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Biomedical subjects

J Dawes

Publications and source records attributed to J Dawes.

At least 91 records · Page 5Linked to original sources
PubMed1985

Platelet-associated proteins in human breast cyst fluids.

Thrombospondin, beta-thromboglobulin and platelet factor 4 were measured by radioimmunoassay in fluids from 50 human breast cysts. beta-Thromboglobulin concentrations were less than 4-330 ng/ml (median 13.5 ng/ml), while normal plasma contained 14-80 ng/ml. Cyst fluids contained 8-225 ng/ml platelet factor 4 with a median of 25 ng/ml (range in plasma 6.5-29 ng/ml). Thrombospondin levels in cyst fluids were 65-55 000 ng/ml (median 2 500 ng/ml) and 92% contained concentrations above the plasma range (58-215 ng/ml). When cyst fluids were classified by their electrolyte composition, group II cysts had similar levels of platelet factor 4 but significantly higher levels of beta-thromboglobulin and thrombospondin than group I. From the remarkably high concentrations of thrombospondin within cyst fluids and the differences between cyst types, it seems that either the protein is synthesised locally or selectively accumulated. Increased platelet activation is unlikely to be the source of the high thrombospondin levels in group II cysts.

Beta-Globulins↗
PubMed1985

The measurement of heparin and other therapeutic sulphated polysaccharides in plasma, serum and urine.

The competitive binding assay described will specifically and accurately measure concentrations of administered heparin in biological fluids with a sensitivity of 60 ng ml-1. Neither endogenous glycosaminoglycans, nor plasma proteins such as ATIII and PF4 interfere in the assay. Semi-synthetic highly sulphated heparinoids and LMW heparin can also be measured. Using this assay heparin clearance followed simple first-order kinetics over the dose range 100-5,000 units, but the half-life was strongly dose-dependent. There was good correlation with heparin activity measurements by APTT and anti-Xa clotting assays. Plasma concentrations were measurable for at least 5 h following subcutaneous injection of 10,000 units of heparin. Excretion in the urine could be followed after all but the lowest intravenous dose. This assay, used in conjunction with measurements of heparin anticoagulant activity, will be valuable in the elucidation of mechanisms of action of heparin and the heparinoids, and in the assessment and management of problems related to heparin therapy.

Antithrombin III↗
PubMed1985

Plasma thrombospondin in patients with chronic renal failure, liver disease and splenectomy.

Thrombospondin (TSP), is a major constituent of human blood platelet alpha-granules. Stimulation of platelets causes the release of TSP in parallel with other alpha-granule constituents such as beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) but the thrombospondin plasma in vivo half life is significantly greater than beta-TG and PF4. The aim of this study was to assay TSP levels in plasma of patients with chronic renal failure (CRF), liver disease (LD) and following splenectomy. The TSP values were then compared to the patients plasma levels of two traditional markers of platelet activation, beta-TG and PF4, and to fibronectin (FN) and von Willebrand factor (VIII:vWF). Plasma TSP levels (67.6 +/- 16.9 ng/ml) assayed in 14 CRF patients were significantly higher (p less than 0.05) than those measured in 28 donors (55.5 +/- 11.7 ng/ml). No correlation was observed, in CRF patients, between the TSP level and PF4 (2.5 +/- 1.5 ng/ml), beta-TG (131.1 +/- 21 ng/ml), FVIII:vWF (252 +/- 85%), or FN (102 +/- 33%) plasma levels. The TSP plasma level in CRF patients was significantly correlated (p less than 0.02) with that of fibrinopeptide A (4.1 +/- 1.9 ng/ml). Although the beta-TG (23.5 +/- 6.9 ng/ml) and PF4 (2.9 +/- 2 ng/ml) plasma levels in six LD patients were normal, the TSP levels (82.5 +/- 39.1 ng/ml) were significantly increased (p less than 0.01). Thrombospondin plasma levels (77.1 +/- 20.1 ng/ml) in 14 patients having undergone splenectomy were significantly increased (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1985

Enhancement of factor VIII-von Willebrand factor ristocetin cofactor activity by monoclonal antibodies.

