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Biomedical subjects

G W Duff

Publications and source records attributed to G W Duff.

At least 91 records · Page 5Linked to original sources
PubMed1993

Cytokines and anti-cytokines.

The cause of RA is unknown but there are several hypotheses: an autoimmune mechanism; a superantigen-driven disease; an infection with an unusual pathogen (e.g. lentivirus); and a non-specific infectious stimulus acting on a particular genetic background. Whatever the cause(s) of RA, it would appear that cytokines are very important mediators of the pathology. Genetic polymorphisms with Mendelian inheritance have been characterized in regulatory regions of cytokines such as IL-1, TNF and related genes. It is possible, therefore, that whatever the initiating stimulus might be, the possession of different cytokine alleles may confer greater or lesser risk of developing a progressive disease.

Arthritis, Rheumatoid↗
PubMed1992

In situ hybridization of interleukin-1 in CD14-positive cells in rheumatoid arthritis.

Interleukin 1 (IL-1) has been implicated as an inflammatory mediator in rheumatoid arthritis (RA). Many cell types, including macrophages, lymphocytes, fibroblasts, and endothelial cells, can produce IL-1 and it is known that IL-1 production is under transcriptional control. It has, however, been difficult to define in vivo the predominant cellular source of this mediator in RA. Here, we have used the combination of in situ hybridization of mRNA and cellular immunophenotyping with monoclonal antibodies to show that the IL-1 beta gene is expressed predominantly by CD14-positive macrophages in synovial tissue from patients with RA. Synovial macrophages were also associated with the immunoreactive IL-1 peptide. These cells appear to be the major source of IL-1 beta within the rheumatoid synovium in vivo and must be regarded as playing a central role in the chronic inflammation and joint destruction of RA.

Antigens, CD↗
PubMed1992

Soluble interleukin-2 receptor and disease activity in Crohn's disease.

An ELISA was used to measure concentrations of soluble interleukin-2 receptor (sIL-2R) alpha chain in the sera of patients with Crohn's disease. In a group of 56 patients, serum concentrations of sIL-2R were significantly raised in patients with active disease compared with patients with inactive disease and age-matched control populations. There was a significant correlation between serum sIL-2R concentration and disease activity as assessed by the Harvey-Bradshaw index (r = +0.60; P less than 0.001) and laboratory measurements of disease activity including C-reactive protein (r = +0.79; P less than 0.001), ESR (r = +0.64; P less than 0.001) and platelet count (r = +0.533; P less than 0.001). We also found a negative correlation between sIL-2R levels and serum albumin (r = -0.66; P less than 0.001). In longitudinal studies, changes in the concentration of serum sIL-2R reflected the changes in disease activity. Soluble IL-2R, therefore, offers a new measure of disease activity in Crohn's disease with a potential advantage over other laboratory parameters currently available in that it may reflect more accurately the underlying immunopathogenic process.

Adrenal Cortex Hormones↗
PubMed1992

In situ hybridization of IL-6 in rheumatoid arthritis.

IL-6, an important mediator of the acute phase response, has been implicated in the pathogenesis of rheumatoid arthritis (RA). Many cell types including macrophages, T cells, B cells, endothelial cells and fibroblasts can produce this cytokine and production is largely regulated at the level of gene transcription or mRNA stabilization. In this paper we have first measured the levels of IL-6 activity in synovial fluid (SF) and serum from patients with RA and then localized IL-6-producing cells in the synovium by in situ hybridization combined with immunophenotyping. Patients with RA had raised levels of IL-6 in both SF and serum compared with patients with osteoarthritis and age-matched healthy controls. In individual RA patients tested serially after admission to hospital, serum IL-6 was initially raised and, unexpectedly, increased with clinical improvement. In situ hybridization of IL-6 mRNA showed positive cells both in the lymphocyte-rich aggregates and adjacent to small blood vessels. With immunophenotyping it was found that cells containing IL-6 mRNA were often in contact with CD14+ tissue macrophages and double immunophenotyping revealed that immunoreactive IL-6 was often associated with synovial T cells.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1991

Purification and characterization of a novel soluble receptor for interleukin 1.

