concanavalin-a and Arthritis--Rheumatoid

Topics: Acrylamides; Adolescent; Adult; Agammaglobulinemia; Age Factors; Antibody Formation; Arthritis, Rheumatoid; B-Lymphocytes; Concanavalin A; Female; Humans; Immunosuppressive Agents; Kidney Transplantation; Lymphocyte Activation; Male; Middle Aged; Nitrobenzenes; Receptors, Antigen, B-Cell; T-Lymphocytes; Trinitrobenzenes

Variable treatment responses to glucocorticoids occur in patients with rheumatoid arthritis (RA). In renal transplantation and asthma treatment responses correlate with in vitro tests of glucocorticoid immunosuppression. We compared in vitro methylprednisolone suppression of concanavalin A stimulated cellular proliferation with clinical responses to methylprednisolone in patients with RA. Patients found to be glucocorticoid sensitive by in vitro testing had significantly greater improvements in joint score and soluble interleukin 2 receptor (sIL-2R) levels compared to control patients, indicating that individual responsiveness to glucocorticoid exists in RA. Similar in vitro and in vivo changes of sIL-2R levels suggests that they reflect cell mediated immune events in vivo.

Topics: Arthritis, Rheumatoid; Cell Division; Concanavalin A; Glucocorticoids; Humans; Immunity, Cellular; Immunosuppression Therapy; Methylprednisolone; Monocytes; Pain Measurement; Prospective Studies; Receptors, Interleukin-2

Gold salts in vitro modulate lymphocyte proliferation to mitogens and antigens and macrophage phagocytosis. These effects are not confined to gold salts; D-penicillamine and chloroquine as well as some of the non-steroidal anti-inflammatory drugs (NSAIDs) have in vivo immunoregulatory effects. Peripheral blood mononuclear cells during treatment with Myocrisin (gold sodium thiomalate, GSTM) show changes that differ from in vitro effects and are related to therapeutic response rather than GSTM administration. This discrepancy between in vitro and ex vivo responses prompted us to measure cellular functions during auranofin therapy. Twenty-nine patients with rheumatoid arthritis took part in a placebo-controlled trial of auranofin. We examined the spontaneous immunoglobulin (IgG and IgM) and IgM rheumatoid factor (IgM RF) production by cultured mononuclear cells, lymphocyte transformation to concanavalin A and macrophage phagocytosis of Candida albicans. There was a significant fall in IgG synthesis (p less than 0.005) and IgM RF synthesis (p less than 0.005) over the first 4 months of treatment, whereas in the control group there were no significant changes. There was no significant change in IgM production. In the auranofin-treated group the lymphocyte response to concanavalin A fell progressively during 6 months of therapy (at 2 months p less than 0.05, at 4 months p less than 0.01, and at 6 months p less than 0.005). Auranofin therapy had variable effects on monocyte phagocytosis of C. albicans. Therefore, in contrast to GSTM, auranofin suppressed both in vitro and ex vivo lymphocyte functions. This effect is probably related to the direct effect of auranofin on lymphocyte membranes.

Topics: Anti-Inflammatory Agents; Arthritis, Rheumatoid; Auranofin; Aurothioglucose; Candida albicans; Clinical Trials as Topic; Concanavalin A; Double-Blind Method; Gold; Humans; Immunoglobulins; Lymphocytes; Macrophages; Phagocytosis

Lymphocyte proliferation is a major factor determining the magnitude of the immune response. Both dexamethasone (DEX) and tofacitinib (TOF) exert marked immunosuppressive effects and are mainstay drugs in the treatment of rheumatoid arthritis (RA). This study was aimed to explore the single and combined anti-proliferative action of DEX and TOF on lymphocytes and their sex differences.. The single-drug effects and dual-drug interactions of TOF and DEX were assessed on the in vitro concanavalin A-stimulated proliferation of lymphocytes isolated from male and female rat and human peripheral blood.. TOF has a promising steroid-sparing potential with the beneficial effects of the combination therapy more likely in males than females.

Topics: Animals; Arthritis, Rheumatoid; Cell Proliferation; Cells, Cultured; Concanavalin A; Dexamethasone; Drug Antagonism; Drug Synergism; Drug Therapy, Combination; Female; Humans; Inhibitory Concentration 50; Lymphocytes; Male; Piperidines; Primary Cell Culture; Pyrimidines; Rats; Sex Factors; Species Specificity

Parvovirus B19 (B19V) infection is associated with various autoimmune diseases. We investigated the levels of pro-inflammatory (IFNᵧ, TNFα, IL-2, IL-12) and anti-inflammatory (IL-4, IL-10) cytokines in the plasma of B19V DNA positive (B19. Blood samples were collected from 118 patients with RA and 49 healthy voluntaries. B19V sequence was determined in whole blood and cell-free plasma DNA by nested PCR. The levels of cytokines in the plasma and cell culture medium from Concanavalin A (ConA) or B19V VP1 protein stimulated PBMC were determined by ELISA.. The levels of IL-4, IL-10, IL-12, IL-2 and TNFα were higher in plasma of RA patients in comparison with control persons. B19. B19V infection could differently modulate the amount of cytokines in the plasma of healthy persons and RA patients. Decreased production of IFNᵧ and raised level of plasma IL-4 in RA patients could lower antiviral clearance.

Topics: Adult; Antibodies, Viral; Arthritis, Rheumatoid; Capsid Proteins; Concanavalin A; Cytokines; DNA, Viral; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunoglobulin G; Interferon-gamma; Interleukin-10; Interleukin-4; Leukocytes, Mononuclear; Male; Middle Aged; Parvoviridae Infections; Parvovirus B19, Human; Polymerase Chain Reaction

Previous studies suggest that selective serotonin reuptake inhibitors (SSRIs) modulate immune system functionality. SSRIs are the preferred treatment for major depressive disorder (MDD). A high rate of MDD is observed in rheumatoid arthritis (RA) patients. The aim of this study was to evaluate immunological effects of SSRIs in a rat model of RA.. Adjuvant arthritis was induced in 8-week-old Lewis rats; in the first set of experiments following the induction, 15.3 or 30.6 mg/kg of sertraline was daily injected into the ankle joint of the left rear leg. Clinical disease activity was evaluated and the findings compared with the 3 untreated legs and with control groups given methotrexate (MTX) or vehicle only at the same site. In a second set of experiments, the effect of 5, 25 and 50 mg/kg daily oral sertraline was evaluated in the same rat model. Splenocyte viability and inflammatory mediators were evaluated.. The sertraline-treated rats showed a significant reduction in clinical arthritis compared to controls, at all doses given, accompanied by a significant increase in interleukin 10 and a decrease in tumor necrosis factor-α levels and cycloxygenase-2 production, without lymphotoxicity. There was no significant difference from MTX, the first-line treatment for RA patients. Oral sertraline had a significant anti-inflammatory effect at all doses. There was no treatment × time effect.. The beneficial effects of sertraline in this rat model of arthritis have clinical implications for its use in humans. Large-scale clinical efficacy trials are needed.

Topics: Analysis of Variance; Animals; Arthritis, Rheumatoid; Concanavalin A; Cyclooxygenase 2; Cytokines; Cytotoxicity Tests, Immunologic; Disability Evaluation; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Freund's Adjuvant; Immunologic Factors; Methotrexate; Rats; Rats, Inbred Lew; Sertraline; Spleen; Thymidine

Altered glycosylation patterns in plasma proteins are found to be associated with the pathogenesis of various malignancies and autoimmune disorders. Our previous studies demonstrated the occurrence of some differentially glycosylated plasma proteins in rheumatoid arthritis (RA) patients. The current study was conducted to evaluate the alterations in expression and glycosylation of major acute phase proteins from wheat germ agglutinin enriched RA patients' plasma. Immunoblotting studies revealed a significant enhancement in the plasma levels of alpha-1 acid glycoprotein (AGP) and haptoglobin (Hp) in RA patients with respect to healthy controls. Monosaccharide analysis by high performance anion exchange-chromatography with pulse amperometric detection showed significant variations in the relative percentage of galactose, glucosamine and mannose in AGP and of mannose in Hp in RA patients. Altered patterns of mannosylation in AGP and Hp were also established by enzyme linked immunosorbent assay and Western blotting using Concanavalin-A lectin. These results could give information for understanding the disease pathogenesis and may provide an insight into the development and progression of the disease.

Topics: Adult; Arthritis, Rheumatoid; Case-Control Studies; Chromatography, Ion Exchange; Concanavalin A; Enzyme-Linked Immunosorbent Assay; Female; Galactose; Glucosamine; Glycosylation; Haptoglobins; Humans; Male; Mannose; Orosomucoid; Reproducibility of Results; Statistics, Nonparametric; Wheat Germ Agglutinins

This study was undertaken to identify the intracellular signaling pathway involved in induction of macrophage migration inhibitory factor (MIF) in human rheumatoid arthritis (RA) synovial fibroblasts.. Human RA synovial fibroblasts were treated with concanavalin A (ConA), various cytokines, and inhibitors of signal transduction molecules. The production of MIF by synovial fibroblasts was measured in culture supernatants by ELISA. The expression of MIF mRNA was determined using reverse transcriptase polymerase chain reaction (RT-PCR) and real-time PCR. Phosphorylation of p38 mitogen-activated protein (MAP) kinase in synovial fibroblasts was confirmed using Western blotting. The expression of MIF and p38 MAP kinase in RA synovium was determined using dual immunohistochemistry.. The production of MIF by RA synovial fibroblasts increased in a dose-dependent manner after ConA stimulation. MIF was also induced by interferon-γ, CD40 ligand, interleukin-15, interleukin-1β, tumor necrosis factor-α, and transforming growth factor-β. The production of MIF by RA synovial fibroblasts was significantly reduced after inhibition of p38 MAP kinase. The expression of MIF and p38 MAP kinase was upregulated in the RA synovium compared with the osteoarthritis synovium.. These results suggest that MIF production was induced through a p38 MAP-kinase-dependent pathway in RA synovial fibroblasts.

Topics: Arthritis, Rheumatoid; Base Sequence; Cells, Cultured; Concanavalin A; Cytokines; DNA Primers; Fibroblasts; Humans; Macrophage Migration-Inhibitory Factors; p38 Mitogen-Activated Protein Kinases; RNA, Messenger; Signal Transduction; Synovial Membrane

A proportion of patients with rheumatoid arthritis (RA) fail to respond adequately to corticosteroid (CS) therapy. Using an in vitro CS sensitivity bioassay, we have subdivided RA patients into steroid-sensitive (SS) and -resistant (SR) subgroups and this correlates with clinical responses to CS therapy. CSs exert their effects via the CS receptor (CR), which exists as two main isoforms, CRalpha and CRbeta. CRbeta can function as a negative inhibitor of CRalpha. We have hypothesized that steroid resistance in RA patients is due in part to a relative over-expression of the CRbeta.. Peripheral blood mononuclear cells (PBMCs) were isolated from SS and SR RA patients. CRalpha and CRbeta mRNA expression was determined by quantitative real time polymerase chain reaction (qRT-PCR). The ratio of CRbeta/CRalpha mRNA expression was determined. CRalpha and CRbeta protein expression by PBMCs was analysed by flow cytometry.. qRT-PCR analysis showed a trend towards higher expression of both CRbeta and basal CRbeta/CRalpha ratio in SR RA patients. Stimulation of PBMCs in vitro with concanavalin-A induced a significantly higher CRbeta mRNA expression, and CRbeta/CRalpha ratio in SR RA patients compared with SS patients, which was not inhibited by hydrocortisone. Flow cytometry showed that the percentage of PBMCs staining for CRbeta protein was significantly lower in the SS RA group (SS 43.3 +/- 14.8% vs SR 88.6 +/- 8.6%; P < 0.0010). The mean intensity of fluorescence CRbeta staining was higher in the SR RA patients (P < 0.001).. We show for the first time that CRbeta is over-expressed in SR RA patients and that hydrocortisone fails to inhibit concanavalin-A stimulated increase in CRbeta mRNA in SR RA patients. This mechanism may contribute in part to the CS hyporesponsiveness seen in some RA patients.

Topics: Adrenal Cortex Hormones; Adult; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Cell Proliferation; Cells, Cultured; Concanavalin A; Dose-Response Relationship, Drug; Drug Resistance; Female; Gene Expression Regulation; Humans; Hydrocortisone; Leukocytes, Mononuclear; Male; Middle Aged; Protein Isoforms; Receptors, Steroid; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

In this study, three single nucleotide polymorphisms (SNPs) located within the promoter of the human interleukin (IL)-10 gene [rs1800896 (position: -1087G>A), rs1800871 (position: -824C>T) and rs1800872 (position: -597C>A)] were investigated in 84 rheumatoid arthritis (RA) patients and 95 age- and sex-matched healthy subjects using polymerase chain reaction-restriction fragment length polymorphism method. Production of IL-10 by peripheral blood lymphocytes from the RA patients and healthy subjects cultured in the presence of Concanavalin A (Con A) was determined by using enzyme-linked immunosorbent assay. The results show that the distribution of the IL-10 genotypes did not differ significantly between RA patients and healthy subjects (P>0.05). However, a significant difference was observed in allele frequencies of -824CT, -824TT, -597CA, and -597AA between the RA patients and healthy volunteers (P=0.04). The -1087A/-824T/-597A (ATA) haplotype, which comprises all mutant alleles, was associated with lower IL-10 production when compared with the other haplotypes. In contrast, the RA patients who did not display the ATA haplotype produced significantly higher levels of IL-10 when compared with those carrying either one (P=0.012) or two (P=0.005) ATA haplotypes. Our findings suggest that there is an association between SNPs in the promoter of the human IL-10 gene and susceptibility to RA.

Topics: Alleles; Arthritis, Rheumatoid; Case-Control Studies; Concanavalin A; Enzyme-Linked Immunosorbent Assay; Female; Gene Frequency; Genetic Predisposition to Disease; Haplotypes; Humans; Interleukin-10; Malaysia; Male; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; Sequence Analysis, DNA

Altered glycosylation of plasma proteins has been directly implicated in the pathogenesis of rheumatoid arthritis (RA). The present study investigated the changes in the Concanavalin-A (Con-A)-bound plasma proteins in the RA patients in comparison to that of the healthy controls. Two proteins (MW approximately 32 kDa and approximately 62 kDa) showed an alteration in expression while an altered monosaccharide profile (high mannose) was observed in the approximately 62 kDa protein in the samples collected from RA patients. The 2-dimensional polyacrylamide gel electrophoresis analysis of the Con-A-bound plasma samples showed a large number of protein spots, a few of which were differentially expressed in the RA patients. Some unidentified proteins were detected in the RA patients which were absent in the control samples. The present study, therefore, enunciates the role of carbohydrates as well as that of the acute phase response in the disease pathogenesis.

