Methylenetetrahydrofolate reductase mutation and coronary artery disease.
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Coronary Disease↗
Biomedical subjects
E G Tuddenham
Publications and source records attributed to E G Tuddenham.
Activation of factor VII during alimentary lipemia occurs in healthy adults and patients with congenital factor XII or factor XI deficiency, but not in patients with factor IX deficiency.
Factor VII activity (FVIIc), a risk marker for coronary heart disease, is increased during postprandial lipemia. Factor VII activation accompanies lipolysis of triglyceride-rich lipoproteins, but the nature of this association and whether it is causal remain uncertain. To explore this issue, four patients with homozygous factor XII deficiency, four with complete factor XI deficiency, six with factor IX deficiency, and their respective age- and sex-matched controls were given two isocaloric dietary regimens, one providing on average 136 g fat and the other 19 g fat. Blood was taken before breakfast, immediately before lunch at 195 minutes, and at completion of the study at 390 minutes. All samples for each subject and matched control were assayed as one batch for FVIIc, activated factor VII, and factor VII antigen (FVIIag). Activation of factor VII was observed with the high-fat regimen but not with the low-fat regimen in all controls, factor XII-deficient patients, and factor XI-deficient patients. No factor VII activation was observed during either regimen in factor IX-deficient patients, but a normal postprandial responsiveness of factor VII to dietary fat was restored in one patient who replicated the study after factor IX therapy. Plasma FVIIag was not altered postprandially in either regimen in any group of patients or controls. Factor IX apparently plays an obligatory role in the postprandial activation of factor VII, although the mechanism remains to be determined.
Adult↗
Severe perinatal thrombosis in double and triple heterozygous offspring of a family segregating two independent protein S mutations and a protein C mutation.
Molecular genetic and phenotypic analyses were performed in a highly unusual case of combined protein S and protein C deficiency manifesting in a family in which a child had died perinatally from renal vein thrombosis. Antenatal diagnosis in a second pregnancy was initially performed by indirect restriction fragment length polymorphism (RFLP) tracking using a neutral dimorphism within the PROS gene and served to exclude severe protein S deficiency. Am umbilical vein blood sample at 22 weeks gestation showed isolated protein C deficiency. This pregnancy proceeded to a full-term delivery without thrombotic complications. Molecular genetic analysis of the PROC and PROS gene segregating in the family then yielded one PROC gene lesion in the father and two PROS gene lesions, one in each parent. These lesions were shown to segregate with the respective deficiency states through the family pedigree. Analysis of DNA from paraffin-embedded liver tissue taken from the deceased child showed the presence of both PROS mutations, as well as the PROC mutation. Genotypic analysis of the second child showed a PROC mutation, but neither PROS mutation consistent with its possession of normal protein S levels and a low/borderline protein C level. Antenatal diagnosis was then performed in a third pregnancy by direct mutation detection. However, although the fetus carried only the paternal PROS and PROC gene lesions, the child developed renal thrombosis in utero. It may be that a further genetic lesion at a third locus still remains to be defined. Alternatively, the intrauterine development of thrombosis in this infant could have been caused, at least in part by a transplacental thrombotic stimulus arising in the protein S-deficient maternal circulation. This analysis may, therefore, serve as a warning against extrapolating too readily from genotype to phenotype in families with a complex thrombotic disorder.
Female↗
The haemophilia A mutation search test and resource site, home page of the factor VIII mutation database: HAMSTeRS.
In order to facilitate easy access to and aid understanding of the causes of haemophilia A at the molecular level we have constructed HAMSTeRS, the third release of the factor VIII mutation database and the first release of this database that may be accessed and interrogated over the internet through a World Wide Web browser. The database also presents a review of the structure and function of factor VIII and the molecular genetics of haemophilia A, a real time update of the biostatistics of each parameter in the database, a molecular model of the A1, A2 and A3 domains of the factor VIII protein (based on the crystal structure of caeruloplasmin) and a bulletin board for discussion of issues in the molecular biology of factor VIII.
