A-stimulated Mononuclear Cells Research Articles

Inhibitory influences of xanthine oxidase inhibitor and angiotensin I-converting enzyme inhibitor on multinucleated giant cell formation from monocytes by downregulation of adhesion molecules and purinergic receptors.

Allopurinol, a xanthine oxidase inhibitor, and captopril, an inhibitor of angiotensin I-converting enzyme, are widely used for hyperuricaemia and hypertension, respectively. There have been reported cases showing that these two agents are effective for the treatment of granulomatous diseases such as sarcoidosis, although the mode of action is not elucidated. We examined the in vitro effects of these agents on the formation of multinucleated giant cells (MGC) from human monocytes by concanavalin A-stimulated mononuclear cell supernatants (conditioned medium). We cultured monocytes with conditioned medium and each agent and compared the rate of MGC formation as well as the expression of adhesion molecules and P2X7 receptor, which are involved in MGC formation. The addition of 25 or 100 microg mL(-1) allopurinol or 0.125-1.0 microg mL(-1) captopril inhibited MGC formation. Monocytes treated with these agents exhibited less expression of intercellular adhesion molecular-1 (ICAM-1) than untreated monocytes. The susceptibility of monocytes cultured in conditioned medium for 24 h to 2'-and 3'-o-(4-benzoyl-benzoyl)adenosine triphosphate-mediated cytolysis was significantly lower in monocytes treated with these agents than in untreated monocytes. Allopurinol and captopril have a therapeutic effect on granulomatous disorders by a direct action on monocyte/macrophage lineage cells partly through downregulation of ICAM-1 and P2X7 receptor.

Inhibitory influences of tranilast on multinucleated giant cell formation from monocytes by supernatant of concanavalin A-stimulated mononuclear cells

Tranilast is an anti-allergic drug that inhibits the release of chemical mediators from mast cells. There have been cases-reports showing that tranilast is effective for the treatment of granulomatous diseases such as granuloma annulare and cutaneous sarcoidosis. Here we examined the in vitro effects of tranilast on the formation of multinucleated giant cells (MGCs) from human peripheral monocytes. Supernatant of concanavalin A (Con A)-stimulated mononuclear cells induced Langhans-type and foreign body-type MGCs and the addition of 10 or 100 microg/ml tranilast inhibited the formation of total MGCs and foreign body-type MGCs. Tranilast decreased the number of MGCs with 16<nuclei and increased that of MGCs with three to five nuclei. Fluorescence-activated cell sorting analysis showed that tranilast-treated monocytes had lower expressions of intercellular adhesion molecule-1 (ICAM-1). These findings suggest that tranilast is effective for cutaneous lesions in some cases of granulomatous disorders partly through a direct effect on monocyte/macrophage-lineage cells.

Effects of propofol emulsion and thiopentone on T helper cell type-1/type-2 balance in vitro.

We have earlier found increased percentages of T helper cells (CD4-positive lymphocytes) in the blood circulation after propofol infusion anaesthesia. Cytokines interferon-gamma (IFN gamma) and interleukin-4 (IL-4) are important in the differentiation of T helper cells into subtypes T helper type-1 (Th1) and type-2 (Th2). To study the effects of propofol emulsion, its solvent Intralipid and thiopentone on Th1/Th2 balance, measurements of IFN gamma and IL-4 production by mononuclear leucocytes were carried out in vitro. As IL-2 has a central role in immune responses to surgery, its production was also measured. Concanavalin A-stimulated mononuclear cells were cultured in the presence of propofol emulsion at 3.5 or 10 micrograms.ml-1, Intralipid 35 or 100 micrograms.ml-1, or thiopentone 3 micrograms.ml-1. Cytokine production was measured from the conditioned media of mononuclear cell cultures. Decreased IFN gamma (p < 0.001) and IL-4 concentrations (p < 0.01) were found in the presence of thiopentone, but IL-2 production was unaffected. By contrast, propofol emulsion or Intralipid had no effects on IFN gamma, IL-2 or IL-4 concentrations. Propofol 10 micrograms.ml-1 increased the IFN gamma/IL-4 ratio from the control value median 243 (162-562) (25th-75th percentile) to 363 (195-1028) (p < 0.01), but thiopentone decreased it to 145 (60-214) (p < 0.01). These findings show that propofol and thiopentone have different effects in vitro on Th1/Th2 balance and suggest that they have different modulating effects on the immune response.

