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Biomedical subjects

S J Machin

Publications and source records attributed to S J Machin.

At least 37 records · Page 2Linked to original sources
PubMed2001

Frequency and prevention of symptomless deep-vein thrombosis in long-haul flights: a randomised trial.

BACKGROUND: The true frequency of deep-vein thrombosis (DVT) during long-haul air travel is unknown. We sought to determine the frequency of DVT in the lower limb during long-haul economy-class air travel and the efficacy of graduated elastic compression stockings in its prevention. METHODS: We recruited 89 male and 142 female passengers over 50 years of age with no history of thromboembolic problems. Passengers were randomly allocated to one of two groups: one group wore class-I below-knee graduated elastic compression stockings, the other group did not. All the passengers made journeys lasting more than 8 h per flight (median total duration 24 h), returning to the UK within 6 weeks. Duplex ultrasonography was used to assess the deep veins before and after travel. Blood samples were analysed for two specific common gene mutations, factor V Leiden (FVL) and prothrombin G20210A (PGM), which predispose to venous thromboembolism. Asensitive D-dimer assay was used to screen for the development of recent thrombosis. FINDINGS: 12/116 passengers (10%; 95% CI 4.8-16.0%) developed symptomless DVT in the calf (five men, seven women). None of these passengers wore elastic compression stockings, and two were heterozygous for FVL. Four further patients who wore elastic compression stockings, had varicose veins and developed superficial thrombophlebitis. One of these passengers was heterozygous for both FVL and PGM. None of the passengers who wore class-I compression stockings developed DVT (95% CI 0-3.2%). INTERPRETATION: We conclude that symptomless DVT might occur in up to 10% of long-haul airline travellers. Wearing of elastic compression stockings during long-haul air travel is associated with a reduction in symptomless DVT.

Aged↗
PubMed2001

Lack of pathogenic mutations in the 5'-untranslated region of the thrombopoietin gene in patients with non-familial essential thrombocythaemia.

Thrombopoietin (TPO) is thought to be the major physiological regulator of thrombopoiesis, and, in general, circulating levels are inversely proportional to megakaryocyte and platelet mass. However, normal or elevated TPO levels are found in patients with essential thrombocythaemia (ET) and the reason for this is not fully understood. Recent studies have shown that four kindreds with hereditary thrombocythaemia (HT) have point mutations in the 5'-untranslated region (UTR) of the TPO gene which lead to increased TPO translation. In order to determine whether similar mutations are present in apparently acquired ET, in particular in those patients with polyclonal myelopoiesis, we have studied this region in 50 ET patients using neutrophil DNA. The known HT mutations were investigated using polymerase chain reaction with mismatch primers and restriction enzyme digestion; only wild-type alleles were detected. Single-stranded conformation polymorphism (SSCP) analysis of exons 1-4 identified a C-->T substitution at nucleotide 3767. However, this appears to be a common polymorphism, as it was present at the same frequency in haematologically normal controls and is unlikely to be of pathological significance. These results demonstrate that mutations in the 5' UTR of the TPO gene are not the cause of the normal or elevated TPO levels in acquired ET.

5' Untranslated Regions↗
PubMed2001

Anti-prothrombin antibodies: assay conditions and clinical associations in the anti-phospholipid syndrome.

Anti-phospholipid antibodies (aPL) are associated with an increased risk of thrombosis and recurrent fetal loss. Antibodies to prothrombin (aPT) have been associated with the anti-phospholipid syndrome (aPS). We assessed variations in aPT assay methodology to optimize an aPT method that was used to screen patients with aPS (n = 66). Detection of aPT using enzyme-linked immunosorbent assay was influenced by the concentration of the capture antigen, the microtitre plate type and the buffer system. The combination of gamma-irradiated plates, a phosphate-buffered saline buffer and coating antigen of 10 microg/ml prothrombin was the most sensitive. Both serum and citrate samples are suitable for the detection of aPT. Under these conditions aPT IgM but not IgG were found to be associated with thrombosis and/or fetal loss.

