87%), 4/8 (50%), and 6/6 (100%) of synovial biopsies stained with the respective marker. There was a wide variation in number of positive cells between patients. CD3+ and CD8+ T cells were seen predominantly in perivascular areas, less often in a diffuse distribution, and not in aggregates. L26+ B lymphocytes were found in much smaller numbers compared with T lymphocytes. A mean of 67 vessels/mm2 was noted. No lymphoid aggregates were seen. In all cases, infiltration of macrophages and lymphocytes was limited mainly to relatively superficial parts of synovium, i.e., within 1 to 2 high power fields of the surface. CONCLUSION: An absence of lymphoid aggregates or dramatic vascularity and a predominantly superficial infiltrate consisting mainly of perivascular T cells with few B cells characterized our patients with very early RA of < 6 weeks' duration. Thus, there appear to be some potentially important differences from findings reported in well established disease.     

Silencing of CD21 expression in synovial lymphocytes is independent of methylation of the CD21 promoter CpG island

The complement receptor II (CD21) recognises the complement component C3d of immune complexes. Expression of the CD21 gene is tightly regulated during B lymphocyte differentiation. Only mature B lymphocytes express CD21 but not pro-, pre-, or plasma B lymphocytes. Previously we found that pro-, pre-, and intermediate B lymphocytes contain a methylated CpG island and do not express CD21. CD21-expressing mature B lymphocytes, plasma B lymphocytes, and nonlymphoid cells carried a demethylated CD21 CpG island. Furthermore, we found that synovial lymphocytes from patients with rheumatic disease show reduced expression of CD21. This observation tempted us to analyse the methylation status of the CD21 CpG island in peripheral blood mononuclear cells and synovial fluid mononuclear cells derived from these patients. While methylation is involved in silencing CD21 in early types of B lymphocytes, we found the CD21-CpG island to be demethylated in peripheral blood mononuclear cells and synovial fluid mononuclear cells of patient DNA.     

Arthroscopic and immunohistologic characterization of knee joint synovitis in osteoarthritis

We studied 10 patients who had arthritis of the knee joint, but no other signs of rheumatic disease. The clinical diagnosis of osteoarthritis was corroborated by arthroscopic evidence of characteristic cartilage degeneration. Signs of inflammation were confined to areas of the synovial membrane that lay near the cartilage; thus, the major part of the joint cavity was not affected. The intensity of the synovial inflammation varied within the areas involved, but was always most pronounced in regions rimming the cartilage. Biopsy samples selected from regions of intensely inflamed synovium contained foci of T lymphocytes, which were bordered by immunoglobulin-carrying B lymphocytes and plasma cells, as well as strongly HLA-DR positive dendritic-like cells adjoined to alpha Leu-3a+ T helper lymphocytes. In tissue samples taken from macroscopically noninflamed areas, only a few infiltrating lymphocytes were seen. Thus, the inflammatory synovial changes found in osteoarthritis appear to be anatomically restricted and of varied intensity but, when present, are microscopically indistinguishable from the changes that have been previously described as indicative of rheumatoid arthritis.     

Cells expressing dendritic cell markers are present in the rheumatoid nodule

OBJECTIVE: To determine if dendritic antigen-presenting cells (DC) are present in rheumatoid nodules, as has been reported in the synovial lesions of rheumatoid arthritis. METHODS: Nodules (n = 14) were examined with monoclonal antibodies (Mab) recognizing the DC differentiation/activation markers CD83, CMRF44, and CMRF56 and an antibody recognizing the CD1a antigen present on epithelial tissue associated DC. Results. Cells expressing CMRF44 were common in rheumatoid nodules, comprising 22% of nucleated cells versus 13% in synovial membranes (n = 10). Cells positive for CD1a (5%) and CD83 (2%) were less common. A majority (86%) of CMRF44 positive cells were also positive for the macrophage marker CD14. This left a significant minority of putative DC that were single stained with CMRF44. CONCLUSION: Cells bearing DC markers are as frequent in the rheumatoid nodule as in the synovial lesions. A majority are "indeterminate" cells that are CD14 positive but a proportion are single stained putative DC. The lack of lymphoid collections containing DC and T and B lymphocytes in the nodule suggests that local presentation of antigen may not occur in the rheumatoid nodule, as is thought to be the case in synovial membranes containing lymphoid follicles. This difference could potentially be explained by different states of activation, and differentiation of DC within the 2 lesions.     

