calcimycin and Arthritis--Rheumatoid

The studies reported here were designed to examine the chemotactic potential, arachidonic acid (AA) metabolism and phospholipid transmethylation in peripheral blood neutrophils from patients with rheumatoid arthritis (RA): a) prior to treatment with BW 91Y (3-deazaadenosine), b) after 4 weeks when half the patients were on active medication and half were on placebo, and c) after 4 weeks at which time all patients were on active medication. The authors demonstrate that BW 91Y in vitro at 600 pg/ml caused a decrease in chemotactic potential as measured by the leading front (LF) assay in neutrophils from both normal volunteers (p less than 0.025) and RA patients. They also demonstrate that BW 91Y caused a significant increase in production of [3H]LTB4 (LTB = leukotriene B) in ionophore-stimulated neutrophils from both normal (p less than 0.025) and RA patients (p less than 0.050) as compared to initial values. BW 91Y caused decreased incorporation and percent distribution of [3H]AA into phosphatidylcholine (PC), with a resultant increase in percent distribution into phosphatidylethanolamine (PE). There was also an increased release of [3H]AA from the PE fraction in BW 91Y-treated cells in response to ionophore stimulation. BW 91Y was found to exhibit a dose-dependent (10(-7) to 10(-4) g/ml) inhibition of the uptake and incorporation of L-[methyl-3H]methionine into the cellular lipids, while at low doses (10(-9) to 10(-5) g/ml) it stimulated the significant uptake and incorporation of [methyl-14C]choline chloride into PC. Although the total cellular content and percent composition of PC remained unchanged, it was found that BW 91Y caused a slight decrease in PC plasmalogens and an apparent increase in the 1,2-diacyl-glycerophosphatidylcholine (-GPC). BW 91Y was found, however, to have no effect on the amount or stimulated metabolism of the ether-linked 1-O-alkyl-2-acyl-GPC. As further evidence for this, the authors demonstrate that BW 91Y has no effect on the ionophore-stimulated production of [14C]acetate-labelled 1-O-alkyl-2-acetyl-GPC, or [14C]PAF.

Topics: Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Chemotaxis, Leukocyte; Chromatography, Thin Layer; Dose-Response Relationship, Drug; Double-Blind Method; Humans; Leukotriene B4; Lipid Metabolism; Lipids; Neutrophils; Phosphatidylcholines; Phosphatidylethanolamines; Phospholipids; Platelet Activating Factor; Tubercidin

Arachidonic acid (AA) metabolism and phospholipase A2 (PLA2) activity were measured in the peripheral blood polymorphonuclear leukocytes (PMNL) from ten patients with rheumatoid arthritis (RA) on treatment with various non-steroidal anti-inflammatory agents (NSAIA). AA metabolism and PLA2 activity were measured both initially and after treatment with either placebo or Clotrimazole, a broad spectrum anti-mycotic agent, as a possible anti-rheumatic drug. AA metabolism was also measured in PMNL from ten patients with active RA untreated with any NSAIA and ten normal volunteers. Using 3H-AA prelabeled cells, we show that there was a significantly higher (P less than 0.025) production of 3H-LTB4 in response to stimulation with the calcium ionophore A23187 in untreated RA patients than in normal volunteers (mean +/- S.D.:4.8 +/- 1.6% and 3.1 +/- 1.0%, respectively). The production of 3H-LTB4 by PMNL from patients on NSAIAs was less elevated (mean +/- S.D.:4.1 +/- 1.5%) and was not significantly different from normal controls. Concurrently we examined PLA2 activity in PMNL-sonicates from ten of our study patients using autoclaved [14C]oleate-labeled E. coli biomembranes as an exogenous substrate. Using linear regression analysis, we demonstrate a significant correlation between in vitro PLA2 activity and the release of 3H-AA from the cellular phospholipids (deacylation) in response to A23187 stimulation (r = -0.526, P less than 0.025). We also demonstrate significant correlations between the overall clinical state of the RA patient, as evaluated by a modified rheumatoid activity index (MRAI), and both the release of 3H-AA from the cellular phospholipids and its production of total [3H]eicosanoids (r = -0.557, P less than 0.025 and r = 0.644, P less than 0.005, respectively). This data suggests that: PLA2 activity may, in part, account for the higher generation of LTB4 by RA PMNL; NSAIAs may be capable of modulating this abnormality; and Clotrimazole may affect the clinical or laboratory data of RA patients already on treatment with NSAIA.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Clotrimazole; Humans; Imidazoles; Leukotriene B4; Neutrophils; Phospholipases A; Phospholipases A2