Abortion, Habitual↗
PubMed2001

Protein S levels are lower in women receiving desogestrel-containing combined oral contraceptives (COCs) than in women receiving levonorgestrel-containing COCs at steady state and on cross-over.

This study aimed to identify specific haemostatic changes that might account for previous observations of higher venous thromboembolic risk among users of combined oral contraceptives (COCs) containing desogestrel (DSG) than levonorgestrel (LNG). Sixty-three current users of monophasic 30 microg oestrogen COCs containing either LNG or DSG omitted one pill-free interval (PFI), switching immediately either to the opposite formulation for one cycle or continuing with the same pill. Venesection followed the initial PFI after one cycle (21 tablets) and two cycles (42 tablets) of continuous pill taking, and after the following PFI. Protein S was lower in users of DSG than LNG formulations after the first PFI (mean +/- SD, 0.67 +/- 0.09 vs 0.76 +/- 0.10, P < 0.001) and after one cycle (0.61 +/- 0.09 vs 0.76 +/- 0.09, P < 0.0001). Protein S decreased when switching from LNG to DSG pills (0.77 +/- 0.07-0.65 +/- 0.06, P < 0.0001), mirrored by an increase at switching from DSG to LNG formulations (0.61 +/- 0.08-0.73 +/- 0.10, P < 0.005). Mean protein S levels remained within the normal range. Three different markers of thrombin generation remained unaltered. Potential explanations for COC-related thrombotic events are 'acquired resistance to activated protein C' or inhibition of fibrinolysis. A potential role has been described for protein S deficiency in both. A further triggering factor is a probable prerequisite for actual thrombosis, but pill-takers whose levels of protein S were in the lowest percentiles may be at greatest risk.

Adult↗
PubMed2001

Anti-tissue factor pathway inhibitor activity in patients with primary antiphospholipid syndrome.

The association between antiphospholipid antibodies and an increased risk of thrombosis in antiphospholipid syndrome (aPS) patients is probably caused by numerous mechanisms, including the effects of antibodies to phospholipid-binding proteins such as beta(2)-glycoprotein I and prothrombin. In this study, we investigated the inhibition of tissue factor pathway inhibitor (TFPI) in 33 patients with primary antiphospholipid syndrome (PAPS). TFPI was measured in PAPS patients using an amidolytic assay, dependent on the generation of activated factor X (Fxa), and this was compared with 55 healthy subjects. Functional levels of TFPI (mean +/- SD) were significantly lower in PAPS patients (0.89 +/- 0.37 U/ml) than the control group (1.05 +/- 0.15 U/ml) (P = 0.02). The difference was caused by a subset of five patients who had TFPI levels below the lower 99% confidence interval of the normal reference range, representing increased FXa generation in the assay system. IgG fractions were isolated from these five patients and five control subjects, then incorporated into normal plasma to measure FXa generation in the TFPI assay system. FXa generation was increased when polyclonal rabbit anti-human TFPI IgG (P < 0.0001) or PAPS IgG (P = 0.0001) were added to normal plasma, demonstrating inhibition of TFPI. The apparent anti-TFPI activity demonstrated in the five subjects with PAPS in this study may represent a significant new mechanism for thrombosis in patients with aPS, as it implies that increased tissue factor FVIIa-mediated thrombin generation might occur.

Adult↗
PubMed2001

Increased circulating platelet-leucocyte complexes and platelet activation in patients with antiphospholipid syndrome, systemic lupus erythematosus and rheumatoid arthritis.

It is possible that platelet activation may play a pathogenic role in the increased risk of thrombosis associated with antiphospholipid antibodies (APA). In this study, levels of in vivo platelet activation were measured in 20 patients with primary antiphospholipid syndrome (PAPS) and 30 systemic lupus erythematosus (SLE) patients (14 of whom had secondary APS) using sensitive flow cytometry. Soluble P-selectin levels were also assayed. Platelet CD63 expression was significantly higher in PAPS than normal controls (P = 0.007), as well as SLE patients with and without secondary APS (P = 0.03 and P = 0.002 respectively). PAC-1 binding was significantly higher in PAPS than the control group (P = 0.007) and SLE patients without APS (P = 0.015). Platelet-leucocyte complexes were significantly higher in SLE patients than both PAPS and the control group, and platelet-monocyte complexes were significantly increased in PAPS compared with the control group. (Platelet-leucocyte complexes were also significantly higher than controls in 10 rheumatoid arthritis (RA) patients without APA). Soluble P-selectin levels were significantly higher in PAPS and SLE patients than the control group. Platelet CD62p expression, annexin V binding and platelet microparticle numbers were not increased in PAPS or SLE patients. We conclude that there is evidence of increased platelet activation in PAPS and SLE, and this is important to note as it may have potential therapeutic implications with respect to use of antiplatelet agents in these patients.

