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Biomedical subjects

M S Enayat

Publications and source records attributed to M S Enayat.

At least 19 recordsLinked to original sources
PubMed2004

Minimizing factor requirements for surgery without increased risk.

The most common type of surgical procedure in patients with haemophilia is orthopaedic surgery on the lower limb. These procedures are of great importance for the improvement of the function and quality of life of patients with haemophilia, but in many countries resources are lacking to provide the factor concentrates needed for these operations. We have attempted to reduce the level of replacement during and after total hip or total knee replacement without jeopardizing the outcome of the procedures. The results from 30 operations in 27 patients are presented. Steady-state levels down to 0.5 IU mL(-1) did not appear to increase the amount of blood loss or cause an increased incidence of complications, compared with higher levels. In addition, we review the possibilities to reduce the amount of factor concentrate by performing a combination of orthopaedic procedures and using local haemostatic agents or antifibrinolytic agents.

Aged↗
PubMed2001

Aberrant dimerization of von Willebrand factor as the result of mutations in the carboxy-terminal region: identification of 3 mutations in members of 3 different families with type 2A (phenotype IID) von Willebrand disease.

The 3' end of the VWF gene was screened in the affected members of 3 different families with type 2A (phenotype IID) von Willebrand disease (vWD). Exons 49 to 52 of the VWF gene were amplified and screened for mutations by chemical cleavage mismatch detection. Mismatched bands were detected in exon 52 of 2 patients and in exon 51 of a third patient. Using direct DNA sequencing, a heterozygous G8562A transition leading to a Cys2008Tyr substitution was found in all the patients in family 1, and a T8561A transversion leading to a Cys2008Ser substitution was found in both patients from family 2. In a patient from a third family, an 8-base deletion from nucleotide 8437 to 8444 was identified in exon 51. The 2 mutations in exon 52 were reproduced by in vitro site-directed mutagenesis of full-length von Willebrand factor (vWF) cDNA and transiently expressed in COS-7 cells. The corresponding recombinant VWFs for these 2 mutations exhibited the typical aberrant vWF:Ag multimer pattern seen in the plasma of the patients. These 3 mutations demonstrate the importance of other carboxy-terminal cysteines in addition to the reported Cys2010 residue, in the normal dimerization of vWF, and their essential role in the assembly of normal multimeric vWF. (Blood. 2001;98:674-680)

Animals↗
PubMed2001

Site and type of mutations in the factor VIII gene in patients and carriers of haemophilia A.

Haemophilia A is an X-linked bleeding disorder caused by reduced or absent FVIII (FVIII) protein caused by mutations in the FVIII gene. We have used Southern blotting and chemical mismatch analysis (CMA) to identify the mutations causing haemophilia A in 59 local or referred patients or carriers of haemophilia A. Southern blot analysis of 87 families with FVIII : C < 5% identified 31 as positive for the intron 22 inversion. Analysis of 19 of the inversion-negative families and a further nine families with mild or moderate haemophilia A by CMA resulted in the identification of a heterogeneous spectrum of mutations in the FVIII gene comprising 21 single base-pair substitutions and nine deletions. Seventeen of the base-pair substitutions are missense, two nonsense, and two are splice-site mutations. Two patients were found to have compound mutations with two mutations identified on a single X chromosome. Six of the point mutations and six of the deletions have not been reported previously in the haemophilia A mutation database. Unusually, a missense mutation, as well as deletion and splice-site mutations, was found to be associated with exon-skipping events.

Factor VIII↗
PubMed2000

A new candidate missense mutation (Leu 1657 IIe) in an apparently asymptomatic type 2A (phenotype IIA) von Willebrand disease family.

Type 2A von Willebrand disease (VWD) is mostly an autosomal dominantly inherited bleeding disorder characterised by a qualitative defect of von Willebrand factor (VWF). Mutation screening was used to screen the whole of VWF gene followed by direct sequencing to detect the mutation in a father and son diagnosed with type 2A (phenotype IIA) von Willebrand disease. A C5219 to A transversion was detected predicting Leucine to Isoleucine substitution in codon 1657. This novel missense mutation which was also identified by MboI restriction enzyme analysis, was found in both patient and his father but not in any other unaffected family member or 50 unrelated normal individuals. This substitution was reproduced by in vitro site directed mutagenesis of full-length VWF cDNA and transiently expressed in COS-7 cells. The corresponding recombinant VWF protein exhibited the full spectrum of VWF multimers, suggesting that the abnormal multimer seen in the patient results from increased proteolysis.