Five monoclonal antibodies to human von Willebrand factor were selected for characterization from 18 produced in murine hybridomas. All showed a high and specific affinity for human von Willebrand factor (vWf) but exhibited little if any cross-reaction with sera from other species. The antibodies defined four epitopes on vWf, none of which were involved in platelet binding. Binding of two distinct antibodies at one of these epitopes was associated with enhancement of the rate of vWf-dependent platelet agglutination in the presence of ristocetin. This effect was more noticeable when cryosupernatant plasma was used in place of normal plasma as the source of vWf, and was not explicable simply in terms of antibody-induced cross-linking of vWf.

Animals↗
PubMed1985

Metabolism of sodium pentosan polysulphate in man measured by a new competitive binding assay for sulphated polysaccharides--comparison with effects upon anticoagulant activity, lipolysis and platelet alpha-granule proteins.

Three human volunteers were injected with a range of doses of pentosan polysulphate, SP54, i.v. or s.c. A competitive binding assay (CBA) for sulphated polysaccharides was used to detect circulating SP54 after doses as low as 1 mg i.v. and a linear relationship was observed between the peak plasma concentration of SP54 measured by CBA and the administered dose. A comparison was made between the clearance of SP54 measured by CBA and its anticoagulant and lipolytic activities. SP54 was detectable by CBA after doses which caused no alteration in activated partial thromboplastin time (APTT) or anti-factor Xa activity but after which a small increase of lipase activity was measurable. After SP54 at 10 mg i.v. or 100 mg s.c. anti-factor Xa activity was 4-6 times greater than would be expected from the in vitro activity of the concentrations of SP54 measured by CBA. Like heparin and other heparin analogues, SP54 caused an increase in plasma concentrations of platelet factor 4 (PF4) without a concomitant rise in beta-thromboglobulin (beta-TG). It is concluded that the newly developed CBA will provide a more sensitive means than conventional bioassays for the determination of plasma concentrations of SP54.

Adult↗
PubMed1985

Comparison of secretion and subcellular localization of von Willebrand protein with that of thrombospondin and fibronectin in cultured human vascular endothelial cells.

Cultured human vascular endothelial cells synthesize von Willebrand protein, thrombospondin and fibronectin. These proteins are secreted in the culture medium and incorporated into the extracellular matrix. We have compared the subcellular localization and the secretion of these proteins in response to stimulants in cultured human umbilical vein endothelial cells. Density gradient centrifugation using colloidal silica showed that the storage and secretion organelle with von Willebrand protein did not contain thrombospondin or fibronectin. Indirect immunofluorescence microscopy indicated that thrombospondin and fibronectin are not located in the rod-shaped organelles containing von Willebrand protein. Thrombin, ionophore A23187 and phorbol myristate acetate did not affect secretion of thrombospondin and fibronectin, while von Willebrand protein secretion was stimulated upon incubation of cells with these agents for 30 min. Prolonged incubation of cultured endothelial cells after a 1-h treatment with phorbol myristate acetate resulted in an increased secretion of von Willebrand protein into the conditioned medium; in contrast, accumulation of thrombospondin and fibronectin in endothelial cell-conditioned medium was decreased. These findings indicate that, unlike in platelets, these major endothelial proteins are not located in the same subcellular compartments. Von Willebrand protein is distinguished from thrombospondin and fibronectin both by its unique subcellular localization and its secretion rate in response to stimuli.

Blood Coagulation Factors↗
PubMed1985

The presence in blood of both glycosaminoglycan and mucosal mast cell protease following systemic anaphylaxis in the rat.