To determine the cytokine profile of CD4+ T cells in the synovial tissue (ST) of rheumatoid arthritis (RA) patients at the single-cell level.. Unseparated ST cells and paired CD4+ T cells separated from the peripheral blood (PB) and ST of RA patients were stimulated for 4 hours with phorbol myristate acetate (PMA) plus calcium ionophore A23187, or for 6 hours with immobilized anti-CD3 plus anti-CD28, in the presence of brefeldin A. Cells were stained for intracellular cytokines such as interferon-gamma (IFNgamma), interleukin-2 (IL-2), IL-4, IL-10, and IL-13, in combination with cell surface markers. The percentages of cytokine-producing T cells were analyzed by flow cytometry.. When ST cells were stimulated with PMA plus A23187 in bulk culture, IFNgamma-producing T cells were more frequently detected in the CD8+ subset, but cells producing other cytokines were found in the CD4+ subset. Purified ST CD4+ T cells, after stimulation with PMA plus A23187, were able to produce higher levels of IFNgamma but lower levels of IL-4 and IL-13, by analysis at the single-cell level, as compared with the PB CD4+, CD45RO+ T cells. The majority of IL-4- or IL-13-producing ST CD4+ cells produced IFNgamma, although PB CD4+ T cells rarely showed this cytokine pattern. IL-10-producing CD4+ T cells were more frequently found in the ST than in the PB. Of interest, most of the IL-10-producing ST CD4+ T cells were able to produce IFNgamma. IL-2-producing CD4+ T cells were similarly present in both compartments. Similar intracellular cytokine patterns were observed with anti-CD3 plus anti-CD28 stimulation, although the number of detected cells was lower.. These data indicate that CD4+ T cells present within the inflamed synovium have apparently distinct cytokine profiles from those of memory CD4+ T cells in the PB, as typified by their ability to secrete both IFNgamma and IL-10.

Topics: Aged; Aged, 80 and over; Anti-Bacterial Agents; Antigens, Surface; Arthritis, Rheumatoid; Brefeldin A; Calcimycin; CD28 Antigens; CD3 Complex; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Separation; Cyclopentanes; Cytokines; Flow Cytometry; Humans; Immunophenotyping; Ionophores; Lymphocyte Activation; Macrolides; Middle Aged; Protein Synthesis Inhibitors; Synovial Membrane; T-Lymphocytes, Helper-Inducer

In the present study, we have demonstrated platelet-type 12-lipoxygenase (12-LOX) expression in human rheumatoid arthritis (RA) type B synoviocytes by reverse-transcription polymerase chain reaction (RT-PCR). The presence of 12-LOX mRNA in these cells was revealed by classical RT-PCR analysis using platelet-type 12-LOX cDNA primers and the PCR fragment (246 bp) was purified, amplified and sequenced. By sequence analysis, this fragment was determined to be 100% identical to that from platelet-type 12-LOX cDNA. Immunofluorescence data demonstrate that interleukin-1beta (IL-1beta) increases cellular 12-LOX protein. Other results associate specific inflammatory cytokines with the activity of 12-LOX in human RA type B synoviocytes. IL-1beta increased 12S-hydroxy-5,8,10,14-eicosatetraenoic acid (12-HETE) production (4-fold) and we also observed an increase in 12-HETE production (2.5-fold) after incubation of human RA type B synoviocytes with TNF alpha. In contrast to the action of IL-1beta on 12-HETE synthesis, IL-4 and IL-6 did not enhance 12-HETE production. This is the first demonstration of platelet-type 12-LOX cDNA derived from the mRNA of cultured human RA type B synoviocytes.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Arachidonate 12-Lipoxygenase; Arachidonic Acid; Arthritis, Rheumatoid; Blood Platelets; Calcimycin; Cells, Cultured; Cloning, Molecular; DNA Primers; Fluorescent Antibody Technique; Gene Expression; Humans; Interleukin-1; Interleukin-4; Interleukin-6; Microscopy, Fluorescence; Polymerase Chain Reaction; RNA, Messenger; Sequence Analysis, DNA; Synovial Membrane; Tumor Necrosis Factor-alpha