Amino Acid Substitution↗
PubMed2000

Identification of type 2 von Willebrand disease in previously diagnosed type 1 patients: a reappraisal using phenotypes, genotypes and molecular modelling.

In order to investigate the possibility that qualitative type 2 defects in von Willebrand factor (VWF) occurred in patients previously diagnosed with quantitative type 1 von Willebrand disease (VWD), the phenotypes and genotypes were reanalysed in 30 patients who exhibited discrepant VWF activity/VWF:Ag ratios of less than 0.7. The capacity of VWF to bind to glycoprotein Ib (GpIb) was reassessed using the ristocetin co-factor activity (VWF:RiCo) assay compared to an in-house and a commercial ELISA assay (based on a mAb directed against the GpIb binding site on VWF). This was supplemented by multimeric analysis and the amplification and sequencing of a 936 bp fragment of exon 28 of the VWF gene with the aim of identifying mutations in the A1 domain. On reappraisal, using the VWF:RiCo assay all patients demonstrated a disproportionately reduced VWF:RiCo/VWF:Ag ratio, indicative of a qualitative defect, while abnormal ratios were detected in only seven kindreds using the in-house ELISA assay and in only one kindred with the commercial ELISA assay. Eight single amino acid substitutions were found in nine kindreds, four of which were novel candidate VWF mutations and four previously described in association with type 2 VWD. In agreement with the phenotype, the novel VWF mutations were located in the VWF-A1 crystal structure at positions that corresponded to potential type 2M defects. This study underlines the difficulties of correct diagnosis of the subtype of VWD and emphasises the importance of using sensitive phenotypic assays, the relevance of the VWF:RiCo/ VWF:Ag ratio, multimeric analysis and molecular modelling analysis.

Amino Acid Substitution↗
PubMed1999

Congenital thrombophilia and thrombosis: a study in a single centre.

AIM: To identify the incidence of congenital thrombophilia in a cohort of children presenting with symptomatic thromboembolism. METHOD: A review of children with thromboembolism investigated for thrombophilia over a 12 month period. SUBJECTS: Thirty children with thromboembolic episodes and 16 of their family members. MEASUREMENTS AND DATA COLLECTION: Data were collected on age at diagnosis, underlying diagnosis, site of thrombosis, associated precipitating factors, occurrence of other thromboembolic events, and family history. Investigations included measurement of protein C activity, total and free protein S antigen, antithrombin III activity, screening for factor V Leiden and prothrombin 20210A, urinary homocysteine estimation, and a screen for lupus anticoagulant. RESULTS: Twenty seven of 30 patients had one or more risk factors present at the time of thromboembolism. Eighty three per cent had acquired precipitating factors present, and 43% had underlying congenital thrombophilia. CONCLUSIONS: There was a high incidence of congenital thrombophilia in this group of patients with symptomatic thromboembolism. These findings emphasise the importance of such defects in the pathogenesis of childhood thrombosis, and suggest that full thrombophilia investigations should be performed on all children presenting with thromboembolic disease.

Adolescent↗
PubMed1998

A novel mutation in exon b (R259C) of the MTM1 gene is associated with a mild myotubular myopathy. Mutation in brief no. 125. Online.

The genetic basis of the relatively mild myopathic symptoms exhibited in a male was investigated. Mutation screening of a candidate gene, MTM1, represented a chance of establishing the molecular defect and the mode of inheritance. SSCA detected variation of the exon b PCR products from the proband and his mother, compared to that observed upon analysis of the PCR products from other members of the family and 159 unrelated X chromosomes. Sequencing revealed a C775 to T transition, in the proband and his mother, but not in his unaffected brother. To confirm the presence of a base change in this region, a Cfol site was introduced into the PCR product of the wildtype allele by using the forward primer 5'-AGAAAATAAGACGGTCATTGcG-3' (mismatch base in small font) with the exon b reverse primer as used by Laporte et al (1996). Analysis of DNA from other members of the family using this method revealed that this is a new mutation in the proband's mother. This mutation would result in a Arg259->Cys substitution.