Adult↗
PubMed2001

The impact of oral anticoagulant therapy, factor VIII level and quality of factor V-deficient plasma on three commercial methods for activated protein C resistance.

Several methods are now available for the laboratory assessment of activated protein C resistance (APCR). In this study, we evaluated two activated partial thromboplastin time-based assays [Coatest activated protein C (APC) and Diagen protein C activator (PCA)], with and without predilution of test plasma in factor V-deficient plasma (FVdp) and an amidolytic assay (Immuno Ltd, Vienna, Austria). Testing plasmas from normal volunteers who had received 1-deamino-8-D-arginine vasopressin (DDAVP) also assessed the effect of elevated factor VIII on APCR. In the unmodified clotting tests, the Coatest kit gave overlapping results for normal and heterozygous FV:Q506 samples; some FV:Q506 samples on oral anticoagulant therapy (OAT) were misclassified as normal, and some normal samples with high factor VIII levels would be classified as APC resistant. The unmodified Diagen kit correctly classified these three types of sample, but had the disadvantage that prolonged PCA clotting times gave serious problems with instrument end-point detection. Both kits modified by diluting the samples in FVdp correctly classified all the samples, as well as samples from patients with lupus anticoagulant (LA) and patients receiving heparin. The Immunochrom kit correctly classified the normal and FV:Q506 samples, but would have misclassified most normal persons on OAT as well as some patients with LA or receiving heparin therapy as APC resistant.

Activated Protein C Resistance↗
PubMed2001

Plasma levels of factor XIIa and factor VIIa are increased but not related in primary hyperlipidaemia.

The lipolysis of triglyceride-rich lipoproteins may provide a surface that supports the activation of factor XII (FXII) with subsequent activation of factor VII (FVII). Plasma levels of activated FVII (FVIIa) but not activated FXII (FXIIa) are increased in the post-prandial state when there is a transient increase in triglyceride levels. We compared plasma levels of FXIIa antigen in control subjects (n = 33) and in patients with chronically elevated lipids (primary hyperlipidaemia, n = 49), with FVIIa and markers of thrombin generation. Results are given as median (first and third quartiles). Plasma levels of FXIIa [2.34 (1.68-3.32) ng/ml versus 1.53 (0.93-1.86) ng/ml, P = 0.0002], FVIIa [3.02 (2.15-4.64) ng/ml versus 2.20 (1.66-2.56) ng/ml, P = 0.0004], thrombin-antithrombin complexes [3.08 (2.16-5.54) microg/I versus 2.13 (1.46-2.84) microg/l, P = 0.005] and prothrombin fragment 1 + 2 (Pro F1 + 2) [1.28 (1.08-1.50) nmol/l versus 0.92 (0.65-1.08) nmol/l, P = 0.0001] were increased compared with controls irrespective of the type of hyperlipidaemia. In hyperlipdaemic subjects, levels of Pro F1 + 2 were correlated with FVIIa (r = 0.56, P = 0.0002) and FXIIa (r = 0.31, P = 0.03). These results suggest increased activation of both FVII and FXII in hyperlipidaemic subjects, which correlates with increased thrombin generation. Given the lack of correlation between levels of FXIIa and FVIIa, it remains to be established whether the increase in FXIIa is responsible for increased FVIIa activity in this subject group.

Adult↗
PubMed2000

Pharmacokinetics and pharmacodynamics of sibrafiban alone or in combination with ticlopidine and aspirin.