The conversion of endogenous arachidonic acid (AA) by polymorphonuclear cells (PMN) from patients with rheumatoid arthritis (RA) was studied before (D0) and one day (D1) after antiinflammatory drug therapy. The biosynthesis of 5,15-diHETE and lipoxins (LXS), were investigated ex vivo, after PMN stimulation by ionophore A23187 without exogenous addition of 15-HETE. The eicosanoids were resolved by RP-HPLC and simultaneously detected at 246 and 302 nm respectively. Large amounts of 5,15-diHETE (50 to 400 ng/10(7) PMN) and significant levels of LXS (from 2 to 20 ng/10(7) PMN) were produced with individual differences between donors. Metabolite levels varied between patients but this work showed for the first time a linear relationship between the amounts of 5,15-diHETE and LXS. Moreover LXS production after treatment may be related to long-term clinical improvement of patients.

Topics: Adult; Anti-Inflammatory Agents; Arachidonic Acid; Arthritis, Rheumatoid; Calcimycin; Chromatography, High Pressure Liquid; Eicosapentaenoic Acid; Female; Humans; Hydroxyeicosatetraenoic Acids; Lipoxins; Male; Middle Aged; Neutrophils; Spectrophotometry, Ultraviolet

PAF-acether (PAF) is a pro-inflammatory phospholipid molecule potentially involved in the pathogenesis of arthritis. PAF and related metabolites have been isolated in the synovial fluid from patients with arthritis. The aim of this study was to determine fluid and blood in patients with rheumatoid arthritis. Blood neutrophils from normal donors were also studied for their capacity to form PAF. Neutrophils were stimulated with the calcium ionophore A23187 (2 microM) for 1 to 60 min. PAF released in the medium and PAF associated to cells were measured. In synovial fluid neutrophils. PAF production began as soon as 1 min of stimulation (16.1 +/- 6.3 pmol per 1 x 10(6) cells) and reached a maximum at 20 min: 29.2 +/- 2.8 pmol per 1 x 10(6) cells (mean +/- SEM, n = 5). The amount of PAF released in the supernatant increased with the length of stimulation, similar amounts of PAF were produced by blood neutrophils isolated from the joint had a lower capacity to produce PAF than blood neutrophils from the same patients. The present results demonstrate the synthesis and release of PAF by synovial fluid neutrophils. They suggest that neutrophils may be source of PAF locally present in the joint. Newly synthesized PAF could participate in the amplification of the local inflammatory reaction.

Topics: Arthritis, Rheumatoid; Calcimycin; Humans; In Vitro Techniques; Ionophores; Neutrophils; Platelet Activating Factor; Platelet Aggregation; Synovial Fluid

5-Lipoxygenase products are pro-inflammatory mediators. Their roles and cellular origin in chronic inflammatory rheumatisms such as rheumatoid arthritis (RA) are poorly understood. The expression of arachidonate 5-lipoxygenase (5-LOX, arachidonate: oxygen 5-oxydoreductase; EC 1.13.11.34) and the 5-lipoxygenase activating protein (FLAP) genes in osteoarthritis and RA synoviocytes was studied at the transcriptional level by reverse transcription-polymerase chain reaction (RT-PCR) methodology. Arachidonic acid metabolism was analyzed by reverse-phase high pressure liquid chromatography. 5-LOX and FLAP mRNA were detectable using RT-PCR in all sources of synoviocytes tested. The expression of 5-LOX and FLAP mRNA led to the synthesis of 5-LOX metabolites. 12- and 15-LOX activities were also present. These LOX products can participate in inflammatory processes leading to joint destruction in RA.

