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Biomedical subjects

D E Woolley

Publications and source records attributed to D E Woolley.

At least 73 records · Page 4Linked to original sources
PubMed1993

IgE antibodies specific for cartilage collagens type II, IX and XI in rheumatic diseases.

Serum samples from 149 patients with RA and other rheumatological diseases, and 57 non-arthritic controls have been assayed for IgE antibodies to the cartilage collagen types II, IX and XI in their native and denatured state. Using an improved ELISA technique together with antigen-binding inhibition studies to confirm specificity, 10 of the 149 (7%) patients showed IgE antibodies to human collagen type II and bovine collagen types II, IX or XI. Some patients responded to only one collagen type whereas others had IgE positive responses to two or all three collagen species. Most of the IgE responses detected were directed towards the denatured collagens. Those sera showing an IgE response to bovine type II collagen produced a similar response to the human equivalent, including two patients with SLE. None of 57 control subjects demonstrated IgE specificity for any of the cartilage collagens. Patients with IgE specificity for the cartilage collagens did not demonstrate IgM or IgA specificity for these antigens, but two of these patients showed IgG responses to type II and XI collagens. Whereas eight patients were exclusively IgE-positive for the cartilage collagens, others expressed specificities for IgG, IgM or IgA. It therefore appears that the specific autoimmune profile for each patient is often different from others, both in terms of the class of immunoglobulin expressed and the collagen antigens recognised. At present no correlations were observed between the IgE-positive patients and their clinical assessment and/or prognosis.

Adult↗
PubMed1991

Serum IgE anti-cartilage collagen antibodies in rheumatoid patients.

The presence of circulating IgG, IgA and IgM antibodies to native cartilage collagens in some patients with rheumatoid arthritis (RA) suggests that an autoimmune response to cartilage collagens may be involved in the pathogenesis of RA. However, the relevance of such antibodies to the pathological process remains unclear, and it is likely that many humoral and cellular derived factors combined to trigger events leading to the chronicity of the rheumatoid lesion. Since histological and biochemical studies have suggested the involvement of mast cells in the rheumatoid joint, we have studied the frequency of IgE antibodies directed against the cartilage collagens type II, IX and XI in patients with active rheumatoid disease. Of the 91 patients' sera tested, 32 had significant levels of IgE anti-cartilage collagen antibodies when compared with non-arthritic controls. Total serum IgE levels did not correlate with the presence of IgE anti-collagen antibodies, nor were any patients positive for IgE antibodies to fibronectin, a widely distributed extracellular matrix component. These results are consistent with an allergic type I hypersensitivity reaction to cartilage antigens in RA involving mast cell and basophil degranulation.

Arthritis, Rheumatoid↗
PubMed1991

Toxicokinetics of Ro 5-4864, lindane and picrotoxin compared.

The effects produced by IP administration of these three agents in the rat were compared because of in vitro evidence that each modulates the picrotoxinin site of the GABAA receptor. For each, hypothermia had the lowest threshold and convulsions the next, with hypophagia produced only by the highest dose of either Ro 5-4864 or lindane. Convulsant effects had a shorter latency and a shorter duration than did hypothermia. Hypophagia, when present, lasted the longest. Myoclonus was the seizure type with the lowest threshold for all three agents. At the highest dose, lindane produced a high incidence of maximal clonic (hopping) seizures, whereas Ro 5-4864 and picrotoxin produced a high incidence of maximal tonic seizures instead. On a mole/kg basis, picrotoxin was 40 times more effective than the other two agents and produced seizures which started later, peaked later, and persisted longest. Ro 5-4864 and lindane were effective at equimolar concentrations and, in combination, produced effects which suggested either dose-addition or synergism. The data are consistent with the hypothesis that the toxic effects of both Ro 5-4864 and lindane may be attributable, at least in part, to an action at a subpopulation of GABAA receptors.

Animals↗
PubMed1991

Ro 5-4864, like picrotoxin, enhances EPSP-spike coupling in the freely behaving rat.

