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A comparison of normal and pathological synovial fluid.

Synovial fluid from 16 normal subjects was compared with that from 149 patients with a variety of rheumatic disorders. Normal fluid had fewer cells and a lower content of beta-glucuronidase than osteoarthritic samples. Particles, including occasional birefringent crystals, were seen in normal fluids as well as pathological samples. Alizarin red staining particles (presumed to contain apatite) were seen in all diagnostic groups; their numbers showed some correlation with radiological calcification in and around the joints and with a hypertrophic subchondral bone response. Lactate levels were highest in septic arthritis. No assay showed disease specificity.

Adult↗

Hidden 19S IgM rheumatoid factor in synovial fluid.

Synovial fluid (SF) of 16 adult patients with rheumatoid arthritis (RA) and 9 children with juvenile rheumatoid arthritis (JRA) was investigated for the presence of hidden 19S IgM rheumatoid factor (RF). SF was incubated with 500 IU hyaluronidase/g of SF, dialyzed against acetate buffers at decreasing pH, centrifuged and subjected to chromatography on a Sephadex G-200 column. The resulting IgM-containing fraction of SF from 5 seropositive patients (3 adult RA, 2 JRA patients) demonstrated RF activity by the complement-dependent hemolytic assay. Hidden 19S IgM RF was not found in the IgM-containing fractions of SF from 13 adult seronegative RA patients. Hidden 19S IgM RF was shown in the IgM-containing fraction of SF in 4 of 7 children with seronegative JRA.

Adult↗

Colony stimulating factor occurs in both inflammatory and noninflammatory synovial fluids.

Synovial fluids (SF) from patients with osteoarthritis (OA) and rheumatoid arthritis (RA) and various other arthritides were examined for the presence of colony stimulating factors (CSF). CSF was found in 7 of 13 (54%) SF from OA patients and in 8 of 12 (67%) SF from RA patients. It was also found in SF from patients with other arthropathies including 5 of 5 samples from patients with septic arthritis. Inhibition studies employing monospecific antisera indicated that in both RA and OA, CSF was of the macrophage type (M-CSF). While CSF was found in both inflammatory and noninflammatory effusions, significantly greater numbers of colonies were stimulated by RA SF than by OA SF and in general greater numbers of colonies correlated with higher SF leukocyte counts. Our data suggest that CSF as well as other cytokines may be involved in the perpetuation of joint destruction that occurs in various rheumatological conditions.

Arthritis, Rheumatoid↗

Cloning and recombinant expression of phospholipase A2 present in rheumatoid arthritic synovial fluid.

Synovial fluid from arthritic patients contains multiple forms of phospholipase A2 (PLA2), as resolved by high performance liquid chromatography (Seilhamer, J.J., Plant, S., Pruzanski, W., Schilling, J., Stefanski, E., Vadas, P., and Johnson, L. K. (1989) J. Biochem. (Tokyo), submitted for publication). Here we describe the cloning of a human 4.5-kilobase gene and 800-base pair cDNA encoding the form representing the major peak of activity and protein mass (peak A). The clones encode a mature peptide of 124 amino acids, which follows a prepeptide of 20 residues. The deduced amino acid sequence constitutes an enzyme of the "Type II" class of PLA2s, and resembles PLA2s from other mammalian sources. This represents the first report of a full length mammalian non-pancreatic PLA2 sequence. Active transcription of this PLA2 gene was detected in two different inflammatory cell sources. Recombinant human peak A PLA2 was expressed in vaccinia as a secreted protein which accumulated in conditioned medium.

Amino Acid Sequence↗

Thymocytotoxic antibodies in synovial fluid.

Synovial fluid (SF), obtained from patients with juvenile rheumatoid arthritis (JRA), osteochondritis, traumatic synovitis and septic arthritis were tested for the presence of thymocytotoxic activity. Such activity, directed against guinea pig thymocytes, was earlier demonstrated in human serum and shown to be mediated by IgM antibodies and a heat labile factor, most likely complement. The cytotoxic activity was demonstrated in all SFs tested, and was abolished by either one of the following procedures: depletion of IgM by use of anti-IgM-antibody coated protein A-Sepharose, or heating to 56 degrees C for 30 min. Activity was regained when samples inactivated by these two procedures were recombined. The cytotoxic SFs produced similar distortion of cell volume distribution of the target thymocytes as previously has been shown with human serum. The findings indicate that the two cytotoxic activities are identical. In most cases the IgM levels and the cytotoxic activity were lower in SF than in normal serum. Although the material does not permit correlation of cytotoxic activity with specific joint conditions, a high SF cytotoxicity was noted in two out of four patients with JRA. The demonstration of IgM-associated thymocytotoxic activity is of interest both in connection with earlier observations of various specific antibodies in SF and in connection with speculations regarding local immune regulatory activity in the joint.