Deficiency of the suppressor inducer subset of T lymphocytes in rheumatoid arthritis

New monoclonal antibodies allow CD4 lymphocytes to be categorized into helper/inducer (HI) CD4+ 4B4+, and suppressor inducer (SI) CD4+ 2H4+ subpopulations. Blood and synovial lymphocytes from 30 patients with rheumatoid arthritis (RA), 13 patients with other articular diseases, and 24 control subjects were exposed to these monoclonal antibodies and analyzed by flow cytometry. CD4:CD8 ratios were similar for the 3 groups. In RA patients, there was a depletion of SI cells in blood and synovial fluid, and the ratio of HI cells to SI cells was elevated, particularly in the synovial lymphocytes. These data provide new evidence for T cell dysregulation in the perpetuation of RA.     

HTLV-I associated arthritis: characteristics of an HTLV-I virus infected T cell line from synovial fluid.

A T cell line from mononuclear cells in the synovial fluid of a patient with polyarthritis was established. The T cell line reacted with serum samples positive for antibodies to human T cell lymphotropic virus type I (HTLV-I) and with monoclonal antibody to HTLV-I p19. In Southern blotting with an env-pX-LTR HTLV-I probe and digestion of T cell line DNA with the restriction enzymes ClaI, DraI, and PstI generated fragments that were identical to those found in two HTLV-I infected T cell lines established from adult T cell leukaemia or HTLV-I associated myelopathy. The T cell line expressed CD2, CD3, CD4, CD45RA, CD29, HLA-DR, CD25, and CD26 antigens, but not CD8 and CD20 antigens. Large amounts of interleukin 6, interferon gamma, and tumour necrosis factor alpha were secreted in the culture supernatants of this cell line. This line helped immunoglobulin production by B cells, but not K562, Raji, and synovial cell lysis.     

Candida arthritis: cellular immune responses of synovial fluid and peripheral blood lymphocytes to Candida albicans.

A case of septic Candida albicans arthritis of the knee in a patient with systemic candidiasis is presented. Systemic and intra-articular cellular immune responses to C albicans and various bacterial antigens were monitored for 15 weeks. It is shown that the candida induced blastogenesis of synovial fluid lymphocytes was much more stimulated than that of peripheral blood lymphocytes, and that the proportion of activated cells expressing HLA class II antigens was markedly increased in the synovial fluid. Strong cellular immune responses to Candida albicans could still be shown many weeks after the synovial fluid aspirates had become sterile. For the first time synovial fluid derived, CD4 positive T lymphocyte clones with specificity for candida antigens were characterised and further propagated in vitro.     

Presence of autoimmunity to pancreatic antigens in a patient with fibrocalculous pancreatic diabetes.

A case of fibrocalculous pancreatic diabetes (FCPD) is reported for which antibody and cellular immune characteristics were determined. The patient, a Thai woman, had serum islet cell antibodies (ICA) that were detected by both immunoperoxidase staining and an indirect enzyme-linked immunosorbent assay (ELISA). Serum anti-human insulin antibodies were negative by a displacement ELISA. Lymphoproliferation assay against pancreatic antigen prepared from a blood group O cadaveric donor was positive. Increased CD8+ lymphocytes were observed using direct immunofluorescence staining and flow cytometry. CD4+ T lymphocytes, B lymphocytes and NK cells were within normal levels. These findings provide evidence for autoimmunity to pancreatic antigens in a patient with fibrocalculous pancreatic diabetes.     