Topics: Adult; Arthritis, Rheumatoid; Blood Proteins; Case-Control Studies; Chromatography, Agarose; Concanavalin A; Electrophoresis, Gel, Two-Dimensional; Glycosylation; Humans; Lectins; Middle Aged; Monosaccharides; Synovial Fluid

We evaluated the pattern of dexamethasone mediated inhibition of concanavalin-A (Con-A) stimulated peripheral blood mononuclear cell (PBMC) proliferation to classify patients with rheumatoid arthritis (RA) as corticosteroid resistant (CR) or sensitive (CS). We also studied the role of T helper 1, (Th1) and Th2 cytokines in the mechanism of glucocorticoid resistance in RA.. PBMC from 21 healthy controls and 15 patients with RA were isolated and cultured for the in vitro glucocorticoid sensitivity assay. Basal and Con-A stimulated PBMC proliferation levels and the inhibitory effect of different doses (10(-8), 10(-6), 10(-4) M) of dexamethasone (Dex) were evaluated. The IC50 was defined as the concentration of Dex that caused 50% inhibition of cell proliferation and subjects with an IC50 > 10(-6) M were considered to be CR. The supernatants were collected for cytokine [interleukin 4 (IL-4), IL-6, IL-10, tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma)] measurement by ELISA.. We observed lymphocyte proliferation after Con-A stimulation, which was inhibited by Dex in a dose-dependent manner in both groups. Two of 21 controls (9.5%) and 7/15 RA patients (53.3%) were CR (p < 0.01). Basal IL-4, IL-6, IL-10, and TNF-alpha levels were similar for both groups; however, basal IFN-gamma levels were slightly higher in patients with RA compared to controls. Con-A stimulation did not increase IL-4 or IL-6 levels compared to basal production but significantly increased IL-10 levels. IL-6 and IL-10 levels were significantly inhibited by Dex 10 M in both the control and RA groups. Con-A stimulation significantly increased TNF-alpha and IFN-gamma levels compared to the basal condition in the control and RA groups, and both cytokines were inhibited only by higher doses of Dex in the RA group.. These findings might reflect a predominance of Th1 cells in RA that might contribute to corticosteroid resistance in patients in RA.

Topics: Adult; Aged; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Biological Assay; Cell Division; Cells, Cultured; Concanavalin A; Cytokines; Dexamethasone; Dose-Response Relationship, Drug; Drug Resistance; Female; Humans; Interferon-gamma; Interleukin-10; Interleukin-6; Lymphocytes; Male; Middle Aged; Tumor Necrosis Factor-alpha

Alpha-1-acid glycoprotein (AGP) is a plasma glycoprotein produced by the liver that undergoes increased production and altered glycosylation in several physiological and pathological conditions including rheumatoid arthritis. To date, although present in the synovial fluid of rheumatoid arthritis patients, there has been no evidence for the separate extra-hepatic production of AGP. This study indicates that there could be a localized production of AGP in rheumatoid synovial fluid by demonstrating that the glycosylation patterns of AGP differed between the serum and synovial fluid in the same rheumatoid patient. Serum AGP was largely composed of fucosylated tri- and tetra-antennary oligosaccharide chains while the synovial fluid contained mainly bi-antennary chains that were fucosylated to a lesser extent. This structural heterogeneity of glycosylation resulted in functional diversity; serum but not synovial AGP is able to inhibit binding to the cell adhesion molecule E-selectin through expression of antigen sialyl Lewis X.

Topics: Arthritis, Rheumatoid; Carbohydrate Sequence; Chromatography, Affinity; Concanavalin A; Glycosylation; Humans; Hydrogen-Ion Concentration; Molecular Sequence Data; Orosomucoid; Synovial Fluid

In monolayer cultures human rheumatoid synovial fibroblasts (HRSF) secrete gelatinase A (MMP-2) and, unlike other human fibroblasts, to a minor extent also gelatinase B (MMP-9) as inactive proenzymes. In this regard HRSF resemble the fibrosarcoma cell line HT-1080. Unlike HT-1080, however, HRSF do not increase the secretion of MMP-9 in response to phorbol-12-myristate-13-acetate. This indicates that in HRSF the protein kinase C pathway for an enhanced MMP-9 secretion is inactive. None of the substances used in our study increased MMP-9 secretion, but some of them inhibited MMP-9 secretion. The secretion of MMP-2 could not be enhanced either, not even by dbcAMP, which has been reported to be effective in Sertoli and peritubular cells. Activation of MMP-2 in HRSF could be induced by treatment with concanavalin A (ConA) or cytochalasin D, as was shown for other cell types. This activation was not accompanied by a significant change in the amount of secreted TIMP-1 and TIMP-2. In contrast to reports on human skin fibroblasts, however, the activation of MMP-2 could not be induced in HRSF by treatment of the cells with monensin or sodium orthovanadate. Moreover, monensin was shown to act as an inhibitor of ConA- or cytochalasin D-mediated activation. Additionally, and in contrast to a report on a rat fibroblast cell line, MMP-2 activation is not mediated via the MAP kinase pathway in HRSF: PD 98059, a specific inhibitor of MAP kinase kinase, did not inhibit the activation of MMP-2. Similarly ineffective were PD 169316, an inhibitor for p38 MAP kinase, other inhibitors for protein kinases as lavendustin A, Gö 6983, wortmannin, rapamycin, as well as the protein tyrosine kinase inhibitors herbimycin A and genistein. Only staurosporin, a broad spectrum inhibitor of protein kinases, and the ionophores monensin and A 23187 effectively inhibited MMP-2 activation in HRSF. Our results demonstrate that MMP-2 can be activated by quite different pathways, and that different cells, even when belonging to the fibroblast family, do not necessarily use the same activating pathways.

Topics: Arthritis, Rheumatoid; Concanavalin A; Cyclic CMP; Cytochalasin D; Enzyme Activation; Enzyme Inhibitors; Fibroblasts; Gelatinases; Humans; Indomethacin; Ionophores; Matrix Metalloproteinase 2; Pentoxifylline; Protein Kinase Inhibitors; Synovial Fluid; Tetradecanoylphorbol Acetate; Tissue Inhibitor of Metalloproteinase-1; Tissue Inhibitor of Metalloproteinase-2; Vanadates

The enzyme ecto 5'-nucleotidase (5'N) was found to be active on 8/14 strains of Mycoplasma fermentans, K(m) (+/-S.D.) 3.8+/-2.8 microM 5'-AMP, and on the type strain of Mycoplasma pulmonis, K(m) 0.63 microM 5'-AMP. The six M. fermentans strains lacking 5'N activity were related by restriction fragment length polymorphism typing. At pH 8.5, the type strains of Mycoplasma arthritidis, Mycoplasma buccale and Ureaplasma urealyticum showed a relatively non-specific phosphatase activity against 5'-AMP but no activity was shown by the type strains of Mycoplasma genitalium, Mycoplasma hominis, Mycoplasma orale, Mycoplasma penetrans, Mycoplasma pneumoniae and Mycoplasma salivarium at this pH. M. fermentans has been reported from rheumatoid joints, which show a raised 5'N activity on their synovial cells and in their fluid which may be associated directly or indirectly with the mycoplasma.

Topics: 5'-Nucleotidase; Animals; Arthritis, Rheumatoid; Colony Count, Microbial; Concanavalin A; Humans; Mycoplasma; Mycoplasma Infections; Nucleic Acid Hybridization; Rats; RNA, Bacterial; Synovial Membrane

Osteoblastic stromal cells are capable of supporting osteoclast formation from hematopoietic precursors in the presence of osteotropic factors such as 1alpha,25(OH)(2)D(3), PTH, and IL-11. Osteoblastic stromal cells produce receptor activator of NF-kappaB ligand (RANKL), a type II membrane protein of the TNF ligand family, in response to these agents. Activated T lymphocytes also produce RANKL; however, the ability of this cell type to support osteoclast formation in vitro is unknown. Human PBMC-derived T cells, extracted using alphaCD3-coated magnetic beads, were cocultured with adherent murine spleen cells in the presence of Con A and a panel of cytokines. In the presence of Con A, bona fide osteoclasts were formed in vitro with activated T cells: IL-1alpha and TGFbeta further enhanced osteoclast numbers. PBMC-derived lymphocytes showed an increase in the mRNA expression of RANKL within 24 h of treatment with the same agents that were used to induce osteoclast formation. In synovial tissue sections with lymphoid infiltrates from RA patients, the expression of RANKL was demonstrated in CD3(+) T cells. The ability of activated T lymphocytes to support osteoclast formation may provide a mechanism for the potentiation of osteoclast formation and bone resorption in disease states such as rheumatoid arthritis.

Topics: Aged; Animals; Animals, Newborn; Arthritis, Rheumatoid; Carrier Proteins; Cell Differentiation; Coculture Techniques; Concanavalin A; Female; Gene Expression Regulation; Hematopoietic Stem Cells; Humans; Interleukin-2; Lymphocyte Activation; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Middle Aged; Osteoclasts; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; RNA, Messenger; Stromal Cells; Synovial Membrane; T-Lymphocytes; Transcription, Genetic; Transforming Growth Factor beta

Acute and chronic inflammation-induced expression of sialyl LewisX has already been shown to occur on alpha1-acid glycoprotein. We now demonstrate that this phenomenon is not restricted to alpha1-acid glycoprotein but also occurs on two other acute-phase proteins. ie on alpha1-antichymotrypsin and on haptoglobin. The level of expression of sialyl LewisX on these proteins was lower than on alpha1-acid glycoprotein, in all likelihood because alpha1-acid glycoprotein is the only acute-phase protein containing tetraantennary glycans. No expression of sialyl LewisX was detectable on alpha1-protease inhibitor, a protein with a high diantennary glycan content. Non-sialylated LewisX was not detectable on these major acute-phase proteins in any of the conditions studied. This indicates that the majority of the a3-linked fucose residues are present as sialyl LewisX on alpha1-acid glycoprotein, alpha1-antichymotrypsin and haptoglobin. The absolute contribution to the total phenotype in plasma of protein containing this determinant in a multivalent form was highest for alpha1-acid glycoprotein. This leads us to propose that alpha1-acid glycoprotein is, among the acute-phase proteins studied, the one with the highest potential for interference with the extravasation of leukocytes by binding to the selectins.

Topics: Acute-Phase Proteins; alpha 1-Antichymotrypsin; alpha 1-Antitrypsin; Antibodies, Monoclonal; Arthritis, Rheumatoid; Blotting, Western; Concanavalin A; Electrophoresis; Haptoglobins; Humans; Inflammation; Lectins; Liver; Oligosaccharides; Orosomucoid; Sialyl Lewis X Antigen; Wounds and Injuries

To determine the proportion of glycosylated ferritin [ferritin bound to concanavalin A (Con-A)] and ferritin subunits in sera and synovial fluids (SF) from patients with rheumatoid arthritis (RA).. Ferritin concentrations were measured by a sandwich ELISA using rabbit IgG F(ab')2 anti-human ferritin antibody as a coating antibody. Proportions of glycosylated ferritin were examined using Con-A Sepharose 4B. Ferritin subunits were tested by Western blot analysis.. Ferritin concentrations in RA SF were significantly elevated compared to those in osteoarthritis (OA) SF (p < 0.01) and those in RA sera (p < 0.01). Percentages of glycosylated ferritin in SF were low in both RA and OA (RA 11.9 +/- 10.7, n = 41; OA 6.9 +/- 11.0, n = 10). However, percentages of glycosylated ferritin in RA sera (65.9 +/- 15.0, n = 20) were significantly higher than in RA SF (p < 0.01). Western blot analysis revealed both G subunit (23 kDa) and L subunit ( 19 kDa) in RA sera, although SF ferritin was composed mostly of L subunit.. Significant differences in ferritin molecule composition were observed between sera and SF from patients with RA, which suggests that in RA most SF ferritin is synthesized locally in the affected joint.

Topics: Adult; Aged; Aged, 80 and over; Arthritis, Rheumatoid; Blotting, Western; Concanavalin A; Enzyme-Linked Immunosorbent Assay; Female; Ferritins; Humans; Male; Middle Aged; Protein Binding; Rheumatoid Factor; Synovial Fluid

Galactose-free immunoglobulin G (IgG), which is known to be higher in the sera of patients with rheumatoid arthritis, was prepared from IgG of healthy volunteers using enzymes. Its reactivity to lectins was analyzed. The galactose-free IgG showed no reactivity to Ricinus communis agglutinin 120 but displayed greater reactivity to concanavalin A and Lens culinaris lectin than did intact human IgG. Then, IgG in serum samples was bound to protein A immobilized on a nitrocellulose membrane, and its reactivity to biotinylated concanavalin A was measured with streptavidin-conjugated horseradish peroxidase. When the reactivity to concanavalin A of IgG in sera from healthy individuals and patients with rheumatoid arthritis (RA), osteoarthritis, systemic lupus erythematosus, or hepatic disease was compared, higher levels were shown in patients with RA, notably in 60% of the seronegative patients and 80% of the early phase patients. Therefore, it was suggested that augmentation of the abnormal IgG in sera was highly specific to patients with RA and that this novel serum test could be very useful for an accurate diagnosis of this disease.

Topics: Arthritis, Rheumatoid; Biotinylation; Carbohydrate Sequence; Concanavalin A; Galactose; Humans; Immunoglobulin G; Lectins; Molecular Sequence Data; Oligosaccharides; Plant Lectins; Protein Binding; Staphylococcal Protein A

alpha 1-Acid glycoprotein (AGP) exists as an heterogeneous population of glycosylated variants (glycoforms) in plasma. The concentration of AGP increases some 2-5 fold in certain pathophysiological states exemplified by the chronic inflammatory disease, rheumatoid arthritis (RA). Moreover, the expressed glycosylation pattern alters in such conditions, indicating functional significance that is likely to be related to the oligosaccharide heterogeneity. We have investigated the heterogeneity of AGP glycosylation using the technique of high pH anion-exchange chromatography (HPAEC). AGP was isolated from the blood of RA sufferers, partially separated by Concanavalin A (Con A) affinity chromatography into bound and non-bound fractions and was enzymatically deglycosylated. Chromatography on the pellicular HPAE resin at pH 13 separated the released oligosaccharides and allowed a comparison of profiles in terms of branching and fucosylation. Results demonstrate an abnormal RA AGP glycosylation, with a tendency towards tri- and tetra-antennary oligosaccharides and enhanced fucosylation, in addition to the possible existence of penta-sialylated RA AGP glycoforms.