Databases, Factual↗
The tissue factor-factor VII complex: recent advances towards elucidating the structure and function of the initiator of haemostasis.
Localised extravascular initiation of mammalian haemostasis is achieved by the anatomical localization of tissue factor (TF) around blood vessels and at organ surfaces. TF avidly binds factor VII from outflowing blood following vascular injury, forming the tissue factor/factor VII (TF.VII) complex. This is rapidly converted to TF.VIIa, the active initiator of blood coagulation. Structural studies have revealed that the extracellular region of TF consists of two C2-type immunoglobulin-like modules rigidly held at an angle of 125 degrees by an extensive buried interface. The regions of TF involved in binding factor VII extend from the binding finger of the domain interface across quite a wide surface of the distal (N-terminal) module. Solution scattering studies show that both TF and factor VII adopt an extended conformation in solution and that the complex is formed by a compact side by side alignment of the two proteins along their long axes.
Amino Acid Sequence↗
High purity factor IX and prothrombin complex concentrate (PCC): pharmacokinetics and evidence that factor IXa is the thrombogenic trigger in PCC.
Recent studies using assays for surrogate markers of thrombogenicity in man have demonstrated that activation of the coagulation system occurs following infusion of clinical doses of prothrombin complex concentrates (PCC) but not after the same doses of high-purity factor IX concentrates (HP-FIX) in patients with haemophilia B. Here we have investigated the mechanism of such thrombogenesis by applying assays that detect early-through to late-events in coagulation system activation in a pharmacokinetic cross-over study of 50 IU/kg PCC and a new HP-FIX product in haemophilia B patients. Satisfactory recoveries and half-lives were observed for both concentrates. HP-FIX caused no increases in thrombin-antithrombin III complex (TAT), prothrombin activation peptide fragment F1+2 (F1+2), factor X activation peptide (FXAP) or factor VIIa (FVIIa). In contrast the same dose of factor IX in the form of PCC was followed by significant increases over pre-infusion levels of TAT, F1+2 and FXAP, but not FVIIa. Elevations of FIXAP occurred after both HP-FIX and PCC but did not reach normal levels and were attributed to normalisation of the FIX concentration in those patients whose levels of FIXAP were initially low. We conclude that the thrombogenic trigger associated with PCC infusion occurs at the level of factor X activation. In the absence of any increase in FVIIa, we would attribute this to the likely presence of FIXa in the PCC.
Adult↗
Factor VIIa and the extracellular domains of human tissue factor form a compact complex: a study by X-ray and neutron solution scattering.
The four-domain structure of human factor VIIa and the two-domain structure of tissue factor form a tight complex to initiate blood coagulation. By solution scattering, the mean X-ray and neutron radii of gyration RG (which determine macro-molecular elongation) were found to be 3.25 nm, 2.13 nm and 3.14 nm (+/- 0.13 nm) for factor VIIa, the extracellular region of tissue factor and their complex in that order. The mean cross-sectional radii of gyration RXS were 1.33 nm, 0.56 nm and 1.42 nm (+/- 0.13 nm) in that order. The mean lengths were 10.3 nm, 7.7 nm and 10.2 nm in that order. The data show that, in solution, the free proteins have extended domain structures, and the complex is formed by a compact side-by-side alignment of the two proteins along their long axes. The high binding affinity of tissue factor for factor VIIa may thus be accounted for by the occurrence of many intermolecular contacts in the complex.
Animals↗
Detection and characterization of seven novel protein S (PROS) gene lesions: evaluation of reverse transcript-polymerase chain reaction as a mutation screening strategy.