Cathepsin B and its endogenous inhibitor cystatin C in rheumatoid arthritis synovium

OBJECTIVE: To compare the expression of cathepsin B and its endogenous inhibitor cystatin C in synovial tissue of patients with rheumatoid arthritis (RA) and to determine the cell type expressing cystatin C. METHODS: The expression of cathepsin B and cystatin C was studied by immunohistochemistry in synovial tissue of 10 patients with RA and compared to healthy controls. Applying double labeling methods, the expression of cathepsin B was compared to that of cystatin C. To determine the cell type expressing cystatin C, double labeling with anti-CD68 (PG-M1) was performed. RESULTS: Both cystatin C and cathepsin B were strongly expressed in synoviocytes of patients with RA. Furthermore, fibroproliferative tissue at the site of cartilage and bone destruction contained fibroblast-like and macrophage-like cells positive for cystatin C and cathepsin B, whereas normal synovial tissue exhibited only limited expression of these molecules. Osteoclasts revealed positive staining for CD68 and cystatin C, but not for cathepsin B. CONCLUSION: Cystatin C is a product of both macrophage-like and fibroblast-like synoviocytes. The strong expression of both the matrix degrading cysteine proteinase cathepsin B and the cysteine proteinase inhibitor cystatin C in rheumatoid synovium, particularly at the sites of bone and cartilage erosion, suggests that cystatin C--although increased--is not sufficient to prevent matrix degradation by cathepsin B.     

Inflammatory infiltrate and interleukin-8 expression in the synovium of psoriatic arthritis – an immunohistochemical and mRNA analysis

Psoriatic arthritis is an inflammatory arthropathy that ultimately can lead to joint destruction. In this study, we investigated the immunophenotype of the inflammatory cells and the expression of interleukin-8 (IL-8), which is the hallmark chemoattractant cytokine of psoriasis in synovial membranes from patients exhibiting active psoriatic synovitis (n=9). The tissue samples were examined by immunohistochemistry, Western blot analysis and in situ hybridisation. The inflammatory infiltrate consisted predominantly of CD3⁺ T lymphocytes, with a higher proportion of CD4⁺ than CD8⁺ T lymphocytes in six cases. CD3⁺ T lymphocytes were focally distributed near small blood vessels and the enlarged synovial intima. CD1⁺ interdigitating reticulum cells were not detected. CD22⁺ B lymphocytes and plasma cells were found in small aggregates without KiM4⁺ follicular dendritic cells. KiM8⁺ macrophages were located in the synovial intima and were distributed in a diffuse pattern near the synovial lining cells. CD15⁺ neutrophil granulocytes were detected in four cases. They were preferentially located in the vicinity of blood vessels and the synovial intima. IL-8 was found at a high level in the synovial lining cells and to a lesser extent in cells located in the perivascular areas. Immunofluorescence double staining showed IL-8 to be expressed in KiM8⁺ multinucleated giant cells, KiM8⁺ macrophages and CD3⁺ T lymphocytes. IL-8 receptor A was demonstrated in the synovial lining and in macrophages and lymphocytes. IL-8 was detected by immunoblot analysis of the synovial tissue at 8.4 kD. Employing in situ hybridisation, IL-8 mRNA was strongly and preferentially expressed in the synovial intima, as well as in macrophages and lymphocytes. The immunophenotype of the psoriatic arthritis inflammatory cells shows great similarity to the inflammatory infiltrate found in the synovial tissue of patients with rheumatoid arthritis. The preferential expression of IL-8 and IL-8 mRNA in the enlarged synovial intima and in lymphocytes and macrophages suggests that IL-8 exerts its action through activated mononuclear cells and T lymphocytes. It seems to play a role in regulating leucocyte traffic into the enlarged synovial intima and may contribute to the aggressive synovitis of patients with psoriatic arthritis. Received: 14 August 1997 / Accepted: 25 August 1997     

Lymphocyte surface marker expression in rheumatic diseases: evidence for prior activation of lymphocytes in vivo.

Expression of major histocompatibility complex (MHC) class II and other lymphocyte activation markers on peripheral blood and synovial fluid T lymphocytes from patients with rheumatoid arthritis (RA), psoriatic arthritis, and Reiter's syndrome were measured and the mean fluorescence intensities of these antigens were assessed. Increased expression of MHC class II antigens of synovial fluid T lymphocytes is not unique to RA, though it is quantitatively greater on RA synovial fluid T cells. There was less expression of other lymphocyte activation markers (4F2, transferin receptor) and a marked discordance between the expression of these markers and the interleukin 2 receptor (IL2r). Synovial fluid T lymphocytes contain a subpopulation of larger cells expressing MHC class II and other lymphocyte activation antigens with the exception of the IL2r. Mean fluorescence intensity of CD3 and CD4 antigens on synovial fluid T lymphocytes was decreased in all three patient groups, suggesting prior in vivo exposure of synovial fluid T lymphocytes to an unknown antigen.     