Arthritis, Rheumatoid↗

T-cell receptor V-gene usage in synovial fluid and synovial tissue from RA patients.

The question of whether there is a preferential use of certain V genes in T cells entering an inflamed joint has hitherto been studied mainly using unfractionated cells from synovial fluid and tissue respectively, and no clear answer to the question has yet been provided. Concomitantly, evidence has been provided that the use of V genes may differ considerably between CD4+ and CD8+ T cells, and consequently that detection of biased V-gene expression within an inflammatory lesion may require separate analysis of the two T-cell subsets. In this paper we have therefore studied T-cell receptor V-gene expression in rheumatoid arthritis by means of double stainings of synovial fluid and blood for available anti-TCR monoclonal antibodies and antibodies to CD4 and CD8, respectively. Double stainings were also performed with anti-TCR antibodies and antibodies to activation markers HLA-DR and IL-2R. A certain bias towards the preferential use of certain V genes was seen particularly in the synovial fluid samples within both the CD4+ and CD8+ T-cell populations, but no uniform pattern was evident among the 35 patients investigated.

Arthritis, Rheumatoid↗

Acidosis of synovial fluid correlates with synovial fluid leukocytosis.

The antibacterial activity of aminoglycoside antibiotics is significantly reduced by lowering the pH of the incubation medium. Since gram-negative septic arthritis responds poorly to aminoglycoside antibiotic therapy, we sought to determine whether synovial fluid acidosis contributes to this poor outcome. Synovial fluid samples from 22 patients with various forms of acute and chronic arthritis were examined for white blood cell count and pH. A close correlation (r = -0.92, p is less than 0.001) between an increasing white blood cell count and a decreasing pH was demonstrated. Since septic arthritis is associated with high white blood cell counts, in synovial fluid, the resultant low pH may contribute to the poor response to gram-negative septic arthritis treated with aminoglycoside antibiotics.

Adult↗

Echovirus-associated polyarthritis. Report of a case with synovial fluid and synovial histologic characterization.

A patient with fever, myalgias, and acute polyarthritis in whom echovirus 9 was isolated from throat and rectal swab specimens is described. All symptoms resolved spontaneously over a one week period except for the arthritis which required three months for complete recovery. A mildly inflammatory synovial fluid and histologic picture were observed. Attempts to isolate the virus from synovial fluid and synovium during acute illness were unsuccessful.

Adult↗

A 10 year retrospective comparison of the diagnostic usefulness of synovial fluid and synovial biopsy examination.

BACKGROUND/AIMS: Synovial fluid examination is thought to be the pathological investigation of choice in most joint disorders, with only a few specific conditions necessitating biopsy, although no evidence based studies are available to support this belief. This study sought to investigate the validity of this assumption. METHODS: One hundred and three cases in which synovial fluid aspiration and synovial biopsy had both been performed at arthroscopy were studied. The amount of diagnostically useful information produced by each investigation was assessed. RESULTS: In most cases, both investigations provided the same amount of information and were generally equally specific or equally non-specific. Overall, the biopsy provided more information than the fluid in 29% of cases and vice versa in 18%. When only those cases in which both tests were adequate were considered, the biopsy provided more specific information than the fluid in a small number (9%) of cases, but these cases could not be predicted. CONCLUSION: The diagnostic usefulness of a biopsy approximates and occasionally exceeds that of a fluid. In the arthroscopic situation, the main advantage of performing both tests is that it provides a "failsafe mechanism" for the rare occasions when one of the samples is inadequate.

Arthritis↗

Differential expression of chemokine receptors on peripheral blood, synovial fluid, and synovial tissue monocytes/macrophages in rheumatoid arthritis.