The molecular genetic analysis of protein S deficiency has been hampered by the complexity of the protein S (PROS) gene and by the existence of a homologous pseudogene. In an attempt to overcome these problems, a reverse transcript-polymerase chain reaction (RT-PCR) mutation screening procedure was developed. However, the application of this mRNA-based strategy to the detection of gene lesions causing heterozygous type I protein S deficiency appears limited owing to the high proportion of patients exhibiting absence of mRNA derived from the mutation-bearing allele ("allelic exclusion"). Nevertheless, this strategy remains extremely effective for rapid mutation detection in type II/III protein S deficiency. Using the RT-PCR technique, a G-to-A transition was detected at position +1 of the exon IV donor splice site, which was associated with type I deficiency and resulted in both exon skipping and cryptic splice site utilization. No abnormal protein S was detected in plasma from this patient. A missense mutation (Asn 217 to Ser), which may interfere with calcium binding, was also detected in exon VIII in a patient with type III protein S deficiency. A further three PROS gene lesions were detected in three patients with type I deficiency by direct sequencing of exon-containing genomic PCR fragments: a single base-pair (bp) deletion in exon XIV, a 2-bp deletion in exon VIII, and a G0to-A transition at position -1 of the exon X donor splice site all resulted in the absence of mRNA expressed from the disease allele. Thus, the RT-PCR methodology proved effective for further analysis of the resulting protein S-deficient phenotypes. A missense mutation (Met570 to Thr) in exon XIV of a further type III-deficient proband was subsequently detected in this patient's cDNA. No PROS gene abnormalities were found in the remaining four subjects, three of whom exhibited allelic exclusion. However, the father of one such patient exhibiting allelic exclusion was subsequently shown to carry a nonsense mutation (Gly448 to Term) within exon XII.
Base Sequence↗
Energetic contributions and topographical organization of ligand binding residues of tissue factor.
Tissue factor is the cellular receptor and macromolecular enzymatic cofactor for the serine protease coagulation factor VIIa. The ligand binding extracellular domain of tissue factor consists of two structural modules which fold similar to fibronectin type III modules, consistent with the classification of tissue factor as a member of the class 2 cytokine receptor family. On the basis of the three-dimensional structure, we here analyze the importance of tissue factor residues for binding of ligand by scanning alanine mutagenesis. The identified significant binding contacts account for as much as 80% of the calculated total free energy of ligand binding. Most residues with energetic contributions to ligand binding are well exposed to solvent, and the area for ligand interaction extends from the cleft formed by the two structural modules (residues Lys20, Ile22, Lys48, Asp58, Arg135, Phe140) to the convex-shaped edge of the three- and four-stranded sheets characterized by a patch of surface-exposed hydrophobic side chains in the amino-terminal module (residues Gln37, Asp44, Trp45, Phe76, Tyr78). The binding residues are dispersed over an extended surface area, indicating adaptation to the recognition of specific structural modules of the macromolecular ligand factor VIIa. This analysis provides detailed insight into the three-dimensional organization of the ligand docking structure of the initiating cofactor for the coagulation pathways.
Aspartic Acid↗
Six point mutations that cause factor XI deficiency.
We have identified six novel types of mutation that cause factor XI deficiency, an inherited bleeding disorder. Two are point mutations that interfere with the normal splicing of exons in the mRNA and four are point mutations that result in amino acid substitutions. One of these amino acid substitutions (Asp 16-->His) is near the amino terminal end of the protein. The other three amino acid substitutions (Leu 302-->Pro, Thr 304-->Ile, and Glu 323-->Lys) are in the fourth apple domain, a region that mediates dimerization of identical subunits of factor XI. All four amino acid substitutions cause a reduction in the amount of factor XI secreted from cells grown in vitro.
Amino Acid Sequence↗
Molecular etiology of factor VIII deficiency in hemophilia A.
Hemophilia is a common X-linked coagulation disorder due to deficiency of factor VIII. The factor VIII gene has been cloned in 1984 and a large number of mutations that cause hemophilia A have been identified in the last decade. The most common of the mutations is an inversion of factor VIII that accounts for nearly 45% of patients with severe hemophilia A. This review lists all the factor VIII mutations identified to date and briefly discusses their functional significance.
Amino Acid Sequence↗
Increase of activated factor VIIA and haemostatic molecular markers in juvenile chronic arthritis.
