Biomedical subjects

I D Walker

Publications and source records attributed to I D Walker.

At least 73 records · Page 4Linked to original sources

PubMed1997

Congenital thrombophilia.

The heritable defects which are at present accepted as proven to be associated with familial venous thrombosis are deficiency of antithrombin (AT), protein C (PC) or protein S (PS) and the FV Leiden mutation. In women from symptomatic kindred each of these defects is associated with increased risk of pregnancy-associated venous thrombosis and increased risk of fetal loss and other vascular complications of pregnancy. The risks appear to be greatest for some types of AT deficiency. These defects are very common but there is growing evidence that congenital thrombophilia is a multigene defect and abnormalities of AT or of the PC-PS system represent only part of the genetic thrombotic predisposition in symptomatic families. Currently it seems reasonable to focus resources on women with AT or PC-PS system abnormalities who are themselves already symptomatic or who come from symptomatic families rather than screen whole populations for these defects. In symptomatic families screening of females around the time of puberty allows time for education and counselling. Pregnancies should be planned, and each pregnancy in each patient managed individually. In general though, women with AT deficiency from symptomatic families require anticoagulant prophylaxis throughout pregnancy and for at least 3 months post-partum, whereas those with PC-PS system defects may require third-trimester plus post-partum prophylaxis or post-partum anticoagulant prophylaxis only.

Antithrombins↗

PubMed1997

Positive direct antiglobulin test due to antiphospholipid antibodies in normal healthy blood donors.

OBJECTIVES: Positive direct antiglobulin tests (DAT) occur occasionally, without obvious cause, among normal blood donors. This is sometimes associated with a false-positive serologic test for syphilis. Positive DATs due to antiphospholipid antibodies (APAbs) are well documented in the antiphospholipid syndrome with systemic lupus erythematosus and the primary antiphospholipid syndrome. METHODS: Using conventional haematologic and serologic methods, we tested 474,545 normal blood donors in the west of Scotland during the years 1991 through 1993. RESULTS: We identified 42 normal blood donors with a positive DAT and observed three who had positive DATs and false-positive VDRL tests, all of whom expressed raised APAbs. CONCLUSION: APAbs may be an incidental cause of positive DATs among healthy blood donors.

Antibodies, Antiphospholipid↗

PubMed1997

Five antithrombin variants, four associated with thrombosis.

We have identified five mutations in antithrombin by direct sequencing of exons amplified using polymerase chain reaction. Four of these mutations are associated with thrombosis, three cause type I antithrombin deficiency and one has features of a type II deficiency. The fifth variant appears to have no functional consequences. The type I mutations are in exon 2, exon 3b and exon 4. The first of these is a nonsense mutation causing substitution of a Tyr-->stop at position -16 within the secretion signal sequence. The second is a missense mutation resulting in the substitution Cys-->Ser at position 247. This disrupts the disulphide bond with Cys 430 leaving a free cysteine residue and the C-terminus unconstrained. The third type I mutation is an in-frame deletion resulting in the loss of Ile 186. This is a highly conserved residue in the serpin superfamily and will predictably result in the disruption of the F-helix. The fourth mutation, in exon 3a, results in the substitution of Ser 162 by Asn. This residue is sited in the E-helix and the replacement of the buried side chain of serine by the larger asparagine side chain will predictably cause structural perturbation. The last example, Val 415-->Asp, was an incidental finding as a follow up investigation of a nephrotic patient. Although one other member of the family also had the mutation there was no linked history of thrombotic disease.

Adult↗

PubMed1997

Asymptomatic coinheritance of heterozygous plasminogen deficiency and the factor VLeiden mutation.

Deficiency of plasminogen has been postulated by some authors as a possible thrombophilic abnormality, though this remains controversial. We have previously identified a cohort of individuals with plasminogen deficiency from a study to determine plasminogen levels within the general population. All were asymptomatic for thrombosis at initial identification. We followed this cohort over a 5-year period, with no venous thrombotic events recorded, although one patient did suffer a myocardial infarction. One family was identified with asymptomatic coinheritance of both plasminogen deficiency and the factor VLeiden mutation. There was no apparent venous thrombotic risk conferred upon any of our cohort of individuals by inheritance of plasminogen deficiency, and, in addition, the combination of the factor VLeiden mutation with plasminogen deficiency which was observed in three individuals did not result in thrombotic events. However, the cohort is small and we cannot entirely exclude plasminogen deficiency as a possible mild thrombophilic defect.