Topics: Arthritis, Rheumatoid; Chromatography, Affinity; Chromatography, Ion Exchange; Concanavalin A; Humans; Monosaccharides; Oligosaccharides; Orosomucoid; Synovial Fluid

High-pH anion-exchange chromatography with pulsed amperometric detection is a highly sensitive technique that can be used for detecting changes in sialylation and fucosylation, as well as different branching patterns of N-linked oligosaccharides in glycoproteins. We examined the N-glycans of alpha1-acid glycoprotein obtained from twelve patients with various inflammatory conditions with this technique, as well as traditional concanavalin A crossed affinity immunoelectrophoresis. We found the chromatographic profiles of N-glycans in all patients with rheumatoid arthritis to be very similar, but significantly different from normal controls. N-glycans from patients with ulcerative colitis also showed specific alterations in their chromatographic profiles. However, some heterogeneity was found between these patients, perhaps reflecting changes in glycosylation secondary to certain states of the disease, or to medical treatment. We conclude that this technique is useful for detailed mapping of glycosylation changes in alpha1-acid glycoprotein in clinical samples, and that it may be used to further increase our knowledge about glycosylation changes in response to inflammatory disease.

Topics: Acute Disease; Adult; Aged; Arthritis, Rheumatoid; Cholangitis; Chromatography, Ion Exchange; Colitis, Ulcerative; Concanavalin A; Female; Glycosylation; Humans; Hydrogen-Ion Concentration; Immunoelectrophoresis, Two-Dimensional; Male; Middle Aged; N-Acetylneuraminic Acid; Orosomucoid; Polysaccharides; Radiochemistry

Topics: Adult; Aged; Arthritis, Rheumatoid; Carcinoma, Hepatocellular; Cell Line; Chromatography, Affinity; Concanavalin A; Female; Humans; Liver Neoplasms; Male; Orosomucoid; Protein Binding; Sepharose; Tumor Cells, Cultured

We developed an assay for the direct determination of selenium in serum with a Perkin-Elmer Model 4100ZL Zeeman atomic absorption spectrometer and Ag-Cu-Mg modifier. We used this assay to analyze concanavalin A-bound selenoprotein (CABSP) in human serum after concanavalin A (ConA) affinity chromatography. The CABSP was identified as a single-chain glycoprotein of 57.3 kDa. Carbohydrate units were N- and O-linked to the protein. The selenium moiety was selenocysteine. Total selenium, glutathione peroxidase (GPX; EC 1.11.1.9), ConA-bound selenium (CABS), and alpha 1-acid glycoprotein (AAG) were determined in normal subjects and patients with various pathological conditions. CABS accounted for 44.1% +/- 6.3% of total selenium in sera from normal subjects and 46.5% +/- 3.9% to 55.1% +/- 8.1% in sera from patients with a variety of diseases. Total selenium in serum was well correlated with serum CABS (r = 0.860), but not with serum GPX activity (r = 0.117), for all patients studied. Serum CABS increased in normal subjects after selenium supplementation. Serum CABSP did not behave as an acute-phase reactant, compared with AAG.

Topics: Adult; Arthritis, Rheumatoid; Chromatography, Affinity; Chromatography, Gel; Concanavalin A; Diabetes Mellitus; Glutathione Peroxidase; Glycosylation; Humans; Kidney Diseases; Middle Aged; Neoplasms; Orosomucoid; Proteins; Reference Values; Selenium; Selenoproteins; Spectrophotometry, Atomic

This study focuses on the consequences of T-lymphocyte activation in chronic polyarthritis in terms of expression of cell surface receptors interacting with extracellular matrix (ECM). The expression of the VLA group of integrins was studied on in vitro-stimulated peripheral-blood T cells, and on peripheral-blood and synovial-fluid mononuclear cells (MNC) of patients with polyarthritis. The VLA expression was measured by flow cytometry using monoclonal antibodies (MoAbs) against alpha-subunits of the VLA family. VLA-alpha 4 and VLA-alpha 5, but not VLA-alpha 1, were expressed on a major fraction of unstimulated peripheral-blood T cells both in the patients with polyarthritis and in healthy individuals. Two distinct populations, VLA-alpha 5-high and VLA-alpha 5-low, were found in resting peripheral-blood T lymphocytes. Two days after stimulation by phorbol 12-myristate 13-acetate (PMA) and concanavalin A, most T cells became VLA-alpha 5-high. In patients with chronic polyarthritis, the expression of VLA-alpha 1 and VLA-alpha 5 was always higher on synovial-fluid T cells than on peripheral-blood T cells. These results give further support to the hypothesis that upon activation the induction of the VLA adhesion-molecule expression may be a factor contributing to the accumulation of T cells in the inflamed synovium.

Topics: Adult; Aged; Arthritis, Reactive; Arthritis, Rheumatoid; Concanavalin A; HLA-DR Antigens; Humans; Lymphocyte Activation; Middle Aged; Receptors, Fibronectin; Synovial Fluid; T-Lymphocytes; Tetradecanoylphorbol Acetate

To determine whether factors related to disease activity determine changes in the glycosylation of alpha 1-acid glycoprotein (AGP) observed at the early stage of rheumatoid arthritis (RA).. Using affinoimmunoelectrophoresis with the lectin concanavalin A (Con-A), the microheterogeneity of serum AGP was studied in patients with early (n = 54) and established (n = 46) RA and the results were expressed as reactivity coefficients (AGP-RC).. When compared with controls (n = 44), AGP-RC values were increased in the patients with very recent RA of no more than 3 months duration (p < 0.05), whereas normal or low AGP-Con-A reactivity was found in the patients with early RA with disease duration exceeding 3 months. In the entire group with early RA, the multiplicative model of regression described the relationship between AGP-Con-A reactivity and disease duration (p < 0.005). AGP variant with low binding affinity to Con-A predominated in the sera of patients with established RA and no relationship between the glycosylation profile of AGP and disease duration was observed in this group. When AGP-RC values were compared in the subgroups of patients with early and established RA with similar disease activity as measured by the Mallya-Mace score or erythrocyte sedimentation rate, there was a significant decrease in AGP-RC with increasing disease activity (p < 0.05).. In view of our findings, early RA begins as an acute inflammation with increase of AGP-Con-A reactivity and becomes chronic during the first year of the disease. Factors related to disease activity appear important in determining the rate at which RA enters a chronic phase.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Arthritis, Rheumatoid; Concanavalin A; Cytokines; Female; Glycosylation; Humans; Immunoelectrophoresis; Male; Middle Aged; Orosomucoid; Regression Analysis; Severity of Illness Index; Time Factors

Using different approaches evidence is provided that aminopeptidase N (APN, EC 3.4.11.2, CD13) is expressed on the surface of resting and stimulated human lymphocytes. 1) 50% of total Ala-pNA hydrolysis of viable cells was found to be due to the ectoenzyme APN as revealed by titration with the new inhibitor probestin in comparison to purified APN from human kidney (Ki value 6.5 nM). After stimulation with ConA an increase of Ala-pNA hydrolysis was observed from 8 pkat up to 15 pkat/10(6) cells. 2) This correlated with an increase of APN (CD13) surface expression as detected by antibody surface labeling of unstimulated (5-12% CD13 positive cells) and stimulated lymphocytes (28-36% CD13 positive lymphocytes) using polyclonal and monoclonal antibodies. 3) In vivo APN was found to be expressed on the surface of up to 50% lymphocytes isolated from the synovial fluid of rheumatoid patients.

Topics: Aminopeptidases; Antibodies, Monoclonal; Arthritis, Rheumatoid; CD13 Antigens; Cells, Cultured; Concanavalin A; Humans; Hydrolysis; Leukocytes, Mononuclear; Lymphocyte Activation; Lymphocytes; Monocytes; Oligopeptides; Synovial Fluid; T-Lymphocytes

We investigated serum levels of interleukin-6 (IL-6), interferon-gamma (IFN-gamma), and tumor necrosis factor alpha (TNF alpha) from patients with systemic lupus erythematosus (SLE) and its various clinical manifestations of disease and from patients with rheumatoid arthritis (RA) and other rheumatic diseases. The serum levels of IL-6 and IFN-gamma were highly elevated from patients with SLE associated with lymphadenopathy (LN) or nephrotic syndrome (NS). On the contrary, the serum levels of TNF alpha were elevated from most patients with SLE associated with thrombocytopenia (TP). However, serum levels of TNF alpha were in the normal range from patients with SLE associated with NS, LN, or central nervous system disease. Of interest, patients with SLE associated with humoral immunodeficiency disorder, hypogammaglobulinemia, had highly elevated levels of serum IL-6. The concanavalin A-stimulated mononuclear cells (MNC) of patients with SLE associated with TP secreted highly elevated levels of TNF alpha compared to other patient groups. We suggest that abnormal production of various cytokines in SLE is an intrinsic defect of MNC and the immune system that may be the key element for a variety of clinical manifestations of this disease.

Topics: Adolescent; Adult; Arthritis, Rheumatoid; Behcet Syndrome; Concanavalin A; Cytokines; Female; Humans; Interferon-gamma; Interleukin-6; Lupus Erythematosus, Systemic; Lymphadenitis; Male; Middle Aged; Nephrotic Syndrome; Polymyalgia Rheumatica; Rheumatic Diseases; Sjogren's Syndrome; Thrombocytopenia; Tumor Necrosis Factor-alpha

The microheterogeneity of alpha 1 acid glycoprotein (AGP) was studied using affinity immunoelectrophoresis with concanavalin A (Con A) in serum samples of 43 patients with early rheumatoid arthritis (RA) without clinical features of intercurrent infection. The results were expressed as reactivity coefficients. Disease activity was measured by clinical (Lansbury's joint index, Mallya-Mace activity score) and laboratory (erythrocyte sedimentation rate, levels of soluble interleukin-2 receptor, C reactive protein, and AGP) indices. In contrast with previous reports, suggesting a decrease in AGP-Con A reactivity in patients with RA, high values of AGP reactivity coefficients were found in patients with disease of short duration, which were similar to those found in patients with acute bacterial infections. Conversely, normal or decreased values of AGP reactivity coefficients were found in patients with disease of longer duration. Regression analysis showed a significant relation between AGP reactivity coefficients and disease duration (multiplicative model). No other indices examined were significantly related to disease duration. These results, taken together with previous findings suggesting that cytokines control the glycosylation of acute phase proteins, indicate that differences in the microheterogeneity of AGP in early and longstanding RA reflect differences in cytokine action at different stages of the disease.

Topics: Adolescent; Adult; Aged; Arthritis, Rheumatoid; Blood Sedimentation; C-Reactive Protein; Concanavalin A; Female; Humans; Immunoelectrophoresis, Two-Dimensional; Male; Middle Aged; Orosomucoid; Receptors, Interleukin-2; Time Factors

The synovial tissue and fluid of patients with rheumatoid arthritis (RA) contain activated T cells that probably have a central role in the disease process which leads to joint destruction. A subset of T cells, gamma delta T cells detected at the site of inflammation, may be important in the pathogenesis of the disease. This study investigated variable (V) gene usage of gamma delta T cell receptors (TcRs) expressed in synovia and peripheral blood of patients with RA by using the polymerase chain reaction (PCR) to amplify TcR gamma- and delta-chain transcripts. Most patients showed no restriction in V gamma gene usage since synovial mononuclear cells (SMC) expressed TcR gamma-chain transcripts which used the same set of V gamma genes as peripheral blood mononuclear cells (PBMC). In contrast, the majority of patients expressed a restricted SMC V delta-chain repertoire biased towards V delta 1, but V delta 2 mRNA transcripts were also detected, albeit at low levels in some patients. The TcR delta-chain repertoires of PBMC from healthy control subjects were also characterized. There was variation in the TcR delta-chain repertoires of PBMC from patients when compared with controls, particularly with respect to expression of V delta 4. V delta 4 mRNA transcripts were expressed in PBMC of only two of seven RA patients in contrast with eight of the nine controls (P = 0.03). These findings are compatible with reports that gamma delta T cells in the rheumatoid synovium are reactive to Mycobacterium tuberculosis and that response to M. tuberculosis is restricted to V gamma 9/V delta 2-bearing T cells, if a superantigen is involved in the pathogenesis of RA.

Topics: Arthritis, Rheumatoid; Base Sequence; Concanavalin A; Gene Expression; Gene Rearrangement, delta-Chain T-Cell Antigen Receptor; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor; Humans; Molecular Sequence Data; Oligodeoxyribonucleotides; Polymerase Chain Reaction; Receptors, Antigen, T-Cell, gamma-delta; RNA, Messenger; Synovial Fluid; T-Lymphocytes

Recently in Japan, one form of vitamin B12, methylcobalamin also known as methyl B12, has attracted the attention of physicians as a therapy for patients with rheumatoid arthritis. However, its immunological actions in vivo are still unknown. In this study, we induced the in vitro production of such cytokines as interleukin-6 (IL-6), interferon-gamma (IFN-gamma), and interleukin-1 beta (IL-1 beta) by adding various mitogens (phytohemagglutinin:PHA, concanavalin A: ConA, or pokeweed mitogen:PWM) as well as recombinant interleukin-2, and we investigated the effects of methyl B12 (final concentration, 8-8,000 ng/ml) on the production of these cytokines by peripheral mononuclear cells. As compared to the controls, IL-6 production induced by PHA and ConA on Day 4 of the culture was suppressed by an average 60-70% when methyl B12 (80-8,000 ng/ml) was added to the medium. IFN-gamma production decreased dose-dependently with methyl B12, i.e., it decreased to 46% of the control when this production was induced by rIL-2, and decreased to 56-66% when it was induced by mitogens. The effect of methyl B12 on IL-1 beta production on Day I of the culture was small. These findings indicate that methyl B12 suppresses mainly the cytokine production of T lymphocytes. Such suppressive effects as shown in the in vitro situation are expected to be expressed also in vivo in patients with rheumatoid arthritis, especially at articulation lesion sites.