OBJECTIVE: Since it is likely that monocytes utilize chemokines to migrate to the rheumatoid arthritis (RA) joint, we investigated the expression of C-C chemokine receptors (CCR) 1-6 and C-X-C receptor 3 (CXCR3) in the peripheral blood (PB), synovial fluid (SF), and synovial tissue of patients with RA as well as in the PB of normal subjects. METHODS: We compared chemokine receptor expression on CD14+ monocytes from normal PB, RA PB, and RA SF using 2-color flow cytometry. Correlations with patient clinical data were determined. Chemokine and receptor expression were investigated in RA synovial tissue by immunohistochemistry and 2-color immunofluorescence to identify CD68+ macrophages. RESULTS: Most normal PB monocytes expressed CCR1 (87%) and CCR2 (84%), but not CCRs 3, 4, 5, or 6 or CXCR3. RA PB monocytes expressed CCR1 (56%) and CCR2 (76%), with significantly more expressing CCR3 (18%), CCR4 (38%), and CCR5 (17%) compared with normal PB monocytes. Significantly fewer SF monocytes from RA patients expressed CCR1 (17%), CCR2 (24%), and CCR4 (6%) while significantly more expressed CCR3 (35%) and CCR5 (47%) compared with RA and normal PB monocytes; CCR6 and CXCR3 were rarely detected. Clinically, the erythrocyte sedimentation rate was inversely correlated with the expression of CCR1 and CCR4 by RA PB, and CCR5 expression by RA SF was correlated with the SF white blood cell count. CCR1-, CCR2-, and CCR5-immunoreactive cells were found in RA synovial tissue and colocalized with CD68+ macrophages. RA synovial tissue RANTES (regulated upon activation, normally T cell expressed and secreted chemokine)- and monocyte chemoattractant protein 1-immunoreactive cells colocalized with CCR1 and CCR2, respectively, on serial sections. Macrophage inflammatory protein 1alpha (MIP-1alpha) was principally restricted to vascular endothelium, and MIP-1beta+ macrophages were found throughout the sections. CONCLUSION: Monocytes mainly express CCR1 and CCR2 in normal and RA PB, CCR3 and CCR5 in RA PB and RA SF, and CCR4 in RA PB. The differential expression of chemokine receptors suggests that certain receptors aid in monocyte recruitment from the circulation while others are important in monocyte retention in the joint.

Adult↗

The elevated ratio of interferon gamma-/interleukin-4-positive T cells found in synovial fluid and synovial membrane of rheumatoid arthritis patients can be changed by interleukin-4 but not by interleukin-10 or transforming growth factor beta.

OBJECTIVES: To quantify the T-helper type (Th) 1 cytokine interferon gamma (IFN-gamma)-positive and the Th2 cytokine interleukin (IL)-4-positive cells in synovial fluid (SF) and synovial membrane (SM) at the single-cell level in rheumatoid arthritis (RA) in comparison to reactive arthritis (ReA), and to manipulate the cytokine pattern of RA patients in vitro. METHODS: Eighteen patients with RA and 17 with ReA were studied. For intracellular staining of cytokines, SF mononuclear cells (MNC) from seven patients with RA, in comparison to eight patients with ReA, were triple stained with anti-IFN-gamma, IL-4 and anti-CD4 or anti-CD8 monoclonal antibodies (mAb) and analysed by flow cytometry. Furthermore, in 13 patients with RA, immunohistology of SM was performed and compared with seven ReA patients. In addition, in six of the RA patients, synovial T cells were grown over 3 weeks in the presence of various cytokines and intracellular cytokine staining analysed by flow cytometry weekly. RESULTS: In SF, the mean percentage of IFN-gamma+/CD4+ T cells in RA was almost 4-fold higher than the number of IL-4+/CD4+ T cells (11.3+/-5 vs 3.02+/-1.04; P=0.0012), while the ratio of IFN-gamma/IL-4+ CD4+ T cells was only 1.59 in ReA (P=0.047 for the ratio difference). A similar result was obtained for SM: the ratio of IFN-gamma/IL-4+ cells in RA was 4.3 (P<0.0001 for the IFN-gamma/IL-4 difference), but only 1.2 for ReA (P=0.02 for the ratio difference). Of the CD3+ cells in SM, 2.8% were positive for IFN-gamma and 0.4% for IL-4 in three RA patients. A decrease in the number of IFN-gamma-positive SF T cells and an increase in the number of IL-4-positive SF T cells could be achieved in vitro through IL-4, but not by IL-10 or transforming growth factor beta. CONCLUSIONS: The Th1 pattern in the joint of RA patients demonstrated at the single-cell level may be important for the pathogenesis of RA and may provide a target for future immunotherapy. Our data suggest a therapeutic role for IL-4.

Adolescent↗

Stimulation by interleukin-7 of mononuclear cells in peripheral blood, synovial fluid and synovial tissue from patients with rheumatoid arthritis.