Topics: 5-Lipoxygenase-Activating Proteins; Arachidonate 5-Lipoxygenase; Arachidonic Acid; Arthritis, Rheumatoid; Base Sequence; Calcimycin; Carrier Proteins; Cells, Cultured; Chromatography, High Pressure Liquid; DNA Primers; Electrophoresis, Agar Gel; Fluorescent Antibody Technique, Indirect; Humans; Hydroxyeicosatetraenoic Acids; Leukotrienes; Membrane Proteins; Molecular Sequence Data; Osteoarthritis; Polymerase Chain Reaction; RNA, Messenger; Synovial Membrane

Phorbol 12-myristate 13-acetate (100 nM), a potent protein kinase C and macrophage activator, has a biphasic affect on 25(OH)D3-1 alpha-hydroxylase activity in synovial fluid macrophages from arthritis patients. After 5 h, 1 alpha, 25(OH)D3 synthesis fell from 5.2 +/- 0.1 to 1.6 +/- 0.2 pmol/h per 10(6) cells, however, after 24 h and 48 h, synthesis increased to 17.4 +/- 0.3 and 22.3 +/- 1.4 pmol/h per 10(6) cells, respectively. Although an independent short-term mechanism is suggested, protein kinase C may promote macrophage activation, thus increasing long-term 25(OH)D3-1 alpha-hydroxylase expression. Intracellular calcium and cAMP are unlikely to activate the enzyme, since 0.1 microM of the calcium ionophore, A23187, and 1 mM dibutyryl-cAMP inhibited synthesis by 87% and 79%, respectively, after 24 h.

Topics: Arthritis; Arthritis, Gouty; Arthritis, Rheumatoid; Bucladesine; Calcimycin; Calcitriol; Cells, Cultured; Dose-Response Relationship, Drug; Humans; Macrophages; Synovial Fluid; Tetradecanoylphorbol Acetate

A phosphonate-containing phospholipid (PL) analogue (Compound 1) designed as a transition-state inhibitor competively inhibits non-human extracellular PLA2 at a mole fraction of 0.003 in the kinetic "scooting mode" (Jain et al., Biochem 28:4135 (1989]. To further profile the activity of Compound 1, we examined its activity with purified human enzyme and in whole cell systems. Compound 1 effectively inhibited a 14 kDa human PLA2 purified from joint synovial fluid of patients with rheumatoid arthritis using 3H-AA labeled E. coli as substrate (IC50 = 1.7 microM) and a high MW PLA2 (110 kDa) isolated from the cytosol of a human monocytic cell line, U-937, which selectively hydrolyzes AA-containing PL (IC50 = 165 microM). It failed to reduce A23187-induced PGE2 or LTC4 production by human adherent monocytes or LTB4 release from human neutrophils which may be due, in part, to poor membrane partitioning.

Topics: Arthritis, Rheumatoid; Calcimycin; Cell Line; Dinoprostone; Eicosanoids; Escherichia coli; Humans; In Vitro Techniques; Male; Monocytes; Neutrophils; Organophosphonates; Phospholipases A; Phospholipases A2; Phospholipids; SRS-A; Synovial Fluid