Adult↗

PubMed1997

Mycoplasma synoviae has two distinct phase-variable major membrane antigens, one of which is a putative hemagglutinin.

Mycoplasma synoviae is a major pathogen of poultry, causing synovitis and respiratory infection. A cluster of 45- to 50-kDa membrane proteins is immunodominant in strain WVU-1853. Four distinct proteins were identified in this cluster by high-pressure liquid chromatography. Monoclonal antibodies and monospecific antisera against each established that they fell into two groups, MSPA and MSPB, each containing two members distinguishable by a difference in hydrophobicity. A 25- to 30-kDa membrane protein (MSPC) was shown to be antigenically related to the MSPB proteins. Considerable variation in the size and expression of MSPA and MSPB was observed among different strains of M. synoviae. Examination of expression in colonies of strain WVU-1853 established that both MSPA and MSPB (and MSPC) were phase variable. Immunostaining of MSPB (and MSPC) with monoclonal antibodies exhibited quantal variation, with three distinct levels observed between and within colonies. Hemadsorption by M. synoviae colonies was also found to be phase variable, with some colonies exhibiting sectorial expression of hemadsorption. Monospecific antisera against MSPA inhibited hemagglutination, but neither monoclonal antibodies nor monospecific antisera against MSPB could inhibit hemagglutination. However, loss of the capacity to hemadsorb by individual clones was associated with loss of expression of both MSPA and MSPB. These findings have elucidated the complexity of structure, function, and expression of the 45- to 50-kDa membrane protein cluster of M. synoviae, and they suggest that all members of the cluster may be involved in adhesion.

Antibodies, Monoclonal↗

PubMed1997

Risk factors for pregnancy associated venous thromboembolism.

In an attempt to reduce the incidence of pregnancy associated venous thromboembolism (PA-VTE), some researchers have advocated screening of all women for the factor V(Leiden) mutation during early pregnancy. We have conducted a large retrospective study (over 72,000 deliveries) to determine if this would be useful. Sixty-two objectively confirmed venous thrombotic events (51 DVT, 11 PE) were recorded at two maternity units in the UK. The incidence of DVT was 0.71 per 1000 deliveries (95% CI 0.5-0.9) with 0.50 occurring in the antenatal period (95% CI 0.34-0.66) and 0.21 in the puerperium (95% CI 0.11-0.31). The incidence of PE was 0.15 per 1000 deliveries (95% CI 0.06-0.24), 0.07 antenatal (95% CI 0.01-0.13) and 0.08 in the puerperium (95% CI 0.02-0.14). Of these 62, 50 attended for follow-up and thrombophilia screening. 28% of all episodes of PA-VTE had no clinical risk factor for thrombosis or an identifiable thrombophilic abnormality. Deficiency of antithrombin was identified in 12% of individuals (95% CI 3-21) and the factor V(Leiden) mutation in 8% (95% CI 0.5-15.5). Based on estimates of the prevalence of the factor V(Leiden) mutation in the population, we estimate that the thrombotic risk for a woman during pregnancy or the puerperium with the defect is approximately 1 in 400-500. This figure would not lend support to the idea of random screening for the mutation in early pregnancy.

Adult↗

PubMed1996

Increased fetal loss in women with heritable thrombophilia.

BACKGROUND: A successful outcome of pregnancy requires an efficient uteroplacental vascular system. Since this system may be compromised by disorders of haemostasis associated with a prothrombotic state, we postulated that maternal thrombophilia might be a risk factor for fetal loss. We studied the relation between heritable thrombophilic defects and fetal loss in a cohort of women with factor V Leiden or deficiency of antithrombin, protein C, or protein S. METHODS: We studied 1384 women enrolled in the European Prospective Cohort on Thrombophilia (EPCOT). Of 843 women with thrombophilia 571 had 1524 pregnancies; of 541 control women 395 had 1019 pregnancies. The controls were partners of male members of the EPCOT cohort or acquaintances of cases. We analysed the frequencies of miscarriage (fetal loss at or before 28 weeks of gestation) and stillbirth (fetal loss after 28 weeks of gestation) jointly and separately. FINDINGS: The risk of fetal loss was increased in women with thrombophilia (168/571 vs 93/395; odds ratio 1.35 [95% Cl 1.01-1.82]). The odds ratio was higher for stillbirth than for miscarriage (3.6 [1.4-9.4] vs 1.27 [0.94-1.71]). The highest odds ratio for stillbirth was in women with combined defects (14.3 [2.4-86.0]) compared with 5.2 (1.5-18.1) in antithrombin deficiency, 2.3 (0.6-8.3) in protein-C deficiency, 3.3 (1.0-11.3) in protein-S deficiency, and 2.0 (0.5-7.7) with factor V Leiden. The corresponding odds ratios for miscarriage in these subgroups were 0.8 (0.2-3.6), 1.7 (1.0-2.8), 1.4 (0.9-2.2), 1.2 (0.7-1.9), and 0.9 (0.5-1.5). Significantly more pregnancy terminations had been done in women with thrombophilia than in controls (odds ratio 2.9 [1.8-4.8]); this discrepancy was apparent in nine of 11 participating centres and for all thrombophilia subgroups. INTERPRETATION: Women with familial thrombophilia, especially those with combined defects or antithrombin deficiency, have an increased risk of fetal loss, particularly stillbirth. Our findings have important implications for therapy and provide a rationale for clinical trials of thromboprophylaxis for affected women with recurrent fetal loss.