Topics: Arthritis, Rheumatoid; Concanavalin A; Cytokines; Humans; In Vitro Techniques; Interferon-gamma; Interleukin-1; Interleukin-2; Interleukin-6; Leukocytes, Mononuclear; Phytohemagglutinins; Pokeweed Mitogens; T-Lymphocytes; Vitamin B 12

Rheumatoid arthritis is a chronic inflammatory disorder with considerable evidence of impaired regulation of the immune response, including defective suppressor cell function, especially in the synovial membrane. We have investigated whether oxidation of cell surface thiols might be responsible for these defects and whether such cell function may be modulated towards normal by treatment with a sulphydryl-reactive drug, D-penicillamine. Using healthy mononuclear cells treated with an impermeant thiol blocker, induction of suppressor activity by incubation with the lectin Con A was not dependent on surface sulphydryl groups but suppressor activity was abolished by thiol blockade after Con A stimulation. Peripheral blood mononuclear cells from patients with active rheumatoid disease showed impaired Con A-induced suppressor activity which was enhanced to near-normal levels by incubating the rheumatoid cells with a sulphydryl reducing agent, 2-mercaptoethanol, or D-penicillamine. Con A-stimulation of cells from patients treated with intramuscular gold or D-penicillamine generated more active suppression than those from patients receiving non-steroidal drugs only. Mononuclear cells from patients with other chronic inflammatory joint diseases showed normal Con A-induced suppressor activity. These data support the conclusion that surface thiols on mononuclear cells in rheumatoid arthritis are reversibly oxidized by the disease process. This gives rise to aberrant cell function including impaired suppressor activity. Such a mechanism may be at least partly responsible for the defective immunoregulation seen in rheumatoid patients and thus be a relevant target for thiol containing antirheumatic drugs.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Concanavalin A; Humans; Immunoglobulin G; Immunoglobulin M; Mercaptoethanol; Monocytes; Oxidation-Reduction; Penicillamine; Phenylmercury Compounds; Sulfhydryl Compounds; T-Lymphocytes, Regulatory

We examined host ability to produce alpha- and gamma-interferon on a large scale by culturing 2 ml of peripheral blood for 20 hr with Sendai virus or concanavalin A as inducer of alpha- or gamma-interferon, respectively. Production of gamma- but not alpha-interferon was lower in females (n = 351) than in males (n = 531) (P less than 0.001). Both alpha- and gamma-interferon production declined gradually with ageing. The production of alpha-interferon (3,233 +/- 1,773 IU/ml) and gamma-interferon (19 +/- 20 IU/ml) in rheumatoid arthritis patients was significantly lower than the values found in total and age-matched healthy donors (P less than 0.01). These results suggest that interferon production is dependent on age and sex and is significantly lower in patients with rheumatoid arthritis.

Topics: Adolescent; Adult; Aged; Aging; Arthritis, Rheumatoid; Cells, Cultured; Child; Concanavalin A; Female; Humans; Interferon Type I; Interferon-gamma; Male; Middle Aged; Parainfluenza Virus 1, Human; Sex Factors

Microheterogeneity of acute phase proteins frequently differs in acute and chronic types of inflammation. However, it is unknown whether these changes depend on the duration of the inflammation in a given disease. We therefore investigated the microheterogeneity of alpha 1-acid glycoprotein (AGP) in sera from patients with acute and chronic bacterial infection in comparison to rheumatoid arthritis and ankylosing spondylitis. In acute bacterial infection Con A-reactivity of AGP was significantly elevated. By contrast, AGP in chronic bacterial infection showed the same glycosylation pattern as rheumatoid arthritis and ankylosing spondylitis being characterized by a decreased reactivity to Con A. Serial measurements in individual patients with bacterial infections showed a transition from the initially elevated to decreased reactivity to Con A as the disease became chronic.

Topics: Acute Disease; Adult; Aged; Aged, 80 and over; Arthritis, Rheumatoid; Bacterial Infections; C-Reactive Protein; Chronic Disease; Concanavalin A; Female; Glycosylation; Humans; Immunoelectrophoresis, Two-Dimensional; Male; Middle Aged; Orosomucoid; Spondylitis, Ankylosing; Staphylococcal Infections; Streptococcal Infections

There are contradictory reports on Interferon Gamma (IFN gamma) production by peripheral blood mononuclear cells (PBMC) in rheumatoid arthritis (RA). Since many patients previously studied were on Gold Sodium Thiomalate (GST), Auranofin (Auf), or D-Penicillamine (D-Pen) we have investigated the effects of these drugs on IFN gamma production using PBMC from normal controls (NC), and RA patients off GST, Auf, and D-Pen. Auf in low concentrations enhanced IFN gamma production by PBMC from NC but not RA; GST, and D-Pen had no effect. In other experiments PBMC were stimulated with concanavalin A (CONA A), or phytohemagglutinin (PHA). Auf, and GST inhibited IFN gamma production by CON A - stimulated NC and RA cells; D-Pen had no effect. Auf in low concentrations enhanced IFN gamma production by PHA - stimulated NC cells, but this effect was not seen with RA cells; GST inhibited both RA and NC cell production of IFN gamma, and D-Pen had no effect. Auf has a biphasic effect on IFN gamma production by NC cells with low concentrations being stimulatory or co-stimulatory, possibly by acting on T helper cells. Higher concentrations of Auf and GST, equivalent to those achieved in vivo in the course of therapy, inhibit IFN gamma production. These results suggest that gold therapy may affect IFN gamma production in RA, and could explain discrepancies noted in previous studies.

Topics: Adult; Aged; Arthritis, Rheumatoid; Auranofin; Concanavalin A; Female; Gold Sodium Thiomalate; Humans; In Vitro Techniques; Interferon-gamma; Leukocytes, Mononuclear; Male; Middle Aged; Penicillamine; Phytohemagglutinins

Synovial fluid (SF) from rheumatoid arthritis (RA) patients were tested for their ability to inhibit the proliferative responses of normal peripheral blood mononuclear cells (PBM) to mitogens and interleukin-2 (IL-2). SF significantly inhibited the responses to concanavalin A (CON A) and phytohaemagglutinin (PHA), but significantly enhanced the responses to IL-2. Similarly, SF mononuclear cells (SFM) were hyporesponsive to CON A and PHA compared with autologous PBM, but hyper-responsive to IL-2. It is concluded that an IL-2 inhibitor in RA SF is unlikely to be the cause of SFM hyporesponsiveness to mitogens.

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Interleukin-2; Leukocytes, Mononuclear; Phytohemagglutinins; Synovial Fluid

Quantitative changes of concanavalin A (Con A)-reactive proteins in serum samples obtained from rats with induced inflammation and from patients with inflammatory and autoimmune diseases were examined by use of lectin blots. Treatment of rats with a single dose of fermented yeast to induce inflammation caused an extensive increase in Con A-reactivity. These changes were time dependent and were similar in both sexes of the animals. When we examined serum samples obtained from patients with various inflammatory disorders for their Con A-reactive proteins as compared with normal donors, we noted that the Con A-reactivity increased in patients with rheumatoid arthritis and systemic lupus erythematosus. Among all the glycoproteins examined by lectin blots with use of Con A, a set of five proteins was selected for detailed analysis by densitometric scanning. These included alpha 2-macroglobulin, P-150, P-95, P-40, and P-35, of Mr 180,000, 150,000, 95,000, 40,000, and 35,000, respectively, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Densitometric scanning analysis of the lectin blots revealed that the Con A-reactivity of these proteins increased during inflammation. Because alpha 2-macroglobulin is not an acute-phase protein in humans, an increase in Con A staining of this protein suggested that altered glycation is associated with autoimmune diseases. Thus, study of changes in Con A-reactive proteins in human sera may facilitate our understanding of the etiology and pathophysiology of autoimmune diseases.

Topics: alpha-Macroglobulins; Animals; Arthritis, Rheumatoid; Autoimmune Diseases; Blood Proteins; Collodion; Concanavalin A; Electrophoresis, Polyacrylamide Gel; Female; Glycoproteins; Inflammation; Kinetics; Lupus Erythematosus, Systemic; Male; Molecular Weight; Rats; Rats, Inbred Strains

Despite chronic inflammation and the presence of hypergammaglobulinemia, rheumatoid factor (RF) is rarely found in the blood of patients with ankylosing spondylitis (AS). We used ELISA to compare spontaneous and pokeweed mitogen (PWM)-induced IgG, IgM and IgM rheumatoid factor (IgM-RF) production in normals and in patients with rheumatoid arthritis (RA) and AS. The IgG and IgM synthesis in these three groups did not differ. However, the IgM-RF level in PWM-induced mononuclear cell cultured supernatants of AS was significantly decreased, compared with normal and RA patients. Furthermore, mixing experiments by co-culture of normal T or B cells with patient's B or T cells in the presence of PWM revealed a deficiency of the helper T cell function in patients with AS. These results illustrate the cellular mechanism of the seronegativity of the rheumatoid factor in patients with ankylosing spondylitis.

Topics: Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Humans; Immunoglobulin G; Immunoglobulin M; Pokeweed Mitogens; Rheumatoid Factor; Spondylitis, Ankylosing; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory

Topics: Arthritis, Rheumatoid; Blood Protein Electrophoresis; Carbohydrates; Concanavalin A; Glycosylation; Humans; Orosomucoid; Sialic Acids

Topics: Arthritis, Rheumatoid; C-Reactive Protein; Carbohydrates; Concanavalin A; Genetic Variation; Humans; Immunoelectrophoresis; Lupus Erythematosus, Systemic; Orosomucoid; Rheumatic Diseases; Spondylitis, Ankylosing

We evaluated the clinical usefulness of determinations of alpha 1-acid glycoprotein (AGP) microheterogeneity in distinguishing patients who have active rheumatoid arthritis (RA) from those who have RA and an intercurrent infection. AGP microheterogeneity was studied by affinity electrophoresis with concanavalin A as the ligand, and the results were expressed as reactivity coefficients (RC). Significant differences were found between the mean RC (+/- SD) in healthy individuals (1.27 +/- 0.16) and the mean RC in RA patients with intercurrent infection (1.74 +/- 0.90), as well as with the mean RC in RA patients with grades III and IV disease activity (0.92 +/- 0.18 and 0.81 +/- 0.25, respectively). Moreover, an additional microheterogeneous form of AGP, similar to that observed in non-RA patients with infections, was noted in RA patients with infections (sensitivity 100%, specificity 100%). The results show that an increase in AGP reactivity with concanavalin A is a sensitive indicator of intercurrent infection in patients with RA.

Topics: Arthritis, Rheumatoid; Bacterial Infections; C-Reactive Protein; Concanavalin A; Electrophoresis, Gel, Two-Dimensional; Humans; Orosomucoid

Interleukin-2 (IL-2) receptor bearing cells and soluble IL-2, measured in a bioassay with IL-2 dependent human T-cell blasts, were recognized in synovial fluid, but not in the peripheral blood of patients with rheumatoid arthritis (RA). After stimulation in vitro with appropriate concentrations of the mitogen concanavalin A (Con-A), comparable proportions of IL-2 receptor (IL-2R) bearing cells were seen in cultures of synovial fluid lymphocytes (SFL) and in cultures of peripheral blood lymphocytes (PBL). On the other hand, higher levels of secreted IL-2 were found in SFL cultures compared to corresponding PBL cultures of RA patients and normal donors. Specificity of the IL-2 bioassay was confirmed by blocking the T-cell blast proliferation (induced by SFL culture supernatants), by 83 +/- 4%, after addition of a monoclonal anti-IL-2R antibody. Despite the high levels of soluble IL-2, only a weak proliferative response was observed in the corresponding SFL cultures.

Topics: Adult; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Interleukin-2; Lymphocyte Activation; Male; Middle Aged; Receptors, Interleukin-2; Synovial Fluid; T-Lymphocytes

Tests for lymphoproliferation and interferon induction in normal blood donors and patients with rheumatoid arthritis (RA) were performed in a whole-blood assay. Patients with high inflammatory RA showed significantly reduced lymphoproliferation and interferon gamma production after stimulation with phytohemagglutinin (PHA) and concanavalin A (Con A) when compared with patients with low inflammatory activity, or with normal control individuals. Similarly, patients with high and low inflammatory RA exhibited a significantly reduced interferon alpha production after stimulation with Newcastle Disease Virus (NDV) when compared with normal blood donors. Our findings may point to an important immunodeficiency of the circulating lymphocytes of RA patients and may explain some of the in vitro immunoregulatory abnormalities reported in this disease.

Topics: Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Humans; Interferon Type I; Interferon-gamma; Leukocyte Count; Leukocytes; Lymphocyte Activation; Phytohemagglutinins; Pokeweed Mitogens

The microheterogeneity of alpha 1-acid glycoprotein (AGP) has been studied in the sera of 48 patients with rheumatoid arthritis and of 12 healthy individuals. For each rheumatoid patient the disease activity has been assessed and each patient has been assigned to one of four activity grades: I, inactive; II, mildly active; III, moderately active; and IV, severe. Concanavalin A-affinity electrophoresis revealed three microheterogeneity variants of AGP: non-reactive with Con A, weakly reactive with Con A and strongly reactive with Con A. The relative amounts of AGP-variants observed in the healthy donors were similar to those observed in the patients with activity grade I, but differed significantly from patients with grades II, III and IV. The differences between the AGP-patterns of patients with activity grades II, III and IV were also statistically significant.

Topics: Arthritis, Rheumatoid; C-Reactive Protein; Concanavalin A; Electrophoresis; Humans; Immunoassay; Orosomucoid

A solid phase radioimmunoassay was set up for direct measurement of the binding capacity of human IgG to three lectins recognizing different carbohydrates of the Fc domain, i.e. peanut agglutinin (PNA), Concanavalin A (Con A) and pokeweed mitogen (PWM) which mainly bind to beta-galactose, alpha-mannose and dimers of N-acetyl-beta-glucosamine respectively. The mean specific binding of the 96 normal IgG tested to PNA and to PWM was statistically higher (P less than 0.001) than that to Con A, whereas no significant differences were observed between the mean specific bindings to PNA and to PWM. A statistically significant linear negative correlation could be established only between the relative bindings (expressed in percentage of the total binding to the three lectins) to PNA and to PWM (r = -0.65, P less than 0.001). The mean specific binding of IgG purified from 34 patients suffering from rheumatoid arthritis (RA) to PNA and to Con A was statistically higher (P less than 0.001) than that reached with PWM, whereas no significant differences were noted between their mean binding capacities to PNA and to Con A. When compared to normal IgG, only four out of 34 RA IgG exhibited a significantly higher binding capacity to PNA, whereas all but one RA IgG possessed a significantly higher binding capacity to Con A. Accordingly, the mean specific binding of RA IgG to Con A was significantly higher than that of normal IgG (P less than 0.001). Besides (and contrary to normal IgG), a statistically significant negative linear correlation was noted between the relative bindings of RA IgG to PNA and to Con A (r = -0.89, P less than 0.001). All the five RA IgG tested exhibited an abnormal circular dichroism. Our data suggest that, by altered steric conformation and glycosylation, mannosyl-residues of RA IgG become prominent or terminal or both, and are therefore able to react more effectively with Con A than normal IgG do.