AIMS: The purpose of this clinical study was to evaluate the effects of a ticlopidine/aspirin combination on the pharmacokinetics and pharmacodynamics of sibrafiban and the tolerability of the combination therapy METHODS: Thirty-eight healthy male volunteers were randomized to receive one of the following treatments for 7 days: sibrafiban (n = 12), ticlopidine/aspirin (n = 12), or the combination treatment sibrafiban/ticlopidine/aspirin (n = 14). Concentrations of the active metabolite of sibrafiban, Ro 44-3888, in plasma and urine were determined by column-switching liquid chromatography combined with tandem mass spectrometry. The pharmacodynamics of sibrafiban and ticlopidine/aspirin were examined by measuring the inhibition of ADP- or collagen-induced platelet aggregation. RESULTS: The addition of ticlopidine/aspirin to sibrafiban did not significantly alter the pharmacokinetic parameters of Ro 44-3888. the geometric mean ratio for AUC(0,12h) was 110 (95% CI 0.82, 1.22). Separately, sibrafiban and ticlopidine/aspirin inhibited ADP-and collagen-induced platelet aggregation and the effects of the two treatments were additive. For example, the average inhibition of ADP-induced platelet aggregation over 12 h was 42% in the sibrafiban treated group, 55% in the ticlopidine/aspirin group and 69% in the sibrafiban/ticlopidine group. The bleeding time was prolonged in the treatments with ticlopidine/aspirin (8.1 min) and sibrafiban/ticlopidine/aspirin (8. 6 min) compared with sibrafiban alone (3.5 min). CONCLUSIONS: This study shows a significant pharmacodynamic interaction between sibrafiban and ticlopidine/aspirin. Consequently, the simultaneous administration of sibrafiban and ticlopidine/aspirin should be carefully monitored to ensure the patient's coverage with an antiplatelet drug without exposure to an excessive bleeding risk.

Adult↗
PubMed2000

The importance of locally derived reference ranges and standardized calculation of dilute Russell's viper venom time results in screening for lupus anticoagulant.

Three commercial dilute Russell's viper venom time (DRVVT) kits were evaluated at four UK centres experienced at performing lupus anticoagulant (LA) tests. Each centre established a normal reference range for the DRVVT ratio calculated against local pooled normal plasma from 20 healthy normal subjects. Plasma from LA-positive patients and LA-negative thrombophilia patients was also tested. DRVVT ratios and the degree of correction were assessed in a variety of ways to reflect not only the UK national Guidelines, but also the manufacturers' recommendations. The reference range data showed a normal distribution in each case, but considerable variation in the mean and SD between the centres and reagents, with the mean +2SD value ranging from 1.06 to 1.19. The use of an arbitrary DRVVT ratio of < 1.1 as the cut-off value for normality, which is applied in many laboratories, is therefore inappropriate. Although no single kit had a clear overall advantage in terms of sensitivity and specificity, the way in which the screen and confirmation data were analysed had a major impact on the interpretation of the results. A data analysis method employing a mean plus two standard deviations (SDs) cut-off for normality, and judgement regarding confirmation of LA based on a percentage correction of DRVVT ratio, was the simplest and most consistent, with overall sensitivity and specificity values of 81% and 94%, respectively, for uncomplicated LA-positive and LA-negative thrombophilia samples. We conclude that the 1991 BSCH Guidelines are in need of revision, each laboratory should establish its own normal reference range for the DRVVT ratio and a common method should be used for calculating the degree of correction with confirmation reagents, so that LA results can be correctly interpreted between laboratories. Standardizing DRVVT interpretation in this way should improve the consistency of LA detection.

Humans↗
PubMed2000

Von Willebrand factor--cleaving protease activity in congenital thrombotic thrombocytopenic purpura.