To determine how interleukin-7 (IL-7) affects the proliferation of T cells in patients with rheumatoid arthritis (RA), we evaluated the response of mononuclear cells (MNC) obtained from their peripheral blood (PB), synovial fluid (SF) and synovial tissue (ST) to stimulation by recombinant IL-7 and interleukin-2 (IL-2). Each cytokine was administered alone or combined with phytohemagglutinin (PHA). Cellular DNA synthesis was assayed by the [3H]-thymidine incorporation method. The stimulatory effect of 500 u/ml IL-7 on PBMNC obtained from 19 patients with RA was significantly lower than on PBMNC from 19 healthy controls. However, the same degree of stimulatory activity of 500 u/ml IL-2 was observed on the PBMNC from both RA patients and control subjects. The response of PBMNC to a suboptimal dose of PHA (0.2 micrograms/ml) was enhanced by adding either IL-7 or IL-2 (100 or 500 u/ml) to the cultures. The enhanced synthesis of DNA by both RA and control PBMNC on exposure to IL-7 following stimulation by a suboptimal dose of PHA was higher than that of IL-2. The effect of IL-7 on RA PBMNC was significantly greater than that of IL-2 at the concentration of 100 u/ml on PBMNC from the same RA patients. The stimulatory activity of IL-2 at the concentrations of 100 and 500 u/ml on SF MNC and ST MNC exceeded that of IL-7. In particular, an IL-2 dose of 500 u/ml had a marked effect on SF MNC. The PHA response of SF MNC was the lowest seen among the MNC from three different compartments.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Interferon production from peripheral blood, synovial fluid, and synovial tissue lymphocytes in patients with rheumatoid arthritis and ankylosing spondylitis.

A previous study demonstrated that interferon was present in the serum of 30% of the patients with systemic lupus erythematosus (SLE), which was significantly higher than the 4.5% found in normal controls. We also recently reported that interferon production was deficient from SLE mononuclear cells, which has been attributed to immunodeficiency of the lymphocytes. In this study, interferon measurement included lymphocytes obtained from peripheral blood (PB), synovial fluid (SF) and synovial tissue (ST) in patients with rheumatoid arthritis (RA) and ankylosing spondylitis (AS). PB from normal subjects (NS) was used as a control. The results showed with PHA stimulation, that the interferon level in PBL (L = lymphocyte) in NS (70.0 +/- 67.5) was significantly higher when compared with PBL in RA (27.9 +/- 21.6). However, there was no difference between PBL in NS and AS. With ConA stimulation, the interferon level was significantly higher in the PBL of NS (130 +/- 59) and as compared with the PBL in RA (83.6 +/- 53.5). The SFL in RA (67.8 +/- 31.1) and the STL in RA (77.2 +/- 93.2) were also significantly different. It is concluded that interferon production was deficient not only in PBL in RA, but also in SF and STL in RA. The reduced interferon production from PB, SF and ST lymphocytes in RA patients may be due to previous release or immunodeficiency. Lymphocyte interferon production was normal in AS, which suggests that the lymphocyte abnormality between RA and AS may be different.

Adult↗

Effects of intraarticular corticosteroid and sodium hyaluronate injections on synovial fluid production and synovial fluid content of sodium hyaluronate and proteoglycans in normal equine joints.

Hyaluronate and cartilage proteoglycan fragments in synovial fluid from normal equine joints were determined after repeated intraarticular injections of beta-methasone, methylprednisolone, hyaluronate, saline and a combination of beta-methasone + hyaluronate and methylprednisolone + hyaluronate. The experimental intraarticular treatments were given at the beginning of the study and after one and two weeks. Synovial samples were obtained at the onset of the study and after 1, 2, 3, 4, 6, and 8 weeks. The synovial volume was determined and showed no significant changes after the repeated arthrocenteses and the experimental treatments. Corticosteroid injections resulted in increased levels of hyaluronate in the synovial fluid which indicated a stimulatory effect on the synoviocytes to produce hyaluronate. The corticosteroids also resulted in high levels (up to 20 times the initial level) of proteoglycans which indicated joint cartilage degradation. A combined injection of corticosteroids and high molecular weight hyaluronate reduced proteoglycan breakdown which indicated a protective effect of hyaluronate on the articular cartilage.

Adrenal Cortex Hormones↗

Subcellular particles in synovial fluids and synovial cells.

Membranous particles of mycoplasma-like appearance (100 to 300 nm in diameter) and non-membranous particles which are viral-like (20 to 80 nm in diameter) were observed in negatively-stained preparations of synovial fluid pellets from patients with rheumatoid arthritis (RA), osteoarthrosis (OA) or psoriatic arthritis (PA). The incidence of positive fluids were: membranous particles--22/28 (RA), 7/11 (OA), 1/3 (PA); non-membranous particles--12/28 (RA), 1/11 (OA), 1/3 (PA). Eight of the RA fluids, one OA and one PA fluid were positive for both types of particle. The observation of an increased frequency of viral-like particles in RA synovial fluids compared to OA and PA fluids is of possible aetiological interest. Viral-like particles were also observed in the nuclei of rheumatoid synovial macrophage syncytia by electron microscopy of ultra-thin sections of cultured cells.