Alleles↗
PubMed1990

Qualitative and quantitative abnormalities of von Willebrand antigen in patients with diabetes mellitus.

Previous reported studies of vVWf antigen (vWf:Ag) in patients with diabetes mellitus have not shown qualitative abnormalities despite frequent documentation of raised vWf:Ag. Twenty two patients with established diabetes mellitus have been studied and qualitative abnormalities of vWf:Ag multimers were found in 10 patients who had poorer glycaemic control (as judged by plasma fructosamine) than the other 12 patients. Seven of the 22 patients were restudied after a 3 month period of improved glycaemic control and the vWf:Ag multimer abnormalities disappeared in 6 of these. Some of the structural abnormalities were accompanied by loss of vWf functional activity. As vWf:Ag is synthesised in the endothelial cell, vWf:Ag abnormalities during periods of poor glycaemic control may reflect endothelial cell damage with vWf:Ag release and possibly subsequent proteolysis.

Antigens↗
PubMed1990

Clinical and laboratory evaluation of the treatment of von Willebrand's disease patients with heat-treated factor VIII concentrate (BPL 8Y).

Adequate rises of vWF:activity and satisfactory haemostasis have been obtained in six von Willebrand patients treated with heated 8Y factor VIII concentrate, despite multimeric analysis showing the concentrate to have an alteration in the proportions of the different vWF:Ag multimers including slight loss of the highest molecular weight multimers and an abnormal triplet. As heat-treated concentrates reduce the risk of transmitting blood borne viruses 8Y concentrate should be considered as a possible first line treatment of vWD patients in the U.K. if they do not have mild forms of type I von Willebrand's disease which can be treated with desmopressin (DDAVP) infusions.

Adolescent↗
PubMed1989

Vascular endothelial cell function and ultrastructure in thrombotic microangiopathy following allogeneic bone marrow transplantation.

We report studies on vascular endothelial function and ultrastructure in 2 cases of fatal cyclosporin (CS)-associated thrombotic microangiography following allogeneic bone marrow transplantation (BMT). Spontaneous vascular release of prostacyclin (PGI2) from a vein sample ex vivo was absent, and scanning electron microscopy (SEM) showed surface changes indicative of vascular endothelial damage (case 1). PGI2 release from cultured human umbilical vein endothelial cells incubated with patients' serum in vitro was normal in both cases. Plasma von Willebrand factor (vWF) antigen and ristocetin cofactor activity levels were raised in both patients, 5.06 and 7.02 (case 1) and 3.60 and 2.01 (case 2) (normal ranges 0.59-1.57 and 0.42-1.74 U/ml), respectively, but multimer patterns were normal. The SEM appearances coupled with the absent PGI2 release and raised vWF levels suggest that vascular endothelial damage is central to the pathogenic process in thrombotic microangiopathy following allogeneic BMT but the mechanisms appear to be distinct from those in the haemolytic uraemic syndrome and de novo thrombotic thrombocytopenic purpura. The precise role of CS in this process remains to be identified.

Adult↗
PubMed1987

Evaluation of monoclonal antibodies to vWf antigen for use in autoradiographing vWf multimer analysis.

Nine monoclonal antibodies (MAb, coded ESvWF 1-5, 7-10) to human von Willebrand's factor (vWf) have been studied for their labelling characteristics with 125I and their ability to demonstrate vWf multimers by autoradiography after discontinuous SDS electrophoresis on agarose and agarose/acrylamide gels in plasma from normal and von Willebrand's disease (vWD) patients. Of ESvWF 1, 2, 4, 7, 8 and 9, ESvWF 2 gave autoradiographs most similar to those obtained with a polyclonal antibody. The others gave much fainter staining of all bands suggesting that the epitope detected by ESvWF 2 is either more common or, less easily altered by SDS than the epitopes identified by the other MAb's. ESvWF 2 gave a different staining pattern with a IIC vWD patient than that obtained using a polyclonal antibody. The most striking result, however, was shown by ESvWF 3, 5 and 10 which failed to stain the lower band of the lower molecular weight multimer. This suggests that the lower triplet band, unlike the central and upper bands of the triplet, does not contain the epitope identified by these three MAb's. With these reagents it has been possible to show, for the first time, that epitope differences do exist between the constituent bands of the vWf triplet.

Antibodies, Monoclonal↗