Abortion, Spontaneous↗

PubMed1996

Origin and development of the gamma delta T-cell system in sheep: a critical role for the thymus in the generation of TcR diversity and tissue tropism.

Ruminant gamma delta T cells are concentrated at epithelial surfaces and share many features in common with species such as mice and humans which contain relatively fewer gamma delta T cells than ruminants. To date no gamma delta T cells with invariant TcR have been found in sheep and the generation of gamma delta TcR diversity which is thymic dependent follows a developmentally regulated sequence. Analysis of thymic export of gamma delta T cells shows that emigration of gamma delta T-cell subsets increases markedly during fetal life and after birth suggesting intrathymic processes leading to mature gamma delta T cells may change during development. Skin homing gamma delta T cells acquire their tissue tropism inside the thymus and pathways of recirculation of gamma delta T cells to skin are laid down during fetal development independent of antigen and remain stable through into adult life.

Animals↗

PubMed1996

Low thrombosis rate seen in blood donors and their relatives with inherited deficiencies of antithrombin and protein C: correlation with type of defect, family history, and absence of the factor V Leiden mutation.

We have previously identified a group of blood donors with inherited deficiencies of either antithrombin (AT) or protein C who appear to have a relatively low thrombosis rate. In the 5 years that have elapsed since initial identification of these individuals, resistance to activated protein C (APC resistance), which is associated with the factor V Leiden gene mutation, has emerged as an important and highly prevalent inherited thrombophilic risk factor. We have followed 28 donors/relatives with deficiency of AT (median age 48 years, range 16-77) and 23 with deficiency of protein C (median age 44 years, range 15-79) over a period of 5 years. During the study period only one individual, who was previously symptomatic, has suffered a thrombotic event which occurred spontaneously whilst on warfarin. We have now excluded coinheritance of APC resistance due to the factor V Leiden mutation in our cohort. Our findings demonstrate that individuals with single inherited thrombophilic defects are not uncommon and are frequently asymptomatic. The absence of the factor V Leiden mutation may in part explain the low thrombosis rate observed, and lends support to the hypothesis that multiple thrombophilic defects may be necessary for the development of thrombosis.

Adolescent↗

PubMed1996

Management of suspected acute venous thromboembolism in a general and maternity hospital.

A retrospective study of the management of patients with suspected acute deep venous thrombosis (DVT) and pulmonary thromboembolism (PTE) in a general and maternity hospital was conducted over a two month period in 1992. Ninety six patients with suspected DVT/PTE were identified, of whom only two were pregnant. Forty four patients had suspected DVT and confirmatory investigations were performed in 84%. The most common risk factor for DVT was intra-venous drug (IVD) use. Unfractionated heparin was prescribed to all patients except one with acute DVT at an average daily dose of 25,000 iu. In patients receiving heparin, 68% had measurements of the activated partial thromboplastin time (APTT) ratio and on 38% of occasions the result was subtherapeutic. Complications of heparin therapy were infrequent. Fifty two patients had suspected PTE and 50 underwent ventilation/perfusion (V/Q) scanning. No patient underwent pulmonary angiography. The management of patients with normal, low and high probability V/Q scans was in keeping with the guidelines, but only 8% [corrected] of patients with an intermediate V/Q scan result had further investigations and 33% received heparin. This study revealed suboptimal anticoagulation of patients with acute DVT and scope for improvement in the management of patients with an intermediate V/Q scan result.

Acute Disease↗

PubMed1996

Type I antithrombin deficiency: five novel mutations associated with thrombosis.