The appearance in blood of rat mast cell protease II (RMCPII) and glycosaminoglycan (GAG) was examined in normal and Nippostrongylus brasiliensis-primed rats challenged intravenously with worm antigen. Systemic release of these two products occurred only in immune recipients of antigen; substantial levels of RMCPII were also present in the intestinal perfusates of these same rats and there was depletion of both RMCPII and mucosal mast cells (MMC) from the intestinal mucosa. Depletion of MMC was evident after staining for proteoglycan or for serine esterase and the mast cell counts with both histochemical techniques were highly correlated. Taken together, the results suggest that MMC are likely to be the principal source of secreted GAG and RMCPII.

Anaphylaxis↗
PubMed1984

Quantitation by radioimmunoassay of thrombospondin synthesised and secreted by human endothelial cells.

Synthesis and secretion of thrombospondin by cultured human endothelial cells was quantified using a sensitive radioimmunoassay. Two endothelial lines were examined; those from umbilical veins and those from saphenous veins. The foetal line secreted approximately 17 micrograms/10(6) cells of the antigen at 24 hr, whereas the saphenous line secreted five times less in the same period. Intracellular values were similar in both lines. Thrombin and calcium ionophore A23187 were added to umbilical vein endothelial cultures. In contrast to platelets where both agents cause a rapid release reaction, no such thrombospondin release was observed in endothelial cells.

Calcimycin↗
PubMed1984

Monoclonal antibodies directed against human alpha-thrombin and the thrombin-antithrombin III complex.

Human alpha-thrombin was poorly immunogenic in Balb/c mice. Nevertheless, following fusion of spleen cells from a responding mouse with NS-1 cells, 8 mouse monoclonal antibodies against alpha-thrombin were isolated, and 6 were characterised. Five of these were isotype IgG2a, and one was IgG1. One, EST 1, bound thrombin only minimally, and was directed against a neoantigen on the thrombin-ATIII (T-AT) complex. This antibody also recognised a site on prothrombin, though with much lower affinity. Its binding was markedly temperature-dependent, indicating a requirement for molecular mobility. A second antibody, EST 4, would not bind the T-AT complex. It inhibited both the clotting and amidase activities of thrombin, and modification of the active site histidine, but not the active site serine, reduced the affinity constant of binding to EST 4. This antibody appears to be directed against an epitope in the vicinity of the enzyme active site. The epitopes for EST 1 and EST 4 were both remote from those of the other monoclonal antibodies, EST 2, 6, 7 and 8. These four competed with each other for binding to thrombin, and all inhibited clotting but not amidase activity. Thrombin binding was not affected by modification of the active site, though formation of the T-AT complex reduced the affinity of binding to EST 6 and EST 8. These monoclonals recognise epitopes in the region of the fibrinogen binding site.

Animals↗
PubMed1984

Detection of enhanced in vivo platelet alpha-granule release in different patient groups--comparison of beta-thromboglobulin, platelet factor 4 and thrombospondin assays.

During the platelet release reaction beta-thromboglobulin (beta TG), platelet factor 4 (PF4) and thrombospondin (TSP) are released from the platelet into plasma and assays of these proteins can be used to monitor in vivo platelet activation. We have assessed their relative merits as markers of the in vivo platelet alpha-granule release reaction in a number of patient groups which have previously been shown to have elevated plasma beta TG and/or PF4 levels. It is concluded that in diseases or conditions not complicated by its reduced clearance, beta TG is the most sensitive marker of in vivo platelet alpha-granule release. However, the TSP assays may be the least ambiguous when monitoring the platelet alpha-granule release reaction in patients with renal failure who are undergoing haemodialysis with heparin anticoagulation. Under these circumstances plasma beta TG, but not PF4 or TSP, levels are elevated because of impaired renal catabolism, and the presence of a heparin-releasable reservoir of PF4 on the endothelium complicates the use of the PF4 assay. In liver failure none of these assays may accurately reflect platelet alpha-granule release because of impaired hepatic or renal elimination of the proteins.

Adolescent↗
PubMed1984

Metabolism of sodium pentosan polysulphate in man--catabolism of iodinated derivatives.