Retinoids have demonstrated antiinflammatory activity in certain animal models and human disease states. The mechanism by which retinoids elicit this activity is unknown. Some retinoids are known to inhibit arachidonic acid (AA) release and metabolism in intact cells in vitro. Retinoids may exert their antiinflammatory effects by inhibiting phospholipase A2 (PLA2) and the resultant production of inflammatory AA metabolites. Retinoids were evaluated in vitro as inhibitors of the PLA2 activity in human synovial fluid (HSF-PLA2). Of the naturally occurring, nonaromatic retinoids tested, all-trans-retinal, all-trans-retinoic acid (all-trans-RA) and 13-cis-RA were the most potent inhibitors (IC50 S 6-15 microM), whereas all-trans-retinol was much less potent. Of the synthetic aromatic retinoids and arotinoids examined, the free carboxylic, sulfonic, and sulfinic acid forms were more than 15-fold more potent inhibitors of HSF-PLA2 than their corresponding ethyl esters. These retinoids also were evaluated as inhibitors of calcium ionophore A23187-induced AA release from rat peritoneal macrophages. All-trans-RA and 13-cis-RA were potent inhibitors of AA release from these cells (IC50 S 4 microM), while the other natural retinoids were inactive. Of the aromatic retinoids and arotinoids tested, the free acid forms (IC50 S 2-6 microM) were 5- to 21-fold more potent inhibitors of AA release from the macrophages than their corresponding ethyl esters. The potencies of the arotinoids as inhibitors of HSF-PLA2 appeared to correlate with their potencies as inhibitors of AA release from A23187-stimulated rat peritoneal macrophages. These data support the hypothesis that one possible mechanism for the known antiinflammatory activity of some retinoids may be by inhibition of phospholipase A2.

Topics: Animals; Arachidonic Acid; Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Depression, Chemical; Humans; Macrophages; Peritoneal Cavity; Phospholipases A; Phospholipases A2; Rats; Retinoids; Structure-Activity Relationship; Synovial Fluid

We studied the effects of a single, oral dose of methotrexate (MTX) on arachidonic acid metabolism in neutrophils from 6 patients with rheumatoid arthritis, which were obtained 1 day before and 1 day after their usual weekly MTX dose. The 6 patients had received a mean weekly MTX dose of 9.6 mg (range 5-15) for a mean of 61.7 months (range 58-64), and none received concomitant corticosteroids. Total generation of leukotriene B4 (LTB4) in neutrophils stimulated ex vivo with 10 microM calcium ionophore A23187 for 20 minutes was significantly suppressed, by a mean of 53%, after the MTX dose compared with the predose levels (mean +/- SEM 13.0 +/- 1.4 ng/10(6) cells versus 6.0 +/- 0.9 ng/10(6) cells; P = 0.0019), reflecting a comparable suppression of both released and cell-retained LTB4. A 49% decrease in omega-oxidation products of LTB4 demonstrates that decreased LTB4 synthesis, rather than increased degradation, is responsible for the decrease in LTB4 generation. The absence of a significant change in either 3H-labeled arachidonic acid release or platelet-activating factor generation indicates that the observed decrease in LTB4 synthesis was apparently not caused by diminished phospholipase A2 activity. A 28% decrease in the total formation of the 5-lipoxygenase products 5-hydroxyeicosatetraenoic acid and the 6-trans-LTB4 diastereoisomers, and a 48% suppression of production of LTB4 plus its omega-oxidation metabolites after the MTX dose suggest inhibition of 5-lipoxygenase activity and possible suppression of leukotriene A4 epoxide hydrolase activity.

Topics: Aged; Arachidonic Acid; Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Chemotaxis, Leukocyte; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Methotrexate; Middle Aged; Neutrophils; Phospholipases; Platelet Activating Factor

Human synovium obtained at arthroplasty from patients with rheumatoid arthritis (RA) and osteoarthritis (OA) were characterized by assessing mast cell morphology, content and function. Histological studies confirmed significant numbers of mast cells in both RA and OA synovium. Electron microscopic data support the morphologic similarity between human synovial mast cells and human mast cells in lung and intestine. Likewise, synovial mast cells do not appear to be functionally different from pulmonary or intestinal mucosal mast cells. Mast cell suspensions with a cellular histamine content of 4.3 +/- 0.5 pg/cell (mean +/- SEM) released histamine following provocation with anti-IgE and calcium ionophore but not compound 48/80, f-met peptide or bradykinin. Prostaglandin D2 (PGD2) and leukotriene C4 (LTC4) were also released in response to anti-IgE. Auranofin inhibited anti-IgE provoked histamine, PGD2 and LTC4 release while gold sodium thiomalate, cromolyn and indomethacin had no effect on histamine release. Theophylline inhibited anti-IgE induced histamine release only at concentrations greater than or equal to 10(-3) M. Our study argues against functional or morphologic mast cell heterogeneity of human intestinal, lung and synovial origin and suggests that mast cells may have a pathogenic role in both RA and OA.