To help determine its mechanism of action, the convulsant benzodiazepine Ro 5-4864 was administered (15-20 mg/kg) intraperitoneally (IP) and electrophysiological and behavioral effects were compared; parallel studies were conducted with picrotoxin (PTX; 1 mg/kg). Both PTX and Ro 5-4864 produced myoclonic seizures, primarily between 15-40 min after administration; myoclonus was followed by more severe seizures after PTX. Both Ro 5-4864 and PTX produced a maximal increase in amplitude and decrease in threshold of the population spike (PS) evoked in the dentate gyrus (DG) by stimulation of the dorsal perforant path prior to peak seizure activity; start latency of the PS and initial slope and amplitude of the population slow wave (SW) were not changed. Amplitude of the PS was already increased by 5 min after administration of Ro 5-4864 and was maximally increased 1.8- to 3-fold, depending on stimulus intensity, usually by 10 min. Similarly, by 20 min after administration, PTX had also increased PS amplitude in the absence of an effect on PS latency or the SW. The increase in PS amplitude without concomitant changes in the SW suggests that Ro 5-4864 enhanced coupling between the excitatory postsynaptic potential (EPSP) and firing of the postsynaptic neurons, i.e., it enhanced E-S coupling, as has also been suggested for PTX. The similarity in the effects of Ro 5-4864 and PTX suggests that antiGABAergic effects, perhaps along feedforward inhibitory pathways, are involved in both the seizures and enhanced E-S coupling.

Animals↗
PubMed1991

Mast cell modulation of tumour cell proliferation in rat mammary adenocarcinoma 13762NF.

Mast cells were shown to accumulate around the periphery of the invasive and metastatic rat mammary adenocarcinoma (MTLn3), and histological evidence of mast cell degranulation was observed during the later stages of this model. To assess the physiological role of mast cells in vivo we have used the mast cell-stabilising compound FPL 55618 applied i.p. daily at 1 mg kg-1 for 23 days. Using groups of 12 rats we have found that this compound inhibited tumour growth at the primary site by as much as 70% in most of the treated animals compared with the control group which received equivalent volumes of saline. When the drug treatment was stopped after 23 days, tumour growth of the test group accelerated over the next 7 days and reached a similar tumour size to that of control animals. Histological studies of the tumour and contiguous host tissue at day 24 of the experiment revealed numerous extra-tumoural mast cells often showing signs of degranulation at several sites around the tumour periphery in the control animals. Such observations were not seen in those animals receiving FPL 55618 where, in contrast to controls, numerous intact mast cells were often seen within the tumour mass. Following cessation of the MC-stabilising treatment progressive mast cell activation was evident within 2-4 days, primarily at the tumour periphery. In vitro studies have shown that drug concentrations equivalent to five times the in vivo dose had no effect on the proliferative rate or viability of the MTLn3 cells. Moreover, the proliferative rate of these cells in culture was significantly increased when exposed to soluble mast cell products. Thus our data indicate that a mast cell-stabilising compound has significant benefits in reducing tumour growth in vivo, an observation which supports the concept that mast cell:tumour cell interactions are important for the growth and invasive properties demonstrated by this model of breast carcinoma.

Adenocarcinoma↗
PubMed1991

Production of matrix metalloproteinase 3 (stromelysin) by cultured ovine endometrial cells.

Ovine endometrial cells (stromal plus epithelial) in primary culture release matrix metalloproteinase 3 (stromelysin, MMP-3), the enzyme that degrades various extracellular components including the basement membrane components collagen IV and laminin. The enzyme was detected in tissue culture medium by specific enzyme assay. A low level of MMP-3 activity was released from the cells (0.12 +/- 0.02 unit per 10(6) cells per 48 h; 1 unit degrades 1 microgram of reduced, carboxymethylated transferrin min-1 at 37 degrees C), but following stimulation by phorbol myristate acetate (PMA) a mean 4.4-fold increase in MMP-3 production was observed (to 0.53 +/- 0.13 unit per 10(6) cells per 48 h). The enzyme was released primarily in the form of its proenzyme, which could be activated in vitro by (4-aminophenyl)mercuric acetate (APMA). Stimulation of enzyme release from cells by PMA was not seen before 24 h and required PMA at a dose of 10 nM or higher. This finding was substantiated by Western blot analysis. Immunocytochemistry of cells that had been treated with PMA for 48 h, and with the ionophore, monensin, for 4 h, confirmed the potential of the endometrial cells to produce stromelysin. The release of MMP-3 from ovine endometrial cells suggests that this enzyme may have a role in assisting the passage of syncytial processes through the basement membrane during the process of implantation.