Thrombotic thrombocytopenic purpura (TTP) is characterized by microangiopathic haemolytic anaemia (MAHA), thrombocytopenia, fluctuating neurological impairment, renal dysfunction and fever. Both acquired and congenital forms are recognized. Recurrent episodes, which may be predictable (occurring every 21-28 d), are seen in congenital disease and may be treated by infusion with fresh-frozen plasma (FFP). Congenital TTP has recently been associated with deficiency of a novel von Willebrand factor (VWF)-cleaving protease. To investigate whether residual protease activity dictates clinical manifestations, we determined protease activity in three patients with congenital TTP of varying severity. Intrinsic VWF-cleaving protease activity of a range of plasma-derived products was also assessed as one patient had been successfully maintained for many years, initially using an intermediate-purity factor VIII concentrate (Kryobulin) and then cryoprecipitate. All three patients had a severe absolute deficiency of VWF-cleaving protease activity (< 3%) up to 5 months after clinical symptoms. Three relatives were also found to have a mild reduction in protease activity (25-50%). Nevertheless, the intrinsic VWF-cleaving protease activity of plasma-derived products correlated with their clinical efficacy: significant (100%) protease activity was found in FFP, cryosupernatant, solvent-detergent-treated plasma, cryoprecipitate and Kryobulin. Two clinically ineffective factor VIII products (Fahndi and Haemate P) possessed only low protease activity (6.25% and 12.5% respectively). Although this suggests that VWF-cleaving protease activity is central to the pathogenesis of congenital TTP, either small differences in protease activity below 3% or hitherto unknown factors have a profound influence on clinical phenotype. The possible use of factor VIII concentrates in the treatment of this condition also warrants further investigation.

ADAM Proteins↗
PubMed2000

Ontogeny of beta 2 glycoprotein I and annexin V in villous placenta of normal and antiphospholipid syndrome pregnancies.

beta2-glycoprotein I (beta2GPI) and annexin V (AV) have been implicated in the pathophysiology of the antiphospholipid syndrome (APS). We investigated their placental expression in normal villous tissues throughout gestation; first trimester n = 10, early second trimester; n = 4, preterm; n = 5) and term; n = 7 and in APS (2 first trimester, 1 preterm and 8 term deliveries). beta2GPI and AV were both expressed by the placenta from as early as seven weeks gestation and were colocalised to the syncytiotrophoblast. beta2GPI staining was also observed in stromal cells, being present in phagocytic Hofbauer cells and surrounding newly formed fetal vessels in a perivascular pattern, from seven to seventeen weeks gestation. An abnormal morphological distribution of AV was noted in one first trimester APS placenta, and for beta2GPI in a further first trimester placenta. When placental proteins were extracted from villous tissue, the concentration of AV/mg protein in term APS placentas (median, interquartile range) (aPS; 8.16, 7.879.72 microg/mg) was significantly higher (p <0.005) than normal term levels (normal; 2.47, 2.28-2.54 microg/mg). beta2GPI increased with advancing gestation (first trimester; 0.93, 0.64-1.26 microg/mg, term; 3.67, 2.58-4.48 microg/mg) in normal pregnancy. Term APS placentas had a reduced beta2GPI content (2.31, 1.87-2.49 microg/mg), p <0.05. The placental role of these proteins remains to be identified.

Abortion, Habitual↗
PubMed1999

A large proportion of patients with a diagnosis of essential thrombocythemia do not have a clonal disorder and may be at lower risk of thrombotic complications.

Essential thrombocythemia (ET) is traditionally considered to be a clonal disorder. No specific karyotypic abnormalities have been described, but the demonstration of clonality using X-chromosome inactivation patterns (XCIPs) has been used to differentiate ET from a non-clonal reactive thrombocytosis. However, these assays may be difficult to interpret, and contradictory results have been reported. We have studied 46 females with a diagnosis of ET according to the Polycythemia Vera Study Group (PVSG) criteria. XCIP results in 23 patients (50%) were uninterpretable due to either constitutive or possible acquired age-related skewing. Monoclonal myelopoiesis could be definitively shown in only 10 patients. Thirteen patients had polyclonal myelopoiesis, and in 8, it was possible to exclude clonal restriction to the megakaryocytic lineage. Furthermore, there was no evidence of clonal progenitors in purified CD34(+)CD33(-) and CD34(+)CD33(+) subpopulations from bone marrow of 2 of these 13 patients. There was no difference between patients with monoclonal and polyclonal myelopoiesis with respect to age or platelet count at diagnosis, duration of follow-up, incidence of hepatosplenomegaly, or hemorrhagic complications. However, polyclonal patients were less likely to have experienced thrombotic events (P =.039). These results suggest that ET is a heterogeneous disorder, and the clinical significance of clonality status warrants investigation in a larger study.