Cobalt-specific T lymphocytes in synovial tissue after an allergic reaction to a cobalt alloy joint prosthesis

Metals such as cobalt and nickel are common contact allergens. We studied the mechanisms underlying an allergic reaction with marked synovial inflammation in a patient with a cobalt alloy arthroplasty. After removing the joint prosthesis the adjacent synovial tissue was examined for cobalt-specific T lymphocytes. Synovial membrane mononuclear cells were expanded in interleukin 2 and cloned using a representative cloning protocol. T cell clones were tested for their proliferative response to cobalt and further characterized with regard to cytokine secretion, phenotype, and HLA restriction. Additionally, synovial fibroblasts were tested for their function as antigen presenting cells (APC). Almost 30% of the T cell clones reacted to cobalt, but not to the control nickel. All these T cell clones were CD4 positive. The cobalt induced proliferative response could be blocked by anticlass II antibodies. Also, synovial fibroblasts expressing class II molecules induced by interferon-gamma were able to serve as APC. However, when testing a panel of APC of HLA class II mismatched donors, no requirement for a certain HLA class II molecule could be defined. Further studies are necessary to determine mechanisms of presentation and recognition of cobalt by T lymphocytes, a prerequisite for improved prevention and treatment of metal induced allergic reactions.     

Novel AICDA mutation in a case of autosomal recessive hyper-IgM syndrome,growth hormone deficiency and autoimmunity

BackgroundThe Hyper-immunoglobulin M syndromes (HIGM) are a heterogeneous group of genetic disorders, which have been rarely reported to be associated with growth hormone deficiency (GHD).Methods and resultsA nine-year-old girl with recurrent urinary tract infections, diarrhoea, sinopulmonary infections, and failure to thrive since the age of six months had normal CD3+, CD4+, CD8 + T lymphocytes, and CD19 + B lymphocytes and natural killer (NK) cells, but extremely elevated IgM and significantly decreased IgG and IgA. In view of the patient's short stature, growth hormone evaluation was carried out and growth hormone deficiency established. The patient underwent Ig replacement therapy and received growth hormone therapy in addition to antibiotics and responded well. Furthermore, the patient developed benign cervical lymphadenopathy, as well as elevated erythrocyte sedimentation rate, positive autoantibodies to SSA-Ro, and severely dry eyes, which partially responded to both the punctate occlusion and systemic corticosteroids, at the age of seven years. Sequencing analysis of the exons from activation-induced cytidine deaminase (AICDA) gene revealed that the patient was homozygous for a single T to C transversion at position 455 in exon 4, which replaces a Valine with an Alanine.ConclusionsTo our knowledge, this is a new AICDA mutation, which has not been reported previously in HIGM. The mutation analysis could improve diagnosis of HIGM patients and also elaborating on the spectrum of AICDA mutations.     

Elbow synovitis related to an intraarticular osteoid osteoma of the humerus, with immunologic and histochemical studies.

An 18-year-old boy presented with elbow synovitis. Investigations disclosed an osteoid osteoma of the coronoid fossa confirmed by histology. The synovium appeared hypertrophic with histologic patterns resembling those seen in synovitis in rheumatoid arthritis. Immunohistochemistry showed lymphoid follicles composed of B and T cells. T lymphocytes were mainly of the CD4 phenotype, showing soluble interleukin 2 receptor (IL-2r) in places but were poorly positive for DR antigen. C3, C4, B factor and CH50 activity were decreased and interleukin 1 and soluble IL-2r were increased in synovial fluid. They were normal in peripheral blood except for a slight decrease in C4. These data suggest a local immunologic activation induced by osteoid osteoma, the mechanism of which remains hypothetic. Immunomodulating mediator diffusion from osteoid osteoma itself or as a secondary response to tumoral antigen release could be advocated. Whether such phenomena are specific to the epiphyseal location of osteoid osteoma needs clarification.