Animals↗
PubMed1990

Contrasting effects of the protein kinase C inhibitor, staurosporine, on cytokine and phorbol ester stimulation of fructose 2,6-bisphosphate and prostaglandin E production by fibroblasts in vitro. Comparative studies using interleukin-1 alpha, tumour necrosis factor alpha, transforming growth factor beta, interferon-gamma and 12-O-tetradecanoylphorbol 13-acetate.

It is known that both interleukin-1 alpha (IL-1 alpha) and 12-O-tetradecanoylphorbol 13-acetate (TPA) promote increases in intracellular levels of the glycolytic regulatory metabolite fructose 2,6-bisphosphate [Fru(2,6)P2] and in the production of prostaglandin E (PGE) by subcultured rheumatoid synovial cells (RSC) and human dermal fibroblasts in vitro. We report here that the protein kinase C inhibitor staurosporine enhanced the IL-1 alpha-induced increase in [Fru(2,6)P2] and PGE production by RSC, whereas in similar concentrations (3-30 nM) this inhibitor decreased the TPA-induced stimulation of these parameters. Staurosporine produced a similar enhancement of the response to IL-1 alpha by normal human dermal fibroblasts. The increased PGE production provoked by tumour necrosis factor alpha (TNF alpha) in RSC was also augmented by staurosporine, but, in contrast, the increases in cellular [Fru(2,6)P2] induced by transforming growth factor beta (TGF beta) and interferon-gamma (IFN-gamma) were diminished. Thus the protein kinase C inhibitor staurosporine discriminates not only between the effects produced by IL-1 alpha and TPA, but also between those of IL-1 alpha and two other cytokines (but not between IL-1 alpha and TNF alpha). These findings suggest that IL-1 alpha and probably TNF alpha act via an intracellular mechanism different from that mediating the action of TPA, TGF-beta and IFN-gamma, and provide evidence that staurosporine is capable of amplifying the IL-1 signal.

Alkaloids↗
PubMed1990

Degradation of cartilage collagens type II, IX, X and XI by enzymes derived from human articular chondrocytes.

Conditioned culture medium derived from Interleukin-I alpha-activated human articular chondrocytes contained both collagen- and proteoglycan-degrading activities. Preparations of soluble type I collagen and the cartilage collagens type II, IX, X and XI were all degraded when incubated with the conditioned culture medium at 35 degrees C. Fractionation of the enzymic activities using column chromatography with Ultragel AcA 34 and Heparin-Sepharose allowed the separation and identification of neutral proteinase, collagenolytic and proteoglycan-degrading activities. Eluant fractions which contained type I collagenase activity effectively degraded collagen type II, but these fractions did not correspond precisely with those which degraded collagen types IX, X and XI. These observations indicate that chondrocytes have the potential to produce a conventional interstitial type II collagenase together with other enzymes having some specificity for the minor collagens. Thus IL-1-activated chondrocytes produce a range of collagenolytic and proteoglycan-degrading enzymes which can process most of the structural components of the cartilage matrix.

Cartilage, Articular↗
PubMed1989

Comparative and combined effects of transforming growth factors alpha and beta, interleukin-1 and interferon-gamma on rheumatoid synovial cell proliferation, glycolysis and prostaglandin E production.