An iodinated derivative of the heparin analogue SP54 has been prepared and used in conjunction with unlabelled SP54 to study the catabolism and organ distribution of this potential antithrombotic agent in healthy human volunteers. As observed previously with 125I-heparin, we found that the 125I-SP54 was rapidly cleared from the circulation, returning later in a desulphated form. Organ distribution studies with 123I-SP54 suggested that the liver and spleen were major sites of desulphation. Gel filtration and Polybrene binding showed the presence of sulphated macromolecular SP54 and desulphated macromolecular and depolymerised SP54 in post-injection urines. No depolymerised material was present in plasma suggesting depolymerisation occurs in the kidney.

Adult↗
PubMed1984

A comparative study using immunological and biological assay of the haemostatic responses to DDAVP infusion venous occlusion and exercise in normal men.

In a group of six normal male volunteers, infusion of DDAVP, venous occlusion and exercise were shown to increase plasma levels of factor VIII and plasminogen activator, activity and antigen, to different extents and at differing rates. Any mechanisms suggested to explain release of these proteins by various stimuli should account for such differences. All three stimuli could also increase plasma levels of prostacyclin metabolites, although this was only significant for high doses of DDAVP. Other potential endothelial markers, such as fibronectin and thrombospondin, showed no specific increase after any of the stimuli.

Arginine Vasopressin↗
PubMed1984

The platelet release reaction in cardiovascular disease: evaluation of plasma betathromboglobulin as a marker of a prothrombotic state.

Plasma betathromboglobulin as a marker of in vivo platelet release was found to be significantly higher in 131 patients with prosthetic cardiac valves and 104 patients with coronary artery disease, compared with 133 normal individuals. Patients with double mechanical valves had a higher mean betathromboglobulin than those with single mechanical valves, who in turn had higher values than those with single heterograft valves. Thirteen patients adequately anticoagulated at the time of a thromboembolic event had significantly higher betathromboglobulin levels than 5 with thromboemboli when not anticoagulated, suggesting platelets were responsible for the emboli in the former group. The presence of coronary risk factors in normal individuals did not affect betathromboglobulin; however, when two or more risk factors were present in patients with ischaemic heart disease, mean betathromboglobulin was higher than when only one or no risk factors were present. This suggests greater platelet activation in the presence of more widespread atherosclerotic disease. Thus mean plasma betathromboglobulin is higher in groups of patients with disorders known to have thrombotic complications, but the wide range of values, which overlap with normals, prevent the clinical interpretation of the plasma betathromboglobulin level found in any individual patient.

Adult↗
PubMed1984

Platelet and coagulation factors in proliferative diabetic retinopathy.

Plasma beta-thromboglobulin, platelet factor 4, fibrinogen, fibrinopeptide A, antithrombin III, factor VIII related antigen, alpha 2-macroglobulin, platelet count, and total glycosylated haemoglobin were measured in three well matched groups of subjects: non-diabetic controls, diabetics without retinopathy, and diabetics with proliferative retinopathy. beta-thromboglobulin and platelet factor 4 concentrations were significantly higher in the diabetics with retinopathy than in the controls and platelet factor 4 was also increased in the diabetics without retinopathy compared with controls. Fibrinogen concentration was raised in diabetics without retinopathy compared with controls, diabetics with retinopathy compared with controls, and diabetics with retinopathy compared with those without. Fibrinopeptide A concentration did not differ significantly between groups. Antithrombin III levels were increased in diabetics with retinopathy compared with controls, and in diabetics with retinopathy compared with those without. Factor VIII related antigen values were higher in both the diabetic groups when compared with the controls. Fibrinopeptide A concentration correlated with both beta-thromboglobulin and platelet factor 4 in each of the three groups. Haemostatic abnormalities in diabetes have been shown, although a hypercoagulable state has not been confirmed. These changes in platelet and coagulation function may be secondary to the development of microvascular disease and their role in the pathogenesis of retinopathy remains uncertain.

Adult↗