The genetic basis of Type I antithrombin deficiency has been investigated in six unrelated kindred with positive histories of thrombosis using a PCR amplification/direct sequencing approach. Four frameshift mutations, all introducing premature translation termination codons were identified. Thus, deletions, of a C at nucleotide position 2599 or 2600, a G at position 2601-2602 and a CT dinucleotide at position 7428-7429 were detected in three kindred and confirmed by restriction enzyme analysis. The identical insertion, of a T at nucleotide 2770, was observed in two apparently unrelated families. This finding may have been due to a founder effect since antithrombin gene polymorphism analysis showed all affected individuals to share a common haplotype. An in frame deletion of 6 bp at nucleotide position 2690-2696 causing the removal of codons 76 and 77 encoding Ile 76 and Phe 77 was also detected indicating that these amino acids are essential for stability of the mature antithrombin.

Adult↗

PubMed1996

Two recombinant tissue factor reagents compared to conventional thromboplastins for determination of international normalised ratio: a thirty-three-laboratory collaborative study. The Steering Committee of the UK National External Quality Assessment Scheme for Blood Coagulation.

Recent advances in recombinant technology have led to the development of prothrombin time (PT) reagents containing recombinant tissue factor which has been lipidated to allow expression of procoagulant activity. In this study we have compared International Normalised Ratios (INRs) determined using two such reagents and conventional thromboplastins in widespread use in the UK. Lyophilised plasma samples from eight different warfarinised patients were distributed to 33 laboratories in the UK. Each participant determined prothrombin times on 20 local fresh normal plasmas (used to derive mean normal PT and calculate INR) and the eight lyophilised samples, using manual technique and the following thromboplastins; Recombiplastin (Ortho Diagnostics Ltd); Innovin (Baxter Diagnostics Ltd); the conventional thromboplastin in local use. For eight plasmas the mean INRs determined with different reagents were as follows: Innovin (33 laboratories) - 3.4; Manchester Reagent (MR = 8 laboratories) - 3.4; Recombiplastin (33 laboratories) - 3.7; Instrumentation Laboratory (IL = 13 laboratories) - 4.4. Mean INR results with Recombiplastin were on average 7% greater than those obtained with Innovin, 8% greater than results with MR and 18% less than INRs with IL thromboplastin. There was no significant difference between results obtained with Innovin and MR. In contrast INRs obtained with IL were markedly (mean 28%) greater than results obtained with Innovin. This study employed lyophilised plasma and it is possible that some of the relationships described are influenced by this. However, the lyophilisation process employed did not influence the relationship between INRs of warfarinised plasmas obtained by the four main reagents described, indicating that the results are relevant to routine clinical practice. In conclusion, our data show some important differences are present between INRs determined using Recombiplastin, Innovin and two conventional thromboplastins.

Humans↗

PubMed1996

Isolated familial plasminogen deficiency may not be a risk factor for thrombosis.

Despite many reports of individuals with congenital plasminogen deficiency and thrombosis, there is still uncertainty whether heterozygous deficiency represents a real thrombophilic risk factor or simply a coincidental finding. We have addressed this issue by testing for plasminogen deficiency in a cohort of 9611 blood donors. Out of 66 donors with reduced plasminogen activity on two occasions 28 were shown to have a familial deficiency state (including 3 with dysplasminogenaemia). Our observed prevalence rate for familial plasminogen deficiency, calculated at 2.9/1000 (95% CI = 1.9-4.2 per 1000), was not significantly different from that calculated from published reports of congenital plasminogen deficiency in thrombotic cohorts (5.4/1000). Furthermore, with only two exceptions, all 80 donors and relatives with familial deficiency were asymptomatic with regard to thrombosis-including a 29 year old donor with suspected compound heterozygous hypoplasminogenaemia. These findings add further weight to the argument that familial heterozygous plasminogen deficiency, at least in isolation, does not constitute a significant thrombotic risk factor. However, it remains uncertain whether plasminogen deficiency, when combined with other thrombophilic conditions, may become more clinically important.

Adult↗

PubMed1995

Characterisation of a novel Kunitz-type molecule from the trematode Fasciola hepatica.