Affinity chromatography and reverse-phase high-performance liquid chromatography was used to purify a soluble interleukin 1 beta (IL-1 beta) specific binding protein from the supernatant of a human B cell line, Raji. The purified protein specifically bound 125I IL-1 beta forming a 60-kD complex in nonreducing conditions and a 70-kD complex in reducing conditions. Binding was found to be displaceable by mature human and murine IL-1 beta and human 31-kD IL-1 beta propeptide, but not displaceable by human and murine IL-1 alpha or human IL-1 receptor (IL-1R) antagonist. Ligand blotting revealed a 47-kD molecule that specifically bound IL-1 beta. Measurement of binding affinity of the cell surface Raji IL-1R (Kd = 2.2 nm) and the Raji soluble (s)IL-1R (Kd = 2.7 nm) demonstrated a similar affinity for 125I IL-1 beta. Purified sIL-1R inhibited binding of IL-1 beta to cell lines with both type I (80 kD) and type II (65 kD) IL-1Rs, but did not interfere with IL-1 alpha binding. This natural sIL-1R may function as an important regulatory molecule of IL-1 beta in vivo.

Cell Line↗
PubMed1991

Failure to detect paramyxovirus sequences in Paget's disease of bone using the polymerase chain reaction.

It is widely believed that Paget's disease of bone is due to a "slow virus" infection of osteoclasts with one of the paramyxovirus group. Controversy continues to surround the identity of the virus involved, however, since at different times evidence has been presented implicating measles virus (MV), respiratory syncytial virus (RSV), and canine distemper virus (CDV) as putative infective agents. In this study we used the technique of reverse transcription and polymerase chain reaction (PCR) to screen for paramyxovirus sequences in ribonucleic acid (RNA) extracted from pagetic bone. We were able to detect viral amplification products of the appropriate size in RNA extracted from as few as 50 cells experimentally infected with a wide range of paramyxoviruses, including measles, canine distemper, parainfluenza 3, and respiratory syncytial virus, but we found no evidence of viral products in RNA extracts of affected bone from 10 consecutive patients with Paget's disease. This study fails to support the hypothesis that active infection with one of these or a related paramyxovirus is involved in the pathogenesis of Paget's disease.

Aged↗
PubMed1991

Soluble CD4 in patients with rheumatoid arthritis and osteoarthritis.

An ELISA was used to measure the soluble form of the leukocyte surface antigen CD4 (sCD4) in the sera and synovial fluids (SF) of patients with rheumatic diseases. Patients with rheumatoid arthritis (RA) had raised levels of sCD4 in both their sera and synovial fluid compared to age-matched healthy controls. In patients with osteoarthritis levels of sCD4 in SF and sera were lower than in RA but higher than in sera of healthy individuals. Mononuclear cells from the synovial fluid of RA patients were found to produce spontaneously high levels of sCD4, but autologous blood cells only produced comparable levels after in vitro stimulation with mitogenic lectin. In individual RA patients with active disease, serial sCD4 levels fell preceding clinical improvement. In three patients where serum sCD4 levels fell and clinical improvement occurred, subsequent small increases in serum sCD4 preceded increased clinical disease activity by up to 5 days. Synovial fluid levels of sCD4 correlated positively with soluble interleukin 2 receptor levels but no correlation was found with sCD8 levels. We conclude that the release of sCD4 reflects the involvement of T helper cells and macrophages in the pathogenesis of joint inflammation, especially in RA.

Adult↗
PubMed1991

Kinetics of IL1 beta mRNA and protein accumulation in human mononuclear cells.

Interleukin 1 beta (IL1 beta) is an inducible polypeptide with many roles in host defence and homoeostasis. It has also been implicated as a mediator of infectious, inflammatory and autoimmune diseases, and the kinetics of its production are relevant to an understanding of the pathogenesis of these conditions. We report here the time-course of IL1 beta production in human adherent monocytes. Both IL1 beta protein and mRNA were measured following cell activation with bacterial endotoxin (lipopolysaccharide; LPS), and pro-inflammatory crystals of monosodium urate (MSU), which cause arthritis and kidney disease. We also tested other crystal types associated with arthritis, namely hydroxylapatite and calcium pyrophosphate dihydrate. IL1 was absent from unstimulated cells, but IL1 beta mRNA accumulated rapidly after LPS or MSU stimulation and was associated with the later appearance of intracellular IL1 beta protein which was subsequently released from the cells (60% at 9 h). The other crystals failed to induce significant IL1 production. Our findings support the view that production of IL1 beta in human mononuclear cells is based on rapid translation of an inducible pool of mRNA and that no pre-formed mRNA or intracellular protein exists in normal blood monocytes. Further, although IL1 beta is translated without a conventional leader sequence, it is translocated extracellularly with the kinetics of a secretory protein.

Blood Proteins↗
PubMed1991

Plasma levels of interleukin-1-alpha in rheumatoid arthritis.