Enhanced cell proliferation, glycolysis and prostaglandin E production are all characteristic features of rheumatoid synovial tissue. The interrelationships of these three cellular parameters have been examined using rheumatoid synovial fibroblasts and their responses to specific cytokines in vitro. Transforming growth factor alpha (TGF alpha) caused a more than threefold increase in synovial cell proliferation whilst transforming growth factor beta (TGF beta), interleukin-1 alpha (IL-1 alpha) and interferon-gamma (IFN-gamma) produced only marginal changes. The combined addition of IL-1 alpha with TGF beta resulted in an enhanced proliferative response comparable with that produced by TGF alpha. Glycolysis, estimated by glucose utilisation and measurements of the glycolytic regulatory metabolite fructose 2,6-bisphosphate was significantly stimulated by TGF beta, IL-1 alpha and IFN-gamma, but less so by TGF alpha. Prostaglandin E production was significantly increased by IL-1 alpha to an extent much greater than that produced by TGF alpha or TGF beta, although the combined addition of IL-1 alpha with either TGF alpha or beta resulted in a synergistic increase in PGE production, a response partly diminished by the addition of IFN-gamma. These findings suggest that the extent to which a cytokine stimulates glycolysis is not consistently related to its mitogenicity, and that cytokine combinations which stimulate high levels of PGE production (a growth inhibitor) will not necessarily be associated with a reduced rate of cellular proliferation in cultured, adherent, rheumatoid synovial fibroblasts.

Biological Factors↗
PubMed1989

"Central" and "peripheral" benzodiazepines and kinetics of lindane-induced toxicity.

Because hypothermia and anorexia were previously found to be more sensitive indices of the effects of lindane than were convulsions, these endpoints were used to quantify the ability of benzodiazepines (BDs) and phenytoin either to ameliorate or exacerbate the toxicity of lindane in the rat. After administration of lindane (40 or 50 mg/kg) in oil per os, toxicity was counteracted by phenytoin and the "central" BD agonists diazepam and clonazepam, but was worsened by Ro 5-4864 a "peripheral" BD agonist. Clonazepam and diazepam were each more effective in counteracting lindane-induced anorexia than in stimulating food intake, presumably because the animals had been fasted and probably even controls ate maximally when food was presented. Diazepam alone (3 injections in 1 day) produced withdrawal-induced decreased food intake the following day. Clonazepam and diazepam alone each transiently decreased colonic temperature, yet effectively blocked the more severe hypothermia produced by lindane. Ro 5-4864 by itself did not produce any measurable effects, yet exacerbated all of the effects, including lethal effects, of lindane. The present findings are compatible with other evidence that lindane and Ro 5-4864 act at the picrotoxinin receptor of the GABAA-activated chloride channel and that systemic administration of agents acting at this site may produce a constellation of effects, including seizures, hypothermia and anorexia.

Animals↗
PubMed1989

Kinetics and thresholds of several indices of lindane-induced toxicity.

The effects of lindane, administered either IP (4, 6 or 8 mg/kg in dimethylsulfoxide, 0.5 ml/kg) or PO (30, 40 or 50 mg/kg in oil, 1 ml/kg), were compared in male rats. Effects appeared later and lasted longer after PO administration. After either route, convulsant effects occurred first and were over before hypothermic effects were maximal. Also, hypothermia subsided before hypophagia ended. After IP administration, hypothermia had the lowest threshold and convulsions the next lowest; reduced food intake was produced only by the highest dose, which also produced 43% deaths. By contrast, after PO administration, all doses reduced food intake and produced hypothermia, but deaths did not occur after even the highest dose. The different time courses and thresholds for the different indices of toxicity suggest that different target sites or mechanisms may be involved. Thus, effects on the gut, in addition to effects on the brain, may account for the longer duration and greater sensitivity of reduced food intake, compared to lethal and hypothermic effects, after PO administration of lindane. After administration of lindane IP, peripheral vasodilation (as demonstrated by an increase in tail temperature) preceded colonic hypothermia. This could be explained if lindane inappropriately activated heat-loss mechanisms.

Administration, Oral↗
PubMed1989

Cleavage of collagen type X by human synovial collagenase and neutrophil elastase.