Cytokine profile in systemic lupus erythematosus, rheumatoid arthritis, and other rheumatic diseases.

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.

Inhibition of bovine peripheral blood mononuclear cell blastogenesis by fractionated mammary secretion.

1. Bovine mammary secretion whey obtained during late involution markedly inhibited mitogen-induced blood mononuclear cell blastogenesis. 2. Whey proteins eluting in the first and second absorbance peaks following molecular exclusion chromatography were associated with greatest inhibition of mononuclear cell blastogenesis. 3. Greatest inhibition of concanavalin A-stimulated mononuclear cell blastogenesis was associated with high concentrations of whey proteins in absorbance peak 1. 4. Whole mammary secretion whey and whey proteins in absorbance peak 2 caused similar inhibition of concanavalin A- and phytohaemagglutinin-treated mononuclear cells. 5. Differential inhibition of mitogen-induced blastogenesis may reflect the presence of immunosuppressive substances in bovine mammary secretion whey which differ in specificity for bovine T-cell subsets.

Separation and purification of lymphocyte chemotactic factor (LCF) and interleukin 2 produced by human peripheral blood mononuclear cells

Two T-cell chemotactic factors, lymphocyte chemotactic factor (LCF) and interleukin 2 (IL-2), were separated and characterized from culture supernatants of concanavalin A-stimulated human peripheral blood mononuclear cells. LCF was purified approximately 7800-fold to homogeneity from culture supernatant using gel filtration and high-performance liquid chromatography (HPLC). LCF was found to be distinct from both IL-2 and interleukin-1. Sephadex G-100 gel filtration of crude supernatants from concanavalin A-stimulated mononuclear cells showed two molecular weight regions of T lymphocyte chemotactic activity. A 10,000- to 25,000-Da region contained both IL-2 and LCF and a 45,000- to 75,000-Da region contained only a high molecular weight form of LCF. Both high and low molecular weight species of LCF eluted with 40-44% acetonitrile from a reversed-phase C18 HPLC column. IL-2 present only in the low molecular weight region eluted from the C18 column with 65-75% acetonitrile. The migration of T lymphocytes to IL-2 was totally inhibited by anti-interleukin 2 receptor antibody while the response of T cells to LCF was unaffected. LCF eluting off the C18 column was purified to homogeneity by two subsequent cycles of gel filtration HPLC. The resultant protein showed a single band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis corresponding to a molecular weight of 10,500. The data presented here demonstrate that IL-2 and LCF are distinct lymphocyte chemotactic factors and although they are not readily separable from crude supernatants by molecular sieve chromatography, they can easily be distinguished by reversed-phase HPLC.

Opioid peptides modulate production of interferon γ by human mononuclear cells

The opioid neuropeptides have previously been shown to bind to and affect leukocyte function including lymphocyte proliferation, NK-cell activity, mononuclear cell chemotaxis, immunoglobulin synthesis, and lymphokine production. The effect of the opioid peptides beta-endorphin and Met-enkephalin on interferon gamma (IFN) production by concanavalin A-stimulated human mononuclear cells was examined. Both beta-endorphin and Met-enkephalin enhanced IFN production by the majority of donor mononuclear cells tested and did so at concentrations between 10(-14) and 10(-10) M. When 10(-12) M beta-endorphin or Met-enkephalin were included in concanavalin A-stimulated mononuclear cell cultures, IFN concentrations were significantly enhanced to 205 +/- 45 and 252 +/- 67% of control, respectively. Although the majority of cell preparations tested exhibited an enhanced production of IFN in response to these opioid peptides, some did not. When beta-endorphin or Met-enkephalin were utilized at 10(-11) M, 10 of 15 and 7 of 11 responded with IFN production greater than 20% above the control (untreated) level. There was not an absolute correlation between an enhanced response to beta-endorphin and Met-enkephalin, suggesting the presence of multiple receptor types on these cells for opioids. The opioid receptor antagonist, naloxone, did not significantly prevent the opiate effect. When 10(-8) M naloxone was included in cultures containing 10(-12) M beta-endorphin or Met-enkephalin no significant inhibition of the effect of either opioid on IFN production was observed.