Topics: Adult; Aged; Arthritis, Rheumatoid; Circular Dichroism; Concanavalin A; Female; Humans; Immunoglobulin G; Lectins; Male; Middle Aged; Peanut Agglutinin; Pokeweed Mitogens

Microheterogeneity of two acute-phase proteins: orosomucoid (alpha1-acid glycoprotein, AGP) and alpha 1-antichymotrypsin (ACHT) were studied in the sera of 48 patients with rheumatoid arthritis and 12 healthy individuals. Each rheumatoid patient was assigned to one of four activity grades. Cross affinoimmunoelectrophoresis (aff-EP) with free Concanavalin A (Con A) revealed three microheterogeneity variants of AGP and four microheterogeneity forms of ACHT. The relative amounts of AGP-variants and ACHT-variants observed in the healthy donors were similar to those observed in the patients with activity grade I, but they changed with the increase in the grade of rheumatoid activity. The differences were most significant for AGP. The comparison of serum C-reactive protein (CRP) levels with AGP-variant-O-non reactive with Con A showed significant correlation in respective rheumatoid activity grades.

Topics: Acute-Phase Proteins; alpha 1-Antichymotrypsin; Arthritis, Rheumatoid; Blood Proteins; Chromatography, Affinity; Chymotrypsin; Concanavalin A; Humans; Immunoelectrophoresis; Orosomucoid; Protease Inhibitors

The Fc-receptor (Fc-R) function of monocytes isolated from 19 control subjects and from 30 patients presenting with a rheumatoid arthritis (RA) was assessed in vitro by a classical rosette assay using IgG-coated sheep red blood cells. In RA patients, the percentage of monocytes forming rosettes was significantly lower than in controls (34.4 +/- 20.4 versus 67.4 +/- 4.5%; P less than 0.001). The blockade observed was reversed by a prior trypsin treatment of RA monocytes, the percentage of recovery being correlated with the IgG plasma levels. Besides, IgG purified from the serum of four RA patients bound a mean of 7.3, 5.2, 1.6, and 1.6 times more than normal IgG did onto concanavalin A (Con A), peanut agglutinin (PNA), phytohemagglutinin (PHA), and pokeweed mitogen (PWM), respectively. Although similar amounts of 125I-labeled normal and RA IgG were bound to normal monocytes, RA IgG inhibited more efficiently than normal IgG the Fc-R function of normal monocytes, for all concentrations tested (10 to 100 micrograms/100 microliters). A prior treatment of RA IgG by alpha-mannosidase, but not by beta-galactosidase, significantly reduced their inhibitory properties. The incubation of monocytes with D-mannose or mannan reduced their capacity to form rosettes. The percentage of monocytes forming rosettes in the presence of both mannan and normal IgG was significantly lower than that measured in the presence of normal IgG only. On the contrary, the rosetting capacity of monocytes in the presence of both RA IgG and mannan was the same as that calculated in the presence of RA IgG only. The inhibitory effect of RA IgG was not related to their abnormal circular dichroism. Our data suggest that the greater ability of RA IgG to block the Fc-R function of monocytes probably depends on the presence of a greater number of accessible mannosyl residues on the glycosidic side chains located in the Fc domain of the molecules.

Topics: Adult; Aged; Arthritis, Rheumatoid; Carbohydrates; Concanavalin A; Female; Humans; Immunoglobulin Fab Fragments; Immunoglobulin G; In Vitro Techniques; Male; Middle Aged; Monocytes; Receptors, Complement; Receptors, Complement 3b; Receptors, Fc

Synovial fluid lymphocytes (SFL) and peripheral blood lymphocytes (PBL) from patients with rheumatoid arthritis (RA) and reactive oligoarthritis were investigated for activated T cells (Ia+SIg-), IL-2 receptor bearing cells (Tac+) and IL-2 production in vivo and in vitro. In contrast to negative results with blood, the synovial fluid of the arthritic joints contains considerable amounts of IL-2 activity (median: 11.8 mu/ml), elevated proportions of Ia+SIg- activated T cells (median: 12.5%) and of IL-2 receptor bearing cells (median: 2.5%). In vitro, after stimulation with several Concanavalin A (Con A) doses, SFL develop proportions of IL-2 receptor cells comparable to PBL. Furthermore, they produce higher values of IL-2 activity than comparable PBL cultures. The proportions of Ia+SIg- activated T cells increase only moderately after Con A stimulation compared to in vivo data, indicating different activated T cell subsets in the synovial fluid (Ia+SIg-, Tac+). The findings are discussed as an expression of an acute hyperactivation of lymphocytes in an inflamed joint.

Topics: Adolescent; Adult; Aged; Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Female; Humans; Interleukin-2; Lymphocyte Activation; Male; Middle Aged; Receptors, Immunologic; Receptors, Interleukin-2; Synovial Fluid; T-Lymphocytes

A new system for lectin-glycoprotein complexes dissociation constants (K) determination is presented. The system is based on agarose affinity electrophoresis where equal lectin (Con A) concentrations are inhibited by variable specific sugar (alpha-methyl-mannoside) amounts. Moreover, the system allows lectin-sugar inhibition constants (Ki) studies. For determination of mechanisms as well as K and Ki values calculations mathematical equations are developed. Values of K for two variants of alpha 1 acid-glycoprotein, two variants of alpha 1-antitrypsin, one variant of alphafetoprotein and Ki for Con A-alpha-MM are calculated according to the introduced equations and compared in a computed system. Moreover, the influence of sugar on lectin-glycoproteins interaction is demonstrated and discussed.

Topics: alpha 1-Antitrypsin; alpha-Fetoproteins; Arthritis, Rheumatoid; Concanavalin A; Electrophoresis, Agar Gel; Glycoproteins; Humans; Kinetics; Lectins; Macromolecular Substances; Orosomucoid; Protein Binding

Activity of short-lived suppressor lymphocytes was studied in 26 patients with rheumatoid arthritis. Of these, 14 patients had diverse systemic manifestations: rheumatoid nodules, polyneuropathy. Sjögren's syndrome, and Felty's syndrome. It was established that attenuation of the suppressor activity in rheumatoid arthritis is characteristic of patients with systemic manifestations, who show a high level of circulating immune complexes of rheumatoid factors. The most informative data were obtained as a result of the use of the suboptimal dose of Con A having suppressor activity.

Topics: Adult; Aged; Arthritis, Rheumatoid; Cell Survival; Concanavalin A; Female; Humans; In Vitro Techniques; Leukocyte Count; Lymphocyte Activation; Male; Middle Aged; Pokeweed Mitogens; T-Lymphocytes, Regulatory; Time Factors

The proliferation of peripheral blood lymphocytes, cultured with Con A, can be inhibited by ionizing radiation. Lymphocytes from patients with conditions associated with autoimmunity, such as rheumatoid arthritis, systemic lupus erythematosus and polymyositis, are more radiosensitive than those from healthy volunteers or patients with conditions not associated with autoimmunity. The nuclear material isolated from the lymphocytes of patients with autoimmune diseases is, on average, lighter in density than the nuclear material from most healthy controls. This difference in density is not related to increased sensitivity to ionizing radiation but the degree of post-irradiation change in density (lightening) is proportional to the initial density, i.e. more dense nuclear material always shows a greater upward shift after radiation. The recovery of preirradiation density of nuclear material, 1 h after radiation exposure, taken as an indication of DNA repair, correlates with the radiosensitivity of lymphocyte proliferation (Con A response); failure to return to pre-irradiation density being associated with increased sensitivity of proliferative response. These results require extension but, taken with previously reported studies of the effects of DNA methylating agents, support the idea that DNA damage and its defective repair could be important in the aetio-pathogenesis of autoimmune disease.

Topics: Adult; Aged; Arthritis, Rheumatoid; Autoimmune Diseases; Cell Division; Cell Nucleus; Concanavalin A; DNA; DNA Repair; Humans; Lupus Erythematosus, Systemic; Lymphocytes; Middle Aged; Myositis; Radiation Tolerance

Topics: Adult; Aged; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Lymphocyte Activation; Male; Middle Aged; Mitogens; Phytohemagglutinins; Pokeweed Mitogens; T-Lymphocytes

The disordered cellular immunity of rheumatoid arthritis (RA) was studied in vitro by assessing the stimulatory effect of tuberculin PPD, a mycoplasma product with major histocompatility complex restricted properties in mice (MAS) and concanavalin A (Con A) on RA and control cells of known tissue type. PPD provided the best discrimination between RA and controls (RA responses 34% of latter). The only significant association with HLA-DR type was that patients with RA who exhibited poor responsiveness to MAS had a significantly increased frequency of HLA-DR4.

Topics: Arthritis, Rheumatoid; Concanavalin A; Female; Histocompatibility Antigens Class II; HLA-DR4 Antigen; Humans; Immunity, Cellular; In Vitro Techniques; Lymphocyte Activation; Male; Middle Aged; Mycoplasma; Tuberculin

Peripheral blood (PBL) and synovial fluid lymphocytes (SFL) from 18 patients with definite rheumatoid arthritis (RA) and from one patient with Reiter's disease (RD) were examined for their capacity to absorb quantitatively interleukin-2 (IL-2) from a standardized, lectin-free IL-2 source. For comparison normal ConA blasts and PBL from various inflammatory and noninflammatory diseases as well as from healthy control persons were studied. IL-2 activity was quantitated by measuring 3H-thymidine-2-deoxyriboside uptake in the IL-2 dependent murine T cell line CTL6. ConA blasts exhibited a high IL-2 absorption capacity and served as a positive control for calibrating the absorption assay. In a population of normal PBL at least 5-10% of ConA blasts were required to detect IL-2 absorption. Significant absorption was assumed if more than 50% of IL-2 activity was removed from 200 microliters of a lectin-free IL-2 standard following incubation with 5 X 10(6) lymphoid cells for 2 h at 4 degrees C; this criterion was fulfilled with 8 out of 20 SFL and 4 out of 14 PBL preparations from RA patients. As a rule SFL absorbed more IL-2 than PBL. Control PBL did not absorb significant quantities of IL-2. PBL from the RD patient apparently produced an IL-2 inhibitor during incubation with the IL-2 standard. IL-2 absorption by ConA blasts and SFL was fully inhibited by preincubation of the absorbing cells with monoclonal anti-TAC antibody, a reagent known to react with the human IL-2 receptor. The results are discussed in view of current concepts of antigen/mitogen induced T cell activation.

Topics: Adult; Aged; Antibodies, Monoclonal; Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Female; Humans; Interleukin-2; Male; Middle Aged; Receptors, Immunologic; Receptors, Interleukin-2; Synovial Fluid; T-Lymphocytes

Several pharmacological agents, some of which are known to have effects on the immune system, decrease the incidence of collagen II-induced arthritis when added to the antigen emulsion. Concanavalin A, which has been reported to exert suppressive effects on the immune system in vivo, consistently reduced the immune response to the collagen antigen. These effects were dose and time dependent. The suppressive effects of pokeweed mitogen, tilorone and carrageenan on anti-collagen II responses were somewhat variable. Suppressive activity could be observed with concanavalin A and levamisole when the drugs were injected at a site distant from the collagen emulsion. These studies indicate that local administration of drugs is an effective approach for demonstrating the activity of some agents that may alter the course of collagen II disease through an effect on the immune system.

Topics: Animals; Antigens; Arthritis; Arthritis, Rheumatoid; Collagen; Concanavalin A; Emulsions; Female; Hypersensitivity, Delayed; Levamisole; Rats

In chronic rheumatoid arthritis, mononuclear cells (MC) accumulate in the subchondral bone and form a prominent part of both destructive lesions and repair reactions. A fraction from human bone matrix extracts (BME) stimulated glycosaminoglycan (gag) and glycoprotein synthesis by fibroblastic cells but its effects on MC metabolism had not been studied. A method was established for the study of incorporation of radioactive precursors into total protein, IgG and gag synthesized and secreted by peripheral blood MC cultured in microwells in the presence or absence of Concanavalin A (ConA). Relatively low concentrations of BME suppressed spontaneous synthesis of radioactive IgG (protein A bound) and TCA precipitable protein but had little effect on gag synthesis. In general, stimulation of the cultures with ConA overcame the inhibitory effects on protein synthesis by the BME. A large stimulation of gag synthesis induced by ConA was not affected by BME. The interactions between the BME and the stimulatory effect of ConA on DNA synthesis were studied in detail and were found to be complex, not immunologically specific and appeared to be due to binding of lectin by the carbohydrate moieties of the glycoproteins in the BME. On the basis of a model of the lectin-BME interaction, the hypothesis is postulated that the carbohydrate moieties of subchondral bone glycoproteins may have the capacity to act as a solid state "trap" for certain circulating antigens which may then also interact with surface glycoproteins of the MC accumulating in the subchondral bone. The physiological role of gag synthesis by MC is not known.

Topics: Arthritis, Rheumatoid; Bone Matrix; Cells, Cultured; Chemical Precipitation; Concanavalin A; DNA; Glycoproteins; Glycosaminoglycans; Humans; Immunoglobulin G; Kinetics; Lymphocytes; Monocytes; Protein Biosynthesis

Nephelometry of serum acute-phase glycoproteins by binding to concanavalin A (con-A) was compared with assays for haptoglobin and alpha 1-acid glycoprotein, and for C-reactive protein. The cutoff points for positive reactions were determined on the basis of results for a random sample (n = 130) from a middle-aged population. The sensitivity of the con-A binding assay compared favorably with that of individual acute-phase glycoproteins in a follow-up cohort of 198 patients with inflammatory joint diseases. Unlike the case in many individual acute-phase glycoprotein assays, the distribution of con-A binding values in healthy subjects is remarkably symmetrical, allowing an easy distinction between abnormal and normal values.

Topics: Acute-Phase Proteins; Adolescent; Adult; Aged; Arthritis; Arthritis, Rheumatoid; Blood Proteins; C-Reactive Protein; Concanavalin A; Glycoproteins; Haptoglobins; Humans; Immunochemistry; Middle Aged; Nephelometry and Turbidimetry; Orosomucoid; Protein Binding

Concanavalin A (Con A)-induced suppressor T cell activity was determined in 10 rheumatoid arthritis (RA) patients with vasculitis, 34 RA patients without vasculitis, and 10 healthy individuals. The percent Con A-induced suppression in RA patients with vasculitis was 24.6. In contrast, it was 68.4% in those RA patients without vascular lesions. Further, the proportion of T cells reactive with OKT8 monoclonal antibody was also decreased in RA patients with vasculitis. Accordingly, the reduced Con A-induced suppressor T cell activity in these RA patients resulted, in part, from the reduction in the number of cells of the suppressor T cell subset. Those patients with vascular lesions also had a higher percentage of positive antilymphocytotoxic antibodies than RA patients without vasculitis. Since the differences in Con A-induced suppressor T cell activity and frequency of positive antilymphocytotoxic antibodies were so great, we believe RA patients with vasculitis could be recognized as a disease group distinct from RA patients without vasculitis.