Topics: Antibodies, Anti-Idiotypic; Arthritis, Rheumatoid; Auranofin; Calcimycin; Histamine Release; Humans; Immunoglobulin E; In Vitro Techniques; Mast Cells; Osteoarthritis; Synovial Membrane

The formation of 5-lipoxygenase products (5-hydroxyeicosatetraenoic acid [5-HETE], leukotriene B4 [LTB4], and leukotriene C4 [LTC4]) by polymorphonuclear and mononuclear leukocytes isolated from peripheral blood of patients with rheumatoid arthritis was evaluated and compared with the data obtained from a group of control subjects. Although the levels of arachidonic acid metabolites via 5-lipoxygenase pathway by stimulated polymorphonuclear cells were comparable between patients and controls, mononuclear leukocytes from patients synthesized, when stimulated, significantly greater amounts of 5-HETE, LTB4, and LTC4 than did cells isolated from normal subjects. In addition, the release of superoxide anion, stimulated by either a particulate or a soluble stimulus, was increased in mononuclear cells from patients. The enhanced capacity of peripheral mononuclear leukocytes isolated from patients with rheumatoid arthritis to generate proinflammatory metabolites of arachidonic acid and oxygenated species with bactericidal and tissue-damaging properties may contribute to the pathogenesis of this complex disease.

Topics: Arthritis, Rheumatoid; Calcimycin; Cell Aggregation; Humans; Hydroxyeicosatetraenoic Acids; Leukocytes, Mononuclear; Leukotriene B4; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Phagocytosis; SRS-A; Superoxides

Peripheral blood lymphocytes from patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), progressive systemic sclerosis (PSS), Reiter's disease, osteoarthritis, and from healthy volunteers were investigated for interferon-gamma (IFN-gamma) production after mitogen activation. Phytohaemagglutinin stimulation revealed an impaired IFN-gamma production in RA, SLE, and PSS but normal levels in Reiter's disease and osteoarthritis. In RA this deficiency was also seen after pokeweed mitogen, OKT3, and concanavalin A activation. No major differences were found in interleukin 2 (IL-2) production and cell proliferation. The IL-2 receptor expression was reduced on stimulated RA lymphocytes. The deficient IFN-gamma production was compensated in RA by co-stimulation of PHA or OKT3 with phorbol myristic acetate (PMA). In addition, the combination of the calcium ionophore A 23187 and PMA induced a strong IFN-gamma secretion in all patient groups and in the controls.

Topics: Adolescent; Adult; Aged; Arthritis, Reactive; Arthritis, Rheumatoid; Autoimmune Diseases; Calcimycin; Drug Synergism; Humans; Interferon-gamma; Lectins; Lupus Erythematosus, Systemic; Lymphocyte Activation; Middle Aged; Osteoarthritis; Receptors, Immunologic; Receptors, Interleukin-2; Scleroderma, Systemic; Tetradecanoylphorbol Acetate

The calcium dependent metabolism of endogenous arachidonic acid (AA) was investigated in 17 patients with rheumatoid arthritis during treatment with dextropropoxyphene alone and in 25 healthy volunteers. Incorporation of [1-14C]AA into intracellular phospholipids of purified neutrophils was achieved by incubation until steady state before activation with ionophore A23187. Analysis of extracellular metabolites was performed by extraction, thin layer chromatography, autoradiography, and laser densitometry. The patients showed a twofold increase in the total capacity for oxidation of AA. Release of leucotriene B4 (LTB4) and its omega oxidation products, 20-OH LTB4 and 20-COOH LTB4, was 29%, range 11-48%, in patients compared with 8%, range 4-12%, in healthy volunteers. Total amounts of radioactivity released and the specific activity of LTB4, as assessed by high pressure liquid chromatography, were equal in experimental and control groups. The demonstrated increased capacity for metabolism of AA to the major proinflammatory metabolite, LTB4, via the 5-lipoxygenase pathway may contribute to perpetuation of inflammation and to tissue destruction in rheumatoid arthritis.