The effect of concanavalin A-stimulated mononuclear cells on low density lipoprotein receptor activity of cultured fibroblasts.

Secretory products of freshly isolated human circulating blood cells such as platelets, monocytes, and B lymphocytes, but not T lymphocytes, have previously been shown to enhance low density lipoprotein (LDL) metabolism by arterial wall cells. This study was undertaken to evaluate how secretory factor(s) from mononuclear cells that had been stimulated by concanavalin A (Con A) alters LDL receptor activity by cultured human skin fibroblasts. Conditioned medium from Con A-stimulated mononuclear cells produced an increase of 125I-LDL degradation accompanied by increased thymidine incorporation into DNA. The effect of conditioned medium from the Con A-stimulated mononuclear cells was mediated by the LDL receptor pathway. Degradation of HDL and methylated LDL, neither of which is taken up by the classical LDL receptor pathway, was not affected. The conditioned medium from these Con A-stimulated cells also failed to stimulate fluid pinocytosis, as measured by the uptake of [14C]sucrose. Some strains of fibroblasts, deficient in LDL receptors, responded to the conditioned medium from the Con A-stimulated mononuclear cells by increasing the very small amounts of LDL degraded by these cells. Fibroblasts from other homozygous familial hypercholesterolemic cell strains were unresponsive, however. The effect on LDL receptors was characterized by an increase in LDL receptor number without a change in the affinity of LDL for its receptor. Thus stimulated mononuclear cells secrete mitogens that also stimulate LDL receptor activity in human skin fibroblasts.

In vitro production and scavenging of hydrogen peroxide by D-penicillamine. Relationship to copper availability.

The capacity of D-penicillamine (DP) to produce or scavenge hydrogen peroxide was investigated. DP added to copper produced H2O2. Greater production was observed with copper sulfate than with copper bound to ceruloplasmin. In contrast, DP in the absence of copper scavenged H2O2, as measured in a direct assay. Furthermore, DP or D-cysteine alone reversed H2O2-mediated inhibition of concanavalin A-stimulated mononuclear cell proliferation. These opposing immunomodulating properties of DP may be relevant to its toxic or therapeutic actions in rheumatoid arthritis.

Migration of human helper/inducer T cells in response to supernatants from Con A-stimulated suppressor/cytotoxic T cells.

Previous studies have shown that supernatants from Con A-stimulated human mononuclear cells are chemotactic for T cells, and that the source of the activity is the Leu-2 (suppressor/cytotoxic) T cell. The specificity of this chemoattractant activity for Leu-2 and/or Leu-3 (helper/inducer) T cells was analyzed with isolated human T cell subsets in an in vitro chemotaxis assay system. These studies demonstrate that both human Leu-2 and Leu-3 cells migrate in response to the lymphocyte locomotor stimulus casein, whereas only Leu-3 cells migrate in response to supernatants from Con A-stimulated mononuclear cells. This migration of Leu-3 cells to the Con A supernatant was reflected in both the distance migrated and the number of migrating cells. When Leu-2 cell locomotion in response to Con A supernatants was analyzed, no significant differences from control were observed in either the distance migrated or the number of migrating cells. Supernatants from Con A-stimulated Leu-2 cells similarly attracted Leu-3 cells but not Leu-2 cells in a concentration-dependent manner. Supernatants from Con A-stimulated Leu-3 cells did not stimulate either Leu-2 or Leu-3 cell migration. These studies demonstrate that Leu-3 T cells migrate in response to supernatants from Con A-stimulated Leu-2 T cells. This specific response may promote cell-to-cell interaction by providing a means for Leu-2 T cells to recruit Leu-3 T cells.