Topics: Adult; Aged; Antibodies, Monoclonal; Antilymphocyte Serum; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Immunoglobulin G; Lymphocytes; Male; Middle Aged; Pokeweed Mitogens; Rheumatoid Factor; T-Lymphocytes; T-Lymphocytes, Regulatory; Vasculitis

Peripheral blood lymphocytes (PBL) from patients with rheumatoid arthritis (RA) and age and sex-matched normal controls were compared in a Concanavalin A (Con A)-induced suppressor cell assay. Suppression induced by pre-incubation with Con A was significantly greater in PBL from RA patients than in PBL from normal controls. Preincubation with thymosin fraction 5, in the absence of Con A, also induced greater suppressor cell activity in PBL from normal controls. Preincubation with Con A and thymosin, simultaneously, induced suppression similar to that seen with Con A alone. These results suggest the presence of an immunoregulatory defect in RA, characterized by an excess of both Con A and thymosin-inducible suppressor cells, that may play a role in disease pathogenesis. The implications of these observations for immunotherapy of rheumatoid arthritis with thymosin are discussed.

Topics: Adult; Aged; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Lymphocyte Culture Test, Mixed; Male; Middle Aged; Rosette Formation; T-Lymphocytes, Regulatory; Thymosin

The ability of peripheral blood lymphocytes from patients with systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and Sjogren's syndrome (SS) to produce interleukin 2 (IL-2) and respond to it in-vitro was examined. Phytohemagglutinin-stimulated lymphocytes from over half of the SLE patients exhibited a decreased ability to produce IL-2 while their concanavalin A-generated blast cells responded normally to exogenous IL-2. Lymphocytes from RA patients not only produced less IL-2 than normals (P less than 0.001), but also responded poorly to exogenous IL-2 (P = 0.011). These abnormalities did not correlate with the patient's age, sex, duration of disease, or disease activity. Production of and response to IL-2 was widely varied among patients with SS and not different from controls. The decreased response of RA lymphocytes to IL-2 may result from a smaller number of cell surface IL-2 receptors since IL-2 adsorption to RA cells was lower than to either SLE or normal cells. These data suggest that IL-2-related abnormalities may play a role in the disordered immunoregulation characteristic of RA and perhaps of SLE.

Topics: Absorption; Adult; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Interleukin-2; Lupus Erythematosus, Systemic; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Receptors, Immunologic; Receptors, Interleukin-2

Lymphocyte responsiveness to various mitogens was studied in patients with palindromic rheumatism. The results show an enhanced lymphocyte reactivity to T-dependent mitogens during the first few days after an acute attack of the disease.

Topics: Adult; Arthritis, Rheumatoid; Concanavalin A; Female; Humans; Lymphocyte Activation; Lymphocytes; Male; Mitogens; Phytohemagglutinins; Pokeweed Mitogens

In patients with rheumatoid arthritis (RA) a significantly decreased autologous mixed lymphocyte reaction (AMLR) was observed which varied widely and did not correlate with disease activity, clinical course or treatment schedules. When supernatants of AMLR cultures were tested for the presence of soluble factors no differences were found in regard to the suppression of allogeneic and mitogen induced lymphocyte proliferation. Furthermore both test groups failed to produce detectable amounts of interferon during the course of the AMLR, in contrast to the allogeneic situation were both RA patients and normal controls exhibited a similar interferon activity in the culture supernatants.

Topics: Adult; Aged; Antigens; Arthritis, Rheumatoid; Autoantigens; Concanavalin A; Humans; In Vitro Techniques; Interferons; Isoantigens; Lupus Erythematosus, Systemic; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Middle Aged; T-Lymphocytes

Isoprinosine (IPS) is a new anti-viral agent which appears to have immunomodulatory activities which include its ability to enhance the in vitro blastogenic responses of normal lymphocytes to mitogens. The present study compares the effects of IPS on the in vitro immune functions of peripheral blood mononuclear cells (PBMC) from systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) patients with its effects on PBMC from normal controls. Each mitogen (Con A, PHA or PWM) was used at its optimal concentration with a range of IPS concentrations (0-25 micrograms/ml). PHA-induced blastogenesis by PBMC from all three groups was enhanced by IPS at or above 5 micrograms/ml. The Con A-induced responses of SLE lymphocytes were significantly enhanced over controls by IPS (P less than 0.02 at 5 micrograms/ml) while those of RA lymphocytes were not. IPS had little effect on PWM-induced blastogenesis by RA lymphocytes but did enhance the blastogenic responses of SLE lymphocytes (P less than 0.01 at 5 micrograms/ml). In contrast, the characteristically high immunoglobulin synthesis by SLE lymphocytes was decreased by IPS. The mechanism responsible for these effects is not known but IL-2 production by patient lymphocytes in vitro which was low for both RA (P less than 0.01) and SLE (P less than 0.02) increased significantly (P less than 0.05) when SLE lymphocytes were cultured with IPS. These data identify IPS as an agent for the study of aberrant immune regulation in autoimmune diseases and suggest that it may have potential therapeutic value in SLE.

Topics: Adult; Arthritis, Rheumatoid; Autoimmune Diseases; Cells, Cultured; Concanavalin A; Humans; Immunoglobulins; Inosine; Inosine Pranobex; Interleukin-2; Lupus Erythematosus, Systemic; Lymphocyte Activation; Lymphocytes; Middle Aged; Mitosis; Phytohemagglutinins

Human peripheral mononuclear cells (MC) secrete 2 soluble activities that modulate the growth of human synovial fibroblastic cells. A growth-suppressive, lectin-dependent activity is elaborated by the non-adherent population and its secretion begins before DNA synthesis is initiated in concanavalin-stimulated MC cultures. The elaboration of this activity is partially dependent on the presence of serum and it appears to be distinct from virus-induced human leukocyte interferon. The second activity is secreted spontaneously by the MC, under a variety of culture conditions including supplementation with homologous human plasma, and it enhances the growth of synovial fibroblasts. The rate of secretion of the growth-enhancing activity by the nonstimulated MC approximately parallels that of the inhibitory activity from the stimulated MC cultures. MC from patients with rheumatoid arthritis and from nonaffected individuals secrete similar concentrations of growth-stimulatory activity for the synovial fibroblasts.

Topics: Arthritis, Rheumatoid; Cell Division; Cells, Cultured; Concanavalin A; Culture Media; DNA; Fibroblasts; Growth Inhibitors; Growth Substances; Humans; In Vitro Techniques; Interferons; Monocytes; Synovial Membrane

Normal controls and patients with rheumatoid arthritis (RA) were investigated with respect to quantitative lymphocyte proliferation (LP) after concanavalin-A (conA) activation and to conA-induced suppressor cell activity (conA-SC). Measurements and assessment of RA activity were made at the beginning and end of a 10-day fast. The controls showed depressed (p less than 0.05) LP at the end of the fast, but no change in conA-SC activity. The RA group showed subnormal (p less than 0.05) LP and conA-SC (p less than 0.01) at the beginning of the experiment. After fasting they showed clinical improvement, the LP was not further depressed, and the initially low conA-SC had become normal.

Topics: Adult; Aged; Arthritis, Rheumatoid; Concanavalin A; Fasting; Female; Humans; Lymphocyte Activation; Male; Middle Aged; T-Lymphocytes, Regulatory

The effects of various chemotactic factor generation conditions and several chemotherapeutic agents on neutrophil migration were determined using in vitro assay systems designed as models for inflammatory processes occurring in the synovial cavities of patients with rheumatoid arthritis. The microtubule-promoting agent concanavalin A and the microfilament-disrupting agent cytochalasin B were shown in these systems to inhibit neutrophil migration towards zymosan-activated serum-derived chemotactic factors. Neutrophils, immunoglobulin G aggregates, and serum were required for maximum generation of comparable chemotactic factors. Insoluble immunoglobulin G aggregates with or without rheumatoid factor produced more chemotactic factor activity on interaction with neutrophils than soluble immunoglobulin G aggregates. Exposure of neutrophils to supratherapeutic levels of the nonsteroidal antiinflammatory agent aspirin decreased neutrophil response to chemotactic factors while exposure to the slow-acting or immunomodulating agents gold, D-penicillamine, or azathioprine had no effect on this neutrophil function. In vitro systems employing neutrophils, insoluble aggregates, and serum may offer useful means for assaying drug effects on important functional components of the rheumatoid inflammatory process.

Topics: Antigen-Antibody Complex; Arthritis, Rheumatoid; Blood Physiological Phenomena; Chemotaxis, Leukocyte; Concanavalin A; Cytochalasin B; Humans; Immunoglobulin G; Inflammation; Macromolecular Substances; Neutrophils

The ability of rheumatoid sera to support concanavalin-A transformation of normal lymphocytes was inversely related to serum C1q binding activity. When C1q binding activity of the sera was removed by absorption with staphylococcal protein A, subsequent lymphocyte response increased to the level found in immune-complex negative sera. Gel filtration of a small number of sera suggested that the suppressive material had a molecular weight in the range 1.8-4.9 x 10(5) daltons. Aggregated human gammaglobulin suppressed con-A transformation of normal lymphocytes in a dose-dependent fashion. These results suggest that immune complexes present in rheumatoid sera can suppress lymphocyte responsiveness. The relevance of this observation to be clinical features of rheumatoid arthritis is discussed.

Topics: Adult; Aged; Antigen-Antibody Complex; Arthritis, Rheumatoid; Complement Activating Enzymes; Complement C1q; Concanavalin A; Female; gamma-Globulins; Humans; Immunosorbents; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Staphylococcal Protein A

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Lymphokines; Suppressor Factors, Immunologic; T-Lymphocytes, Regulatory

Topics: Adult; Aged; Antibodies, Monoclonal; Arthritis, Rheumatoid; B-Lymphocytes; Concanavalin A; Female; Humans; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Male; Middle Aged; Pokeweed Mitogens; Receptors, Antigen, T-Cell; Rosette Formation; T-Lymphocytes; T-Lymphocytes, Regulatory

Peripheral blood lymphocytes from 30 patients with rheumatoid arthritis and 14 controls were examined for suppressor activity by two different assays. These were the Concanavalin-A-induced and the short-lived suppressor cell assays. There was no difference in suppressor activity between patients and controls, the suppressor activity of HLA-DR3 positive patients was no less than that of non-DR3 patients. However, patients with nodules showed reduced suppression in the short-lived suppressor cell assay when compared with patients without nodules.

Topics: Adult; Arthritis, Rheumatoid; Autoimmune Diseases; Concanavalin A; Female; Histocompatibility Antigens Class II; HLA-DR3 Antigen; Humans; Immunoassay; Male; Middle Aged; T-Lymphocytes, Regulatory

Patients with rheumatoid arthritis (RA) have diminished peripheral blood cellular immune responses in vivo and in vitro. To determine whether these changes reflected excessive peripheral blood suppressor function, we compared suppressor function in 52 patients with RA with 57 normal subjects. We examined suppressor cell function for (i) preincubation sensitivity, (ii) glass adherence, (iii) indomethacin sensitivity, (iv) concanavalin (con) A inducibility, (v) aggregated IgG inducibility and (vi) quantitated Tmu and Tgamma cell populations. Con A-generated suppressor cell function was further studied by evaluating unseparated-, T-cell, glass adherent-, indomethacin sensitive-, radiosensitive-, and radioresistant-suppressor cell effects upon mitogen-stimulated allogeneic and autologous unseparated, T-cell, B-cell and IM9 lymphoblastoid cell line responses, the latter representing a newly devised and simplified assay system. We found that patients with RA exhibited: (i) significantly increased incubation-sensitive suppressor function; (ii) generally normal nonspecific suppression function in other assays, although individual patients were strikingly abnormal; and (iii) significantly increased sensitivity of B-cell proliferation to aggregated-IgG-induced T-cell suppressor function.

Topics: Arthritis, Rheumatoid; B-Lymphocytes; Cell Adhesion; Cells, Cultured; Concanavalin A; Humans; Indomethacin; Lymphocyte Activation; Mitogens; T-Lymphocytes; T-Lymphocytes, Regulatory

A simplified, sensitive assay has been devised to examine suppressor cell function in normal persons and patients with rheumatic and atopic diseases. Cultured human lymphoblastoid (IM9) cells were used as responders. Induced suppressor cells were treated initially with mitomycin (mit) C, to prevent DNA synthesis, then incubated with concanavalin (con) A in microtiter plates for 40 hr. Responder cells were added directly to suppressor cells in plates. Suppression was determined by comparing effects of con A-induced with noninduced (control) cells on responder 3H-thymidine (3HTdR) uptake. This assay is simple and conserves time, reagents, and cells and uses standardized, available responder cells. It also obviates problems of con A in cultures with responder cells and autologous or allogeneic mixed-lymphocyte types of reactions and reduces needs for blood donation. Moreover, IM9 cells proved suitable for detecting spontaneous, con A-generated, glass-adherent, or prostaglandin-secreting (indomethacin-sensitive) suppressor cells.

Topics: Arthritis, Rheumatoid; Cell Line; Cell Survival; Cells, Cultured; Concanavalin A; Humans; Immunologic Techniques; Mitomycin; Mitomycins; T-Lymphocytes, Regulatory

Suppression of the blastogenic response by autologous cells precultured with Concanavalin A (Con A) or Phytohemagglutinin (PHA) was studied in an in vitro system. The mean mitogen response of cells from normal individuals and from patients with Rheumatoid Arthritis (RA) in isologous plasma was similar. In isologous plasma there was no significant difference between the activity of Con A suppressor cells of normals and those of RA. Incubation of the cells in RA plasma abrogated the suppression in both normals and RA patients. In isologous plasma PHA suppressor cells were also induced in both groups, the levels being lower in the normals than in the RA group. PHA suppressor cell activity against PWM induced blastogenesis could not be engendered in normals. RA plasma abrogated PHA suppressor cell activity, especially in the RA group. The abrogating activity of RA plasma was heat stable, nondialysable and could be correlated with the presence of anti-lymphocyte antibodies in the RA plasma.