Adolescent↗
PubMed1999

Pharmacokinetics and pharmacodynamics of Ro 44-3888 after single ascending oral doses of sibrafiban, an oral platelet aggregation inhibitor, in healthy male volunteers.

AIMS: This study constituted the first administration of the oral platelet inhibitor, sibrafiban, to humans. The aim was to investigate the pharmacokinetics and pharmacodynamics of Ro 44-3888, the active principle of sibrafiban, after single ascending oral doses of sibrafiban. Particular emphasis was placed on intersubject variability of the pharmacokinetic and pharmacodynamic parameters of Ro 44-3888. METHODS: The study consisted of three parts. Part I was an open ascending-dose study to determine target effect ranges of sibrafiban. Part II, a double-blind, placebo-controlled, parallel-group study, addressed the intersubject variability of pharmacokinetic and pharmacodynamic parameters of the active principle at a sibrafiban dose achieving an intermediate effect. Part III was a double-blind, placebo-controlled, ascending-dose design covering the complete plasma concentration vs pharmacodynamic response curve of sibrafiban. RESULTS: At sibrafiban doses between 5 mg and 12 mg, the pharmacokinetics of free Ro 44-3888 in plasma were linear whereas those of total Ro 44-3888 were non-linear because of the saturable binding to the glycoprotein IIb-IIIa receptor. Saturation of the GP IIb-IIIa receptor was reached at plasma concentrations of 15.9 ng ml-1. At sibrafiban doses up to 2 mg, ADP-induced platelet aggregation was inhibited by 50%, whereas the inhibition of TRAP-induced platelet aggregation was about 20-30%. At the higher doses, ADP-induced platelet aggregation was almost completely inhibited while a clear dose-response could be observed with TRAP-induced inhibition of platelet aggregation at sibrafiban doses of 5 to 12 mg. Ivy bleeding time increased very steeply with dose with a significant prolongation observed at doses of 5 to 7 mg of sibrafiban (5-7 min, >30 min in one case). At a sibrafiban dose of 12 mg, the stopping criterion for dose escalation (prolongation of the Ivy bleeding time >30 min in three out of four subjects per dose group) was reached. The interindividual coefficients of variation of the integrated pharmacokinetic and pharmacodynamic parameters (AUC and AUE) were below 20%, thus lying well within the pre-set level of acceptance. CONCLUSIONS: With a low intersubject variability of its pharmacokinetic and pharmacodynamic parameters, linear pharmacokinetics and pharmacodynamic effects closely related to its plasma concentrations, Ro 44-3888 has good pharmacological prerequisites for a well controllable therapy of secondary prevention of arterial thrombosis in patients with acute coronary syndrome.

Adenosine Diphosphate↗
PubMed1999

Platelet activation responses in vitro to human mast cell activation.

Mast cells are thought to play an important role in atherogenesis and plaque rupture, but their role in the subsequent platelet activation and thrombus formation is unclear. Tryptase positive cells (KU812T+) were established from the KU812 cell line as an in vitro model of human mast cells and used to study the effect of mast cell activation on human platelets. Overnight incubation of KU812T+ with IgE and subsequent challenge with anti-IgE caused the release of heparinoid substances which inhibited 1 microg/ml collagen-induced platelet aggregation. KU812T+ challenged with compound 48/80 produced a releasate that had no apparent heparinoid content but caused full platelet aggregation. These findings showed that, although activation of KU812T+ via FcepsilonR1 partially abrogated collagen-induced platelet aggregation, activation of the C5a receptor signalling pathway, by compound 48/80, caused the release of potent platelet-activating substances. This cell culture model offers a unique insight into the role of platelet-mast cell interactions in arterial thrombogenesis.

Cell Line↗