A low molecular mass monomeric protein termed Fh-KTM (Fasciola hepatica Kunitz-type molecule) was isolated from the trematode Fasciola hepatica. Fh-KTM is a single polypeptide of 58 amino acids and a Mr of 6751. The complete amino acid sequence of Fh-KTM was determined and revealed significant similarity to the Kunitz-type (BPTI) family of proteinase inhibitors. Several polymorphisms were observed suggesting that more than one Fh-KTM molecule may be expressed by this parasite. Modified proline residues were shown to occur at all four positions in this protein as 3-hydroxy derivatives. This is the first report of 3-hydroxyproline residues in a Kunitz-type molecule. Indirect immunofluorescence and immunogold labelling revealed that Fh-KTM is an abundant molecule within the parasite localised to the gut, the parenchymal tissue and the tegument of adult F. hepatica. Serine protease inhibition assays revealed that Fh-KTM exhibited little or no inhibition against chymotrypsin, kallikrein, urokinase or key serine proteases of the blood coagulation pathways. However, Fh-KTM was able to inhibit trypsin even though the P1 reactive amino acid of Fh-KTM was a leucine residue.

Amino Acid Sequence↗

PubMed1995

Two novel antithrombin variants, Asn187Asp and Asn187Lys, indicate a functional role for asparagine 187.

Three unrelated families have been identified with mutations involving asparagine 187. Two of these families are asymptomatic and were identified during the screening of random blood donors, whilst the third has a history of recurrent thromboembolic disease. In two families the mutation (6460 AAC-->GAC) results in an asparagine to aspartate substitution and is associated with normal immunological levels of antithrombin but a reduction in functional activity. In the third family the mutation (6462 AAC-->AAA) results in an asparagine to lysine substitution at residue 187 and is associated with a parallel reduction in both immunological and functional antithrombin levels. Asparagine 187 is located in the middle of the F helix of antithrombin and forms the major link between the F helix and strand 3 of the A sheet. The F helix is seen to overlie the A sheet of the molecule and moves with strands 2 and 3 of this sheet as they open to allow entry of the reactive site loop to form strand 4. Substitutions of asparagine 187 are, therefore, likely to disrupt this sliding movement leading to a loss of inhibitory activity.

Adult↗

PubMed1995

Thrombocytopenia, antithrombin deficiency and extensive thromboembolism in pregnancy: treatment with low-molecular-weight heparin.

Pregnancy limits the therapeutic options for managing thrombocytopenia which occurs in 5% of patients on heparin. We describe a case of extensive thromboembolism associated with antithrombin (AT) deficiency complicated by thrombocytopenia which resolved when low-molecular-weight heparin was instituted. A primigravid woman presented at 11 weeks gestation with bilateral femoral occlusive thrombi extending above the renal veins. Investigations revealed AT deficiency, thrombocytopenia and renal infarction. After low-molecular-weight heparin was substituted for unfractionated heparin, the thrombocytopenia resolved and although the pregnancy was lost, the patient made a full recovery.

Adult↗

PubMed1995

Expression studies on four members of the pMGA multigene family in Mycoplasma gallisepticum S6.

A large family of related genes known as pMGA exists in the avian pathogen Mycoplasma gallisepticum but only a single member of this family was previously found to be expressed in one strain of this bacterium. In this work two unrelated strains of M. gallisepticum were also shown by amino-terminal sequencing to express a unique pMGA polypeptide in both cases. To investigate pMGA gene selection in M. gallisepticum, mRNA expression was analysed in M. gallisepticum strain 56 using reverse transcription-PCR (RT-PCR) and Northern blot techniques with probes for several members of the pMGA multigene family. It was shown that the pMGA message is 2.2 kb in size and is monocistronic. RT-PCR detected four different pMGA mRNA molecules but their relative yields were significantly affected by magnesium concentration. By quantitative Northern analysis, the relative abundances of the four pMGA mRNAs in M. gallisepticum S6 total RNA was determined: the pMGA1.1 mRNA predominated [1.88 ng (micrograms total RNA)-1] but at least three other pMGA genes were found to be transcribed but at much lower levels (20 to 40-fold lower). The pMGA1.1 mRNA is expressed at a level five times higher than the tuf gene, known to be one of the most abundantly expressed proteins in the prokaryotic cell. The start point of transcription for pMGA1.1 was determined and probable promoter assigned. From these data it appears likely that transcriptional control plays a major role in the selection of pMGA gene expression in the M. gallisepticum cell.

Amino Acid Sequence↗

PubMed1995

Coronary artery bypass surgery in patients with inherited antithrombin deficiency.

Antithrombin (AT) replacement in coronary artery bypass grafting procedures in three individuals with inherited antithrombin deficiency is described. All three had a significant personal or family history of thrombotic disease. All patients achieved satisfactory AT levels throughout bypass and in the postoperative period. All received heparin prophylaxis in the postoperative period. None suffered thrombotic or bleeding complications.

Aged↗