Chick-derived native cartilage collagen type X and the pepsin-resistant 45 kDa fragment were susceptible to attack by human synovial collagenase and neutrophil elastase at 25 degrees C and 35 degrees C. Synovial collagenase cleaved type X collagen at two sites which were equally susceptible to the enzyme. In contrast, elastase produced three cleavages, but the sensitive loci showed different susceptibilities as judged by the sequential appearance of specific breakdown products. Both enzymes produced a major, enzyme-resistant fragment of approximately 32 kDa at 35 degrees C, and both of these end-products co-migrated in SDS polyacrylamide gels. Human chondrocyte-derived collagenase also degraded native, 59 kDa collagen type X in a similar manner to that shown by the synovial collagenase. From amino acid sequence data the enzyme cleavages probably occur at three regions of sequence imperfection. The specific cleavages brought about by synovial or chondrocyte collagenase, or neutrophil elastase, may have a functional catabolic role in vivo, and in vitro might provide useful tools with which to further analyse specific properties of the native collagen type X molecule.

Animals↗
PubMed1989

A new cell line (8701-BC) from primary ductal infiltrating carcinoma of human breast.

A cell line, designated 8701-BC, was established in culture from tissue fragments of primary ductal infiltrating carcinoma of human breast. The cell cultures after the sixth passage were devoid of contaminating fibroblasts as judged by the positive staining of all cells with the specific epithelial cell markers carcinoembryonic antigen (CEA), tissue polypeptide antigen (TPA) and cytokeratin 8. The epithelial nature of these cells was confirmed by ultrastructural analyses which demonstrated the retention of specific structural properties characteristic of the original tumour. The cells possessed an abnormal karyotype with 55-60 chromosomes per cell with numerous rearrangements. They do not express HLA antigens and the c-myc gene was not amplified. The 8701-BC cells have a doubling time of approx. 29h and have been maintained in culture for more than 100 passages. These properties suggest the establishment of a human neoplastic cell line which, with its ability to produce homotrimer collagen in vitro, will provide a useful model system for the study of tumour cell:stromal matrix interactions.

Breast Neoplasms↗
PubMed1988

Susceptibility of cartilage collagens type II, IX, X, and XI to human synovial collagenase and neutrophil elastase.

The action of purified rheumatoid synovial collagenase and human neutrophil elastase on the cartilage collagen types II, IX, X and XI was examined. At 25 degrees C, collagenase attacked type II and type X (45-kDa pepsin-solubilized) collagens to produce specific products reflecting one and at least two cleavages respectively. At 35 degrees C, collagenase completely degraded the type II collagen molecule to small peptides whereas a large fragment of the type X molecule was resistant to further degradation. In contrast, collagen type IX (native, intact and pepsin-solubilized type M) and collagen type XI were resistant to collagenase attack at both 25 degrees C and 35 degrees C even in the presence of excess enzyme. Mixtures of type II collagen with equimolar amounts of either type IX or XI did not affect the rate at which the former was degraded by collagenase at 25 degrees C. Purified neutrophil elastase, shown to be functionally active against soluble type III collagen, had no effect on collagen type II at 25 degrees C or 35 degrees C. At 25 degrees C collagen types IX (pepsin-solubilized type M) and XI were also resistant to elastase, but at 35 degrees C both were susceptible to degradation with type IX being reduced to very small peptides. Collagen type X (45-kDa pepsin-solubilized) was susceptible to elastase attack at 25 degrees C and 35 degrees C as judged by the production of specific products that corresponded closely with those produced by collagenase. Although synovial collagenase failed to degrade collagen types IX and XI, all the cartilage collagen species examined were degraded at 35 degrees C by conditioned culture medium from IL1-activated human articular chondrocytes. Thus chondrocytes have the potential to catabolise each cartilage collagen species, but the specificity and number of the chondrocyte-derived collagenase(s) has yet to be resolved.

Arthritis, Rheumatoid↗
PubMed1988

Plasminogen activator release from cultured murine mast cells.

Mast cells from the Furth murine mastocytoma tumour line were found to contain significant levels of plasminogen activator (PA). Cultured cells released PA activity into the culture medium in parallel with the release of histamine, and both were proportionately increased following exposure to degranulating agents. Pretreatment of the mast cells with cycloheximide did not alter their total PA content or their ability to release PA. These studies suggest that PA is a prestored granule constituent. The ability of PA to generate plasmin from plasminogen suggests an important role for mast cell PA in fibrinolysis and tissue degradation, observations that have been associated with mast cell degranulation and infiltration in vivo.

Animals↗