Topics: Adult; Aged; Arachidonic Acid; Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Chromatography, Thin Layer; Female; Humans; Leukotriene B4; Male; Middle Aged; Neutrophils

Microscopic analysis of synovial specimens from 35 patients with rheumatoid arthritis (RA) and 7 patients with osteoarthritis revealed mast cell hyperplasia in perivascular regions, in fibrous interstitial areas, and clustered around the periphery of lymphoid aggregates. Metachromatic staining, immunofluorescence studies, and ultrastructural analysis revealed a single population of connective tissue-type mast cells with surface IgE receptors. Total extractable histamine of synovial tissue was 4.15 +/- 2.30 micrograms/gm (n = 8) for RA synovium and 0.53 +/- 0.23 microgram/gm (n = 7) for OA synovium. Mast cell secretion was assessed and specific release of histamine from RA synovial mast cells was observed following stimulation with anti-IgE (32.3%), compound 48/80 (40.1%), calcium ionophore A23187 (25.2%), and a partially purified lymphokine with histamine-releasing activity (23.9%).

Topics: Antibodies, Anti-Idiotypic; Arthritis, Rheumatoid; Calcimycin; Fluorescent Antibody Technique; Histamine; Histamine Release; Humans; Immunoglobulin E; Lymphokines; Mast Cells; Microscopy, Electron; Osteoarthritis; p-Methoxy-N-methylphenethylamine; Synovial Membrane

Prostaglandins (PG) and related eicosanoids which derive from essential fatty acids are important mediators and modulators of inflammation. Macrophages (M phi), which derive from peripheral blood monocytes (PBM), are prominent cells in the synovium of patients with rheumatoid arthritis (RA), and are a major source of synovial PGE2. In addition, fresh and cultured PBM from RA patients produce more PG than normal control cells. When allowed to mature in culture PBM exhibit many characteristics of macrophages (M-M phi). We examined uptake by M-M phi of eicosanoid precursor fatty acids (FA), their incorporation into cellular phospholipid (PL), and mobilization of FA after cell stimulation. Cultured M-M phi from treated and untreated RA patients (RA M-M phi) took up significantly more linoleic acid (LA), dihomogammalinolenic acid (DHLA) and arachidonic acid (AA) than M-M phi from normal volunteers (N M-M phi). The enhanced uptake of FA observed in 12-day cultures of RA M-M phi was similar to uptake seen in normal human peritoneal macrophages (PM phi). After uptake FA were incorporated mainly into phosphatidylcholine (PC). M-M phi from untreated RA patients incorporated a smaller proportion of [14C]LA into PC (37.0 +/- 12.7% of total PL label) than normal cells (86.0 +/- 4.2%), and a greater proportion of [3H]AA into PC (57.1 +/- 7.1%) than normals (23.9 +/- 6.9%). Stimulation of M-M phi with calcium ionophore A23187 resulted in significantly greater hydrolysis of LA and AA from PC in RA M-M phi from both treated and untreated patients than from PC in N M-M phi. The data indicate that M-M phi from RA patients mature more rapidly in vitro than M-M phi from controls as uptake of FA by RA M-M phi increases with duration of culture and by 12 days in culture equals uptake by normal human peritoneal M phi. Also, RA M-M phi exhibit differences from N M-M phi in uptake, PL distribution, and hydrolysis of eicosanoid precursor FA. Such changes in FA metabolism might influence cell function and inflammatory responses.