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Immune Tolerance; Lymphocyte Activation; Middle Aged; Phytohemagglutinins; T-Lymphocytes, Regulatory

Topics: Adult; Arthritis, Rheumatoid; Ascorbic Acid; B-Lymphocytes; Chediak-Higashi Syndrome; Concanavalin A; Cyclic GMP; Humans; Hypersensitivity, Delayed; Immunity, Cellular; Immunoglobulins; In Vitro Techniques; Lymphocytes; Middle Aged; Monocytes; Pokeweed Mitogens; T-Lymphocytes, Regulatory

Topics: Adult; Arthritis, Rheumatoid; Concanavalin A; Humans; Indomethacin; Male; T-Lymphocytes, Regulatory

Studies were performed using an in vitro assay system to determine whether or not methyl-B12 could affect human T-cell function. When T cells were stimulated with phytohemagglutinin and allogeneic B cells, methyl-B12 did not enhance T-cell proliferation. In contrast, remarkable enhancing effects of methyl-B12 on the proliferative response to concanavalin A (Con A) and autologous B cells at suboptimal concentrations were observed, ranging from 0.1 to 10 micrograms/ml. Concentrations of methyl-B12 sufficient to enhance cellular proliferation were able to enhance the activity of helper T cells for immunoglobulin synthesis of B cells by pokeweed mitogen. Furthermore, the presence of methyl-B12 significantly potentiated the induction of suppressor cells in Con A-activated cultures. These results suggest that methyl-B12 could modulate lymphocyte function through augmenting regulatory T-cell activities.

Topics: Antibody-Producing Cells; Arthritis, Rheumatoid; B-Lymphocytes; Cell Differentiation; Concanavalin A; Humans; Immunity, Cellular; Immunoglobulin M; Lymphocyte Activation; Phytohemagglutinins; Pokeweed Mitogens; T-Lymphocytes; T-Lymphocytes, Regulatory; Vitamin B 12

A nephelometric assay of concanavalin-A binding of serum acute phase proteins (con-A binding) has been used in cross-sectional and sequential studies of disease activity in rheumatoid arthritis (RA). Con-A binding correlated well with blood viscosity, C-reactive protein, and other individual acute phase reactants in patients with active RA. Twenty-four patients were treated for 6 months with D-penicillamine and assessed clinically and seriologically. Clinical improvement was accompanied by significant falls in both C-reactive protein and con-A binding, although the serological changes did not always occur in parallel in individual patients. The advantages of this simple, cheap assay of acute phase proteins are discussed.

Topics: Adolescent; Adult; Aged; Arthritis, Rheumatoid; Blood Viscosity; C-Reactive Protein; Concanavalin A; Female; Humans; Male; Middle Aged; Penicillamine; Protein Binding

Topics: Arthritis, Rheumatoid; Cell Migration Inhibition; Collagen; Concanavalin A; Fibroblasts; Humans; Lung; Lymphocyte Activation; Lymphocytes; Lymphokines

Topics: Arachidonic Acids; Arthritis, Rheumatoid; Cell Separation; Concanavalin A; Female; Humans; Lymphocyte Activation; Lymphocytes; Male; Monocytes; Prostaglandins; Prostaglandins E; Prostaglandins F; Thromboxane B2

Whereas the two isomers of histidine equally enhanced the dextran and Con A anaphylactoid reactions in rats, their copper complexes were potent inhibitors of these reactions. On the other hand, the two isomers of cystine and their copper complexes inhibited the oedema reactions in rat paws induced by dextran, carrageenan and concanavalin A.

Topics: Amino Acids; Animals; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Carrageenan; Ceruloplasmin; Concanavalin A; Copper; Cystine; Dextrans; Histamine Release; Histidine; Isomerism; Rats; Tryptophan

Concanavalin A-induced suppressor cell activity was found moderately but significantly (P less than 0.05) decreased in RA patients treated with nonsteroidal antiinflammatory drugs (31 +/- 7% suppression) as compared to patients on remission-inducing drugs, such as gold, penicillamine, or chloroquine (51 +/- 6%) or to healthy individuals (50 +/- 6%). Also, lymphocytes from patients with antibodies to collagen mediated lower suppression (33 +/- 7%) than lymphocytes from patients without evidence for these autoantibodies (61 +/- 11%). No significant difference between patients and controls or between individual groups of patients were observed in regard to IgM and IgG secretion induced by pokeweed mitogen. Thus, although no indication for a severe derangement of regulatory cells in peripheral blood of RA patients could be observed in this study, a slight deficiency of ConA-inducible suppressor cells that may be reverted by remission-inducing drugs seems to be present in RA.

Topics: Adult; Aged; Arthritis, Rheumatoid; Concanavalin A; Female; HLA Antigens; Humans; Immunoglobulin G; Immunoglobulin M; Immunoglobulins; In Vitro Techniques; Male; Middle Aged; T-Lymphocytes, Regulatory

Lysosome proteins derived from peripheral blood granulocytes of patients with multiple sclerosis (MS), lupus erythematosus (LE), rheumatoid arthritis (RA) and recurrent uveitis cause the impairment of Con A-induced suppressor cell activity in two-step culture in vitro. This effect is independent of lysosome protease activity. Such a phenomenon, observed in vitro, may cause disturbances in suppressor cell function in the course of the above diseases.

Topics: Adult; Arthritis, Rheumatoid; Autoimmune Diseases; Concanavalin A; Female; Humans; Isoflurophate; Lupus Erythematosus, Systemic; Male; Middle Aged; Multiple Sclerosis; Neutrophils; Proteins; Recurrence; T-Lymphocytes, Regulatory; Uveitis

Suppressor cell activity was investigated in peripheral blood lymphocytes from twenty patients with rheumatoid arthritis (RA) and twenty patients with juvenile rheumatoid arthritis (JRA) using a concanavalin A/mixed lymphocyte culture assay. The mean suppression in the RA patients was slightly reduced compared with the suppressor cell activity in adult controls (25 +/- 5% suppression compared with 37 +/- 5%; P less than 0.05, Student's t test), whereas the JRA patients had normal suppressor cell activity (mean 46 +/- 5% versus 43 +/- 5% in healthy children matched for age and sex). The RA patients had normal proportions of T-cell subpopulations, 13.3% T gamma cells and 49.8% T mu cells, compared with 13.8% and 58.0% in controls. The JRA patients, however, had a significantly reduced mean percentage of T gamma cells, 6.6%, compared with 13.8% in healthy children (P less than 0.05, Mann-Whitney U-test). The mean percentage of T mu cells was 53.7%, versus 56.2% in the controls. The relation between suppressor cell activity and suppressor cells enumerated by membrane markers is discussed.

Topics: Adult; Arthritis, Juvenile; Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Humans; Immunoglobulin kappa-Chains; Immunoglobulin lambda-Chains; Lymphocyte Culture Test, Mixed; T-Lymphocytes; T-Lymphocytes, Regulatory

The mitogenic response of peripheral blood lymphocytes from 21 patients with rheumatoid arthritis to concanavalin-A (con--A), phytohaemagglutinin (PHA), and pokeweed mitogen (PWM) was significantly lower than in 30 normal subjects. After 15--24 weeks' treatment with sodium aurothiomalate (GST) the response to these mitogens rose to within the normal range. Improvement over pretreatment values was significant for con-A and PWM measured as area under the dose response curve but only for con--A if response at optimal mitogen concentration is the sole criterion. The improvement in PHA response was not significant with either method of measurement. There was an improvement in disease activity by 15--24 weeks as measured by a fall in serum C-reactive protein (CRP), IgM rheumatoid factor (RF), Clq binding activity (ClqBA), and Ritchie articular index. Con--A lymphocyte responsiveness was inversely related to serum CRP levels, but measurements of disease activity were otherwise unrelated to lymphocyte mitogen responsiveness. The observed improvement in peripheral blood lymphocyte responsiveness during gold treatment contrasts with the suppressive effect of gold in vitro. We suggest that the improvement in lymphocyte function is due to the lessening of rheumatoid disease activity during gold treatment, and that the low serum gold levels in our patients were insufficient to mask this effect. Sera from some of our patients were capable of suppressing the function of normal lymphocytes, and this was less apparent after treatment. The suppressive effect of sera correlated with ClqBA. Suppressive factors in serum, including possibly immune complexes, may be one factor leading to suppression of lymphocyte function during rheumatoid arthritis. Such an inverse relationship between humoral and cellular immune mechanisms might influence the clinical expression of rheumatoid arthritis.

Topics: Arthritis, Rheumatoid; Complement C1; Concanavalin A; Female; Gold; Gold Sodium Thiomalate; Humans; Immunity, Cellular; Lymphocyte Activation; Male; Phytohemagglutinins; Pokeweed Mitogens

We studied T cell subpopulations and their immunoregulatory circuits in the peripheral blood of 16 patients with rheumatoid arthritis (RA) who were receiving no medications that might interfere with the results. We found normal T cells with receptors for the Fc portion of IgG or IgM as well as autologous rosette-forming T cells (Tar cells), a subpopulation of T cells we have found to have the properties of human post-thymic precursors. We also found that peripheral blood cells of RA patients have normal concanavalin A-induced or spontaneously-expanded suppressor cell functions. Also normal were the characteristic functions of the Tar cells; feedback inhibition and the generation of suppression. The normal state of these T cell subpopulations and immunoregulatory circuits in the peripheral blood of patients with RA contrasts with their various abnormalities in other connective tissue diseases. This may either mean that the immunoregulatory aberration in RA involves primarily B cells, or, if it involves T cells, that it does so primarily in the synovial membrane.

Topics: Arthritis, Rheumatoid; Concanavalin A; Feedback; Humans; Rosette Formation; T-Lymphocytes

Topics: Animals; Antibodies; Antigens, Surface; Antilymphocyte Serum; Arthritis, Rheumatoid; Complement System Proteins; Concanavalin A; Humans; Immunoglobulin G; Immunoglobulin M; Lymphocyte Activation; Pokeweed Mitogens; Rabbits; Time Factors

The present study was designed to characterize leukocytes of patients with rheumatoid arthritis (RA) with regard to proliferation of peripheral blood lymphocytes (PBL) and prostanoid release from circulating monocytes (M phi. Compared to cells of healthy individuals, PBL from RA patients exhibited a reduced mitogenic response to concanavalin A (Con A) which was associated with an increased capacity of circulating M phi to synthesize PGE and thromboxane B2 (TXB2). Addition of synovial fluid exudates of RA patients (RA-SFE) to peripheral blood leucocyte cultures produced three effects: A spontaneous proliferation of normal and RA-PBL, a reduction of the Con A response of normal and RA-PBL, a reduction of the Con A response of normal and RA-PBL, and an enhanced release of PGE and TXB2 from RA-M phi only. To elucidate the cellular origin of these activities, normal and RA-PBL were incubated with supernatants (SNT) os synovial cell cultures from RA patients and patients with non-RA joint diseases. SNT from Con A-stimulated synovial lymphocytes of both RA and control patients induced a spontaneous proliferation of normal and RA-PBL. In contrast, SNT from non-lymphoid adherent synovial cells of RA and control patients reduced the Con A response of normal and RA-PBL but a striking difference was noted in that an enhanced PGE and TXB2 release occurred only from M phi of RA patients.

Topics: Arthritis, Rheumatoid; Cell Division; Concanavalin A; Exudates and Transudates; Humans; In Vitro Techniques; Lymphocytes; Mitogens; Prostaglandins; Prostaglandins E; Synovial Fluid; Thromboxane B2; Thymidine

Adherent synovial cells (ASC) were obtained from minced synovium from patients with rheumatoid arthritis by dissociating the lining cells from the extracellular matrix and dispersing them with proteases. These cells produce high levels of latent collagenase in primary culture. With passage of ASC, collagenase levels decrease but can be stimulated by a soluble factor (MCF) released in vitro from cultured human blood monocyte-macrophages. Monocyte cultures exposed to aggregated immunoglobulin, Fc fragments of immunoglobulin or concanavalin A (Con A) increase production of MCF several-fold more than the control cultures. MCF production by monocytes exposed to Fab fragments does not differ from controls. Although aggregated immunoglobulin, Fc fragments, and Con A stimulate PGE2 synthesis and secretion by human monocytes, the effects on MCF production are not mediated by PGE2 since concentrations of indomethacin that completely block PGE2 production do not inhibit MCF production. These findings of increased production of MCF by monocytes in response to substances that probably exert their effects via surface receptors could be relevant in interpreting the in vivo role of immune complexes in diseases associated with connective tissue destruction.

Topics: Arthritis, Rheumatoid; Cell Adhesion; Cells, Cultured; Concanavalin A; Humans; Immunoglobulin Fab Fragments; Immunoglobulin Fc Fragments; Immunoglobulin G; Lectins; Macrophages; Microbial Collagenase; Monocytes; Synovial Membrane

Peripheral blood lymphocyte mitogen responsiveness was studied in 21 patients with rheumatoid arthritis being treated with sodium aurothiomalate. There was a significant increase in lymphocyte response to concanavalin A and pokeweed mitogen but not to phytohaemagglutinin. This observed increase in lymphocyte response contrasts with the suppressive effect of gold salts in vitro. We propose that this apparent contradiction may be explained by the relatively low serum gold levels measured in our patients, compared with expected levels in synovial membrane. Thus gold could suppress rheumatoid inflammation in the "target tissue" while having little suppressive action in the peripheral blood compartment, where a removal of suppressive influences due to active disease might then be seen as a net improvement in lymphocyte responsiveness.

Topics: Adult; Arthritis, Rheumatoid; Concanavalin A; Female; Gold Sodium Thiomalate; Humans; In Vitro Techniques; Lymphocyte Activation; Male; Middle Aged; Pokeweed Mitogens

We have studied the primary in vitro antibody response toward a hapten in cultures of peripheral blood lymphocytes from twenty-two patients suffering from regular rheumatoid arthritis (RA). These patients were not receiving immunosuppressive drugs or corticosteroids and had not taken Aspirin or non-steroidal anti-inflammatory agents for at least 72 hr. The control groups included thirty-two healthy subjects and twenty-seven control patients. The mean anti-TNP response of the RA patients was significantly lower than that of both control groups. No pre-existing anti-TNP or IgG response could be detected. A search for suppressor cells in co-cultures of RA and normal lymphocytes was negative. On the contrary, the extent of allogeneic enhancement in such co-cultures was comparable to that observed when control lymphocytes were co-cultured. RA serum added to normal lymphocytes cultures showed a dramatic inhibitory effect in only two out of nine cases. A follow-up study has strongly suggested that RA lymphocytes could increase their in vitro antibody response upon treatment.