Arthritis↗

Interleukin-6 activity in paired samples of synovial fluid. Correlation of synovial fluid interleukin-6 levels with clinical and laboratory parameters of inflammation.

Paired synovial fluid (SF) samples obtained from the knees of 12 arthritis patients were studied to establish a relation between parameters of local inflammatory activity and SF interleukin-6 (IL-6) levels. Local disease activity was scored using joint temperature, swelling and pain as clinical parameters of inflammation. SF samples were assayed for laboratory parameters of inflammation such as leucocyte content, the percentage polymorphonuclear cells, the pH, and for immunoglobulin levels (IgG, IgM). SF IL-6 concentrations were determined using the B9-bioassay. Within individual patients the local activity of inflammation as measured using clinical parameters was found to be related to the local SF IL-6 level. When considering the total group of patients, a correlation (P less than 0.001) was found between the clinical parameters of local inflammation and the SF IL-6 levels. Furthermore, IL-6 levels were found to correlate with leucocyte counts (P less than 0.02), the percentage of polymorphonuclear cells (P less than 0.10), the pH value (P less than 0.01), but not with SF IgM and IgG concentrations.

Adult↗

Ultrastructural demonstration of spirochetal antigens in synovial fluid and synovial membrane in chronic Lyme disease: possible factors contributing to persistence of organisms.

To perform the first systematic electronmicroscopic (EM) and immunoelectron microscopy (IEM) study of the pathological changes and the evidence of spirochete presence in synovial membranes and synovial fluid (SF) cells of patients with chronic Lyme arthritis. EM examination was performed on four synovial membrane and eight SF cell samples from eight patients with chronic Lyme disease. Spirochetal antigens in the samples were sought by IEM using monoclonal antibody to Borrelia burgdorferi outer surface protein A (OspA) as the immunoprobe. Prominent ultrastructural findings were surface fibrin-like material, thickened synovial lining cell layer and signs of vascular injury. Borrelia-like structures were identified in all four synovial membranes and in two of eight SF cell samples. The presence of spirochetal antigens was confirmed by IEM in all four samples studied (one synovial membrane and three SF cell samples). OspA labelling was in perivascular areas, deep synovial stroma among collagen bundles, and in vacuoles of fibroblasts in synovial membranes; and in cytophagosomes of mononuclear cells in SF cell samples. Electron microscopy adds further evidence for persistence of spirochetal antigens in the joint in chronic Lyme disease. Locations of spirochetes or spirochetal antigens both intracellulary and extracellulary in deep synovial connective tissue as reported here suggest sites at which spirochaetes may elude host immune response and antibiotic treatment.

Adolescent↗

Enrichment of differentiated CD45RBdim,CD27- memory T cells in the peripheral blood, synovial fluid, and synovial tissue of patients with rheumatoid arthritis.

OBJECTIVE: To delineate in greater detail the phenotype of T cells that reside in the synovial tissue (ST) and synovial fluid (SF) of patients with rheumatoid arthritis (RA), in order to determine their precise differentiation status, and to determine whether the accumulation of these specific T cell subsets in these synovial compartments could be related to their capacity for transendothelial migration. METHODS: Lymphocytes from normal subjects or from the peripheral blood (PB), ST, and/or SF of RA patients were phenotypically analyzed by flow cytometry. Normal PB CD4+ T cells were also characterized using an in vitro assay of transendothelial migration. RESULTS: ST and SF were found to be enriched with memory (CD45RA-,CD45RO+,CD11abright,CD44bright and activated (CD69+) T cells. Moreover, ST and SF cells from RA patients were enriched in differentiated CD4+,CD45RBdim,CD27- T cells, a subset of mature memory T cells that develops after prolonged antigenic stimulation. In addition, PB of some RA patients contained an increased number of CD4+,CD45RBdim,CD27- T cells. The CD4+,CD11abright,CD44bright memory T cells, which included the CD45RBdim,CD27- more mature memory cells, exhibited an enhanced capacity for transendothelial migration that is likely to contribute to their enrichment in the rheumatoid synovium. CONCLUSION: RA patients manifest an increased number of mature memory T cells in the SF and ST, and some also have an increased number of these cells in PB that is likely to reflect chronic antigenic stimulation. The enrichment of these cells in the SF and ST reflects, in part, an enhanced capacity to migrate from the vascular space into inflamed tissue.

Arthritis, Rheumatoid↗