Topics: 8,11,14-Eicosatrienoic Acid; alpha-Linolenic Acid; Arachidonic Acid; Arachidonic Acids; Arthritis, Rheumatoid; Calcimycin; Cell Differentiation; Cells, Cultured; Fatty Acids; Humans; Kinetics; Linoleic Acid; Linoleic Acids; Linolenic Acids; Macrophages; Membrane Lipids; Monocytes; Palmitic Acid; Palmitic Acids; Phospholipids; Prostaglandins

The production of three kinds of oxygen radicals (superoxide, hydrogen peroxide, and hydroxyl radicals) by neutrophils from patients with bacterial infection or rheumatoid arthritis was measured. The stimulators used in this study were opsonized zymosan (1 mg/ml), phorbol myristate acetate (20 ng/ml), A23187 (1 microM), and platelet activating factor (1 microM). Oxygen radical production by neutrophils from patients with rheumatoid arthritis was not significantly different from that of the control group. Hydrogen peroxide production by the neutrophils from patients with bacterial infection was significantly enhanced by only opsonized zymosan, but the production of the other kinds of oxygen radicals was not. Cytochalasin B reduced the production of hydrogen peroxide induced by opsonized zymosan more markedly than that of any other kind of oxygen radical. The measurement of hydrogen peroxide is suggested to be the most accurate indicator of the enhancement of intracellular production of oxygen radicals by neutrophils during infection.

Topics: Arthritis, Rheumatoid; Bacterial Infections; C-Reactive Protein; Calcimycin; Cytochalasin B; Free Radicals; Humans; Hydrogen Peroxide; Leukocyte Count; Neutrophils; Oxygen; Platelet Activating Factor; Superoxides; Tetradecanoylphorbol Acetate; Zymosan

Leukocyte suspensions containing basophils were obtained from 23 patients with rheumatoid arthritis (RA). When these cells were incubated with leukocyte nuclei from normal persons, histamine release was seen in 11 of the 15 patients with active disease, but not in the quiescent group or in normal individuals. The dose-response curve for histamine release was similar to that obtained by specific antigen in type I allergy. By removal from and refixation to the cells of surface Ig, the release of histamine was respectively, abolished and restored, just as in similar experiments in hay fever patients. The dependence of pH for removal was also identical with that found in type I allergy. Antinuclear antibodies of the IgE class (IgE ANA) mainly directed against the granulocyte nuclei were often found in serum and on the cell surface of the RA patients, but not in normal individuals. A correlation was found between these titres in serum and on the cell surface. No correlation was found between ANA in serum and on the cell surface, on the one hand and disease activity and histamine release on the other. In a group of 12 patients with another joint disease, osteoarthrosis, only two patients showed histamine release, and in contrast to the other patients they showed swelling of more than two joints. The present investigation supports our hypothesis of an involvement of an autoimmune type I reaction directed against nuclear components in the RA disease.

Topics: Adult; Aged; Animals; Antibodies, Anti-Idiotypic; Antibodies, Antinuclear; Antigens, Nuclear; Arthritis, Rheumatoid; Basophils; Calcimycin; Female; Histamine Release; Humans; Immunoglobulin E; Leukocytes; Male; Middle Aged; Nucleoproteins; Osteoarthritis; Rabbits; Receptors, Antigen, B-Cell

The mechanism of poor lymphocyte transformation to mitogens was studied in selected patients with rheumatoid arthritis and systemic lupus erythematosus. Low lymphocyte response to PHA and Con-A in media containing autologous and homologous sera was usually associated with poor response to the calcium ionophore A23187, which induces blastogenesis by a different mechanism. The low lymphocyte response to mitogens in patients with rheumatoid arthritis could be restored by in vivo treatment with the anthelmintic drug, levamisole. The present findings suggest that intrinsic defects are responsible for the decreased cellular response in patients with rheumatoid arthritis and systemic lupus erythematosus.

Topics: Adolescent; Adult; Arthritis, Rheumatoid; Calcimycin; Concanavalin A; Humans; Lectins; Levamisole; Lupus Erythematosus, Systemic; Lymphocyte Activation; Middle Aged