Topics: Adult; Antibody Formation; Antibody-Producing Cells; Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Humans; Lymphocytes; Male; Middle Aged; T-Lymphocytes, Regulatory; Trinitrobenzenes

The suppressor activity of mononuclear cells of the peripheral blood (MCPB) during rheumatoid polyarthritis (RP) was studied using experimental protocole. The MCPB stimulated in vitro by concanavaline A (Con A) are capable of suppressing the mitogenis response of autologus cells; moreover, the short-lived spontaneous suppressor cells disappear during 24 hour in vitro incubation that determines an increase in the proliferative response of the incubated cells for 24 hours. In two of the six RP studied, the suppressor activity generated Con A and the spontaneous suppressor activity are nul. Culture experiments with the MCPB of PR and control subjects show that this defect in suppressor activity is more related to a problem in the generation of suppressor cells than to a deficiency in the response to suppressor signals.

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Lymphocyte Activation; T-Lymphocytes, Regulatory

We have studied the in vitro antibody response to a hapten of peripheral blood lymphocytes from 26 patients with rheumatoid arthritis and 7 ankylosing spondylitis. These patients had never received immunosuppressor drugs before or corticosteroids during the month before the test. They had failed to receive aspirin or non-steroid anti-inflammatory drugs for 72 hours before blood sampling. The control groups included respectively 38 healthy subjects and 24 patients hospitalized for non inflammatory disease. The antibody response of ankylosing spondylitis patients is comparable to that of controls ; on the opposite the response of patients with rhumatoid arthritis is significantly depressed in comparison with the three other groups. The weak response of lymphocytes in arthritis is not due to increased cell death in culture or to modified kinetics of the antibody response or to the appearance of a IgG secondary type response or a in vivo pre-activation. The lymphocytes of arthritis patients do not inhibit the response of normal lymphocytes when they are co-cultured. The observed response is identical to that obtained when control patient lymphocytes are co-cultured with normal lymphocytes. The function of suppressor T cells induced by Con A seems normal in spondylitis and arthritis.

Topics: Adult; Aged; Antibody Formation; Arthritis, Rheumatoid; Cells, Cultured; Concanavalin A; Female; Humans; Lymphocytes; Male; Middle Aged; Spondylitis, Ankylosing; T-Lymphocytes, Regulatory; Trinitrobenzenes

Synovial lymphocytes eluted by enzyme treatment from eleven patients with rheumatoid arthritis (RA) were investigated for the presence of concanavalin A (Con A)-activated suppressor cell activity as compared with that of peripheral blood lymphocytes of twenty normal donors. In addition, two patients with psoriatic arthritis and juvenile rheumatoid arthritis (JRA) were also investigated. Synovial lymphocytes from the eleven RA patients showed a mean augmentation of 28 +/- 13.30, and thus clearly lacked suppressor activity, whereas the mean suppression in the lymphocytes from the twenty normal donors was 13 +/- 14.40. Synovial lymphocytes from one patient with JRA and one with psoriatic arthritis showed a normal suppressor activity.

Topics: Arthritis, Juvenile; Arthritis, Rheumatoid; Concanavalin A; Humans; Lymphocyte Activation; Lymphocytes; Synovial Membrane; T-Lymphocytes, Regulatory

The response of peripheral blood and synovial fluid lymphocytes to three non-specific mitogens has been studied. The paired samples were taken from patients with a range of inflammatory arthritides. Unstimulated synovial fluid lymphocytes (SFL) tended to have a greater uptake of tritiated thymidine than had unstimulated peripheral blood lymphocytes (PBL). This background uptake of tritiated thymidine by SFL showed a positive correlation with the response these SFL then showed to the mitogens. A significant depression was observed in the SFL response to phytohaemagglutinin when compared with the paired PBL response; this was seen in both the rheumatoid arthritis and other inflammatory joint diseases groups. SFL responses to concanavalin A and pokeweed mitogen, although depressed in individual cases, failed to show a significant depression overall. Attempts to restore the SFL response to that of the paired PBL by removal of any possible blocking substance from the cell surface either by pre-incubation of SFL in tissue culture medium or by enzyme treatment were unsuccessful. This suggested that cell surface blockers were possibly not the reason for deficient SFL reponses and that other factors were involved.

Topics: Arthritis, Rheumatoid; Cells, Cultured; Complement C3; Concanavalin A; Humans; Hyaluronoglucosaminidase; Lymphocyte Activation; Neuraminidase; Phytohemagglutinins; Pokeweed Mitogens; Synovial Fluid; Trypsin

The effect of clotrimazole, an imidazole derivative with anti-rheumatic properties, on lymphocyte stimulation by phytohaemagglutinin, concanavalin-A and pokeweed mitogen was investigated in an in vitro culture system. Evidence was obtained to show that the drug could either enhance or inhibit lymphocyte stimulation, the response depending on the concentration of the drug in the culture system and the mitogen used, as well as on individual variability. At a concentration of the drug corresponding to therapeutic serum levels, clotrimazole inhibited lymphocyte response to the three mitogens in all the normal volunteers studied. In addition, the effect of clotrimazole on in vitro mitogenic responsiveness of lymphocytes of a group of patients with rheumatoid arthritis taking this drug was compared to that of the proprionic acid derivative, ketoprofen. Patients taking clotrimazole showed a significant reduction in lymphocyte responsiveness, when compared to pre- and post-treatment levels, whereas there was no significant difference in those patients taking ketoprofen. Although cortisol levels tended to be higher in the groups of patients taking clotrimazole there was no correlation between lymphocyte responsiveness and cortisol concentration.

Topics: Arthritis, Rheumatoid; Clotrimazole; Concanavalin A; Humans; Imidazoles; In Vitro Techniques; Ketoprofen; Lectins; Lymphocyte Activation; Phytohemagglutinins; Pokeweed Mitogens

Lymphocytes were highly purified from synovial fluid and peripheral blood of 10 rheumatoid arthritis patients and assessed for responsiveness to PHA-P and Con-A. In all cases, both synovial and blood lymphocytes showed a marked reduction in response to these mitogens compared with normal blood lymphocytes. The factors responsible for this low T cell responsiveness are discussed.

Topics: Arthritis, Rheumatoid; Chronic Disease; Concanavalin A; Humans; Lymphocyte Activation; Phytohemagglutinins; Synovial Fluid

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Lectins; Lymphocyte Activation; Lymphocytes; Lymphotoxin-alpha; Synovial Fluid

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

Mitogenic transformation of lymphocytes from patients with rheumatoid arthritis (RA) has revealed divergent results in different laboratories. Since the sera used for medium supplementation in these previous studies might have influenced the results, we investigated lymphocyte transformation in serum-free medium in 28 RA-patients and 25 controls. It was shown that a majority of patients with RA responded weakly to PHA. Comparing some of these serum-free cultures with cultures that were set in parallel in 10% ABserum, no difference between patients and controls was observed after transformation in supplemented medium. It is concluded that lymphocyte reactivity to PHA (however not to ConA and PWM) is diminished in patients with RA and that this defect--at least partially--is reversible by addition of serum. The use of parallel cultures in serum-free and serum-containing medium is recommended for evaluation of the lymphocyte mitogenic response.

Topics: Adult; Aged; Arthritis, Rheumatoid; Blood; Concanavalin A; Culture Media; Humans; Lymphocyte Activation; Middle Aged; Mitogens; Phytohemagglutinins

Topics: Arthritis, Rheumatoid; Concanavalin A; Humans; Lectins; Lymphocyte Activation; Synovial Fluid

The criteria for selecting and establishing a animal model for arthritis were described. Rats are the most frequently used animals. Adjuvant and Myocobacterium induced arthritis provide a model of chronic joint inflammation, although significant differences exist when compared with human disease. A better model of arthritis in rats and mice can be induced by the injection of one strain of Mycoplasma arthritidis. An even better model is presented by rabbits first immunized against a protein such as heterologous fibrin or albumin, and then challenged by the same protein injected directly into a joint. This results in a localized chronic arthritis pathologically similar to that of man. Arithritis can also be induced in rabbits by the injection of polymers such as chitinor or Concanavalin A into the joint. Although there is no lack of arthritis animal models, there is no animal model which gives a true replication of rheumatoid arthritis.

Topics: Animals; Arthritis; Arthritis, Infectious; Arthritis, Rheumatoid; Concanavalin A; Disease Models, Animal; Dog Diseases; Dogs; Haplorhini; Mice; Mycobacterium; Mycobacterium tuberculosis; Mycoplasma Infections; Rabbits; Rats; Rodent Diseases

The influence of synovial fluid from four patients with Reiter's syndrome on lymphocyte responsiveness was studied. On synovial fluid was found which specifically depressed the responsiveness of lymphocytes from patients with Reiter's syndrome to the non-specific mitogen phytohaemagglutinin (PHA). The ratio of the responsiveness of lymphocytes cultured in the presence of foetal calf serum (FCS), compared to those incubated in the Reiter's synovial fluid, was used as a measure of the depression induced by the Reiter's synovial fluid. The mean ratio for eight normals stimulated with PHA was 0-70 (range 0-35-0-96), while for eight patients with Reiter's syndrome, it was 0-13 (range 0-07-0-19). Similar studies done with concanavalin A (con A) showed no difference between lymphocytes from normals (0-73) or patients with Reiter's syndrome (0-67). Chromatography of the Reiter's synovial fluid on a Sepharose 4-B column resulted in the separation of three major fractions, one of which exhibited the inhibitory activity. When this active fraction was absorbed with Reiter's lymphocytes, a loss of the inhibitory activity of the fraction was seen. A similar absorption with normal lymphocytes had no effect. These studies demonstrate that a factor present in the synovial fluid of a patient with Reiter's syndrome reacted specifically with lymphocytes from patients with Reiter's disease and not with lymphocytes from normals. The interaction of this factor with lymphocytes from patients with Reiter's syndrome inhibited the responsiveness of these lymphocytes to PHA but not to con A.

Topics: Adult; Arthritis, Reactive; Arthritis, Rheumatoid; Chromatography; Concanavalin A; Humans; Lectins; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Synovial Fluid

Concanavalin A (Con A) froms precipitates with carbohydrate-rich protein such as IgM, IgD, IgE, and IgA. Since IgG contains little carbohydrate and does not react with Con A, the activity of IgG-rheumatoid factor (RF) can be measured in the supernate of the Con A-treated serum. When the latex fixation test (LFT) and the sensitized sheep cell agglutination test (SSCA) were perfromed in the supernate for the detection of IgG-RF, LFT was positive in 32-1% of sera, out of 137 sera originally positive for LFT, and SSCA was positive in 18-5% of sera, out of 119 sera originally positive for SSCA. IgG-RF exhibited lower complement fixing ability than IgM-RF and correlated with agglutination titres of IgG-RF, while the CH50 of the original serum did not correlate with haemolytic activities of either IgM-RF or IgG-RF.

Topics: Arthritis, Rheumatoid; Complement Fixation Tests; Concanavalin A; Humans; Immunoglobulin A; Immunoglobulin G; Immunoglobulin M; Methylglucosides; Rheumatoid Factor

Topics: Adolescent; Adult; Arthritis, Rheumatoid; Autoimmune Diseases; Concanavalin A; Humans; Lectins; Lupus Erythematosus, Systemic; Lymphocytes; Middle Aged

Lymphocytes were eluted from synovial tissues of 17 patients with classical rheumatoid arthritis, using a procedure previously reported. Stimulation was obtained with the unspecific mitogens phytohemagglutinin (PHA), pokeweed mitogen (PWM), and concanavalin A (Con A) as well as with purified protein derivative of tuberculin (PPD) and mitoycin-C-treated allogeneic lymphocytes, whereas candida antigen usually gave a low response. The pattern of reactivity to unspecific mitogens was similar to that obtained with lymphocytes from peripheral blood of rheumatoid arthritis patients. Two different PPD preparations usually gave transformation of the same magnitude as seen with PHA. This was in contrast to the reactivity of the peripheral blood lymphocytes. It could be demonstrated that the elution procedure initiated some degree of lymphocyte transformation, mainly potentiating the responses to PHA and Con A.

Topics: Antigens; Arthritis, Rheumatoid; Concanavalin A; Humans; Immunoglobulin Fc Fragments; Lectins; Lymphocyte Activation; Lymphocytes; Mitogens; Mitomycins; Receptors, Antigen, B-Cell; Synovial Membrane; Tuberculin

Daily intraperitoneal doses of concanavalin A (Con A) produced a dose-related inhibition of adjuvant-induced arthritis in rats. Con A was also effective on established arthritis, markedly relieving the disease after only three doses. The inhibitory effect of Con A was neutralised by pre-incubation with ovalbumin, although this treatment did not modify the delayed phlogistic action of Con A in rat paws.

Topics: Animals; Arthritis, Rheumatoid; Concanavalin A; Dose-Response Relationship, Drug; Edema; Female; Foot; Hindlimb; Immunosuppressive Agents; Ovalbumin; Rats; Time Factors

Patients with rheumatoid arthritis (RA), 36 cases, and normal subjects, 49 cases were studied by lymphocyte cultures stimulated by phytohemagglutinin (PHA), Concanavalin A (Con A), pokeweed mitogen (PWM) and Con A convalently bound to Sepharose 4 B (Con A-S). The comparisons between the two groups showed a significant difference between the RA lymphocytes and the control lymphocytes stimulated by PHA and Con A. However, no statistical difference was found between the two lymphocyte populations stimulated by PWM and Con A-S. In order to determine the lymphocyte population stimulated by each mitogen, separation of B and T cells from peripheral blood was performed according to the ability for the T cell population to bind the sheep red blood cells (rosette-forming cells). The T cell-rich population was only stimulated by PHA, Con A and PWM. Although the T cell-depleted population showed no response to these mitogens, a response to Con A-S was elicited.

Topics: Adult; Aged; Arthritis, Rheumatoid; B-Lymphocytes; Cell Separation; Concanavalin A; Humans; Immune Adherence Reaction; Lectins; Lymphocyte Activation; Middle Aged; Mitogens; T-Lymphocytes

Topics: Arthritis, Rheumatoid; Binding Sites, Antibody; Chloramines; Concanavalin A; Dose-Response Relationship, Drug; Epitopes; Iodine Radioisotopes; Lectins; Lymphocytes

Topics: Arthritis, Rheumatoid; B-Lymphocytes; Cell Separation; Concanavalin A; Humans; Immune Adherence Reaction; Lectins; Lipopolysaccharides; Lymphocyte Activation; Lymphocytes; Mitogens; T-Lymphocytes; Thymidine; Tritium