Activated factor VIIa (FVIIa), von Willebrand factor antigen (vWF:Ag), D-dimer and thrombin-antithrombin III complex (TAT) were measured to monitor coagulation status in patients with juvenile chronic arthritis (JCA). Subjects included 14 patients with systemic JCA, 16 with pauciarticular JCA and 16 with polyarticular JCA without disseminated intravascular coagulopathy, thrombosis or liver dysfunction. All types of JCA showed an increase of FVIIa, D-dimer and TAT, indicating enhanced activation of coagulation. In systemic JCA only there was also characteristically an elevation of vWF:Ag. We conclude that all types of JCA constitute a state of subclinical hypercoagulopathy caused by tissue damage and that additionally systemic JCA involves a prothrombotic state associated with or precipitated by vasculitis.
Antifibrinolytic Agents↗
Identification of two novel mutations in non-Jewish factor XI deficiency.
We have studied two heterozygous unrelated CRM- non-Jewish FXI-deficient patients. Neither of the patients carries a previously described mutation. Their FXI genes were screened by SSCP analysis following PCR amplification of each exon and the flanking intronic sequences. DNA fragments showing aberrant mobility were cloned and sequenced. The following mutations were identified: in case 1, a T to G transition in exon 12 results in the substitution of Phe-442 by Val (FXI-F442V); in case 2 a C to A transition in exon 5 results in the substitution of Cys-128 by a nonsense codon (FXI-C128X). The missense mutation results in a substitution within the protease domain of FXI. Molecular modelling locates this residue in a structurally conserved region of the protease domain and the amino acid substitution may therefore interfere with either chain folding and subsequent secretion or the stability of the protein in plasma. We conclude that the mutations which we have identified are responsible for the inherited abnormality in these patients.
Adult↗
Characterization of mutations within the factor VIII gene of 73 unrelated mild and moderate haemophiliacs.
To screen for mutations within the factor VIII gene of 101 patients (85 unrelated), we used denaturing gradient gel electrophoresis (DGGE) after DNA amplification of target regions, including all coding regions except for the middle part (amino acid 757 to amino acid 1649) of the B domain. With this method, missense mutations were identified in 86% of unrelated patients. 41 different mutations were identified: 25 of them have not been described previously. Five of the genotypes are associated with CRM+ and 26 with CRMred status. Patients who are definitely related to each other showed no differences in DNA sequence. One patient showed two different base pair alterations, the first at amino acid 469 [ala(GCA-->gly(GGA)] and the second at position 473 [tyr(TAT)-->cys(TGT)]. One patient with an amino acid change at position 1689 [arg(CGC)-->his(CAC)] has developed an inhibitor against factor VIII.
Base Sequence↗
Definition of the bleeding tendency in factor XI-deficient kindreds--a clinical and laboratory study.
Individuals with severe factor XI deficiency are prone to excessive bleeding after injury or surgery, but the existence of a haemorrhagic tendency in partial factor XI deficiency is controversial. In this study, 172 members of 30 kindreds (20 non-Jewish) transmitting factor XI deficiency in North West England were interviewed and a bleeding history questionnaire completed. Blood was taken for coagulation assays. The questionnaires were categorised independently by two assessors to determine presence or absence of a bleeding tendency, in the absence of information about the factor XI level or family history. Analysis shows that 48% of heterozygotes have a bleeding tendency. Eighteen (60%) families came to attention because of bleeding problems in heterozygotes. Comparison of histories between partially deficient and non-deficient individuals demonstrated a higher incidence of menstrual problems, an increase in significant bruising, and an increased likelihood of excessive bleeding after tonsillectomy and dental extractions. The incidence of von Willebrand's disease was not increased, but individuals with heterozygous factor XI deficiency who were bleeders tended to have lower levels of factor VIIIc and von Willebrand factor, and were more commonly of blood group 0. These features may contribute to the bleeding tendency. There was no evidence of alteration in factor VII activity (as defined by the ratio of activity to antigen) between the bleeders and non-bleeders. This is convincing evidence for abnormal bleeding in factor XI deficiency which is not confined to severely deficient patients.
Adolescent↗
Inherited factor VII deficiency: genetics and molecular pathology.
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Amino Acid Sequence↗
Gene therapy for the haemophilias.
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Factor IX↗