Interleukin-1-beta (IL-1 beta) has been implicated as an inflammatory mediator in rheumatoid arthritis (RA) but little is known about the related cytokine, IL-1 alpha, in this disease. IL-1 alpha has biological properties similar to IL-1 beta but, unlike IL-1 beta, remains mostly cell-associated. In this study plasma IL-1 alpha was measured by radioimmunoassay in patients with RA and in healthy controls. Plasma levels were compared with conventional measures of disease activity. The mean levels in the two groups were not significantly different and, within the patient group (n = 53), the only significant cross-sectional correlation was between plasma IL-1 alpha and ESR. In longitudinal studies, some individual patients had plasma IL-1 alpha levels that correlated with different measures of disease activity. We conclude that while IL-1 alpha may be involved in the immunopathogenesis of RA, its measurement in plasma seems to offer little of clinical value.

Adult↗
PubMed1991

Urate crystals stimulate production of tumor necrosis factor alpha from human blood monocytes and synovial cells. Cytokine mRNA and protein kinetics, and cellular distribution.

Crystals of monosodium urate (MSU) provide a dose-dependent stimulus for the production by human blood monocytes of tumor necrosis factor (TNF), a cytokine with proinflammatory properties; TNF activity was inhibited selectively by monoclonal antibody to TNF alpha. Biologically active cell-associated TNF activity peaked at 3 h and was exceeded at 6 h by extracellular activity, which peaked at 12-18 h. Comparable kinetics were observed with immunoreactive TNF alpha. TNF alpha mRNA accumulation in monocytes stimulated with MSU crystals appeared as a single peak at 2-4 h, kinetics compatible with rapid production of a short half-life transcript. In contrast, crystals of calcium pyrophosphate or of hydroxyapatite did not stimulate significant production of TNF or of message. Fresh tophaceous material from a patient with gout contained significant levels of TNF alpha and cells cultured from the tophus produced TNF alpha in vitro. In rheumatoid synovial cells, spontaneous release of TNF alpha was increased by in vitro exposure to MSU crystals. Taken together with earlier work, these results support an expanded view of gouty inflammation in which the crystal-stimulated production of cytokines provides a crucial link between crystal deposition and many of the clinical and pathological facts of both acute and chronic gouty arthritis.

Blotting, Northern↗
PubMed1990

A soluble form of the interleukin-1 receptor produced by a human B cell line.

A soluble protein that binds specifically to interleukin-1 (IL-1)beta was released from a B cell line (Raji). The covalently cross-linked binding protein/[125I]IL-1 beta migrated at 60 kDa by SDS-PAGE. The IL-1 receptor (IL-1R) on Raji cells had the same ligand specificity. Stimulation of Raji with dexamethasone increased surface expression of the IL-1R and the rate of release of soluble binding protein. A serine protease inhibitor prevented release of the binding protein and increased IL-1R expression on the cells. These results suggest that the soluble IL-1 beta binding protein is a proteolytically cleaved form of the novel B cell IL-1R.

Aprotinin↗
PubMed1990

Heat-shock protein 65 as a beta cell antigen of insulin-dependent diabetes.

The primary beta-cell antigen of insulin-dependent diabetes is thought to be a protein with a molecular weight of approximately 64 kD. Hyperthermic incubation and cytokines such as interleukin 1 beta, gamma interferon, and tumour necrosis factor induce synthesis of 64 kD protein by insulinoma cells. By western blot techniques, cross-reactivity was found between this 64 kD protein and monoclonal antibodies directed against Mycobacterium tuberculosis heat-shock protein 65, but not with antibodies directed against a similar epitope of M leprae heat-shock protein 65. Binding of M tuberculosis heat-shock protein 65 antibodies to interleukin-1 beta-treated cells was inhibited by prior addition of serum from insulin-dependent diabetic patients which contained antibodies to 64 kD beta-cell antigen. It is suggested that heat-shock protein 65 may be the 64 kD beta-cell antigen and that autoreactivity to an epitope of heat-shock protein 65 may confer susceptibility to insulin-dependent diabetes mellitus.

Animals↗
PubMed1990

Identification of an interleukin-1 beta binding protein in human plasma.

A covalent cross-linking technique was used to bind iodinated interleukin-1 (IL1) alpha and beta to plasma proteins. One specific IL1 beta binding protein was observed, that when cross-linked to 125I-IL1 beta migrated to approximately 60 kDa on SDS-PAGE. The protein did not bind IL1 alpha. The 43 -kDa protein was partially purified using a wheat germ agglutinin affinity column. The isolated factor again specifically bound IL1 beta, and appeared to consist of single chain glycoprotein. The protein was heat stable and had a rapid association time with IL1 beta. This protein may be an important carrier molecule for IL1 beta in vivo.

Animals↗