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Biomedical subjects

B Winchester

Publications and source records attributed to B Winchester.

At least 55 records · Page 3Linked to original sources
PubMed1998

Prenatal diagnosis of the carbohydrate-deficient glycoprotein syndrome type 1A (CDG1A) by a combination of enzymology and genetic linkage analysis after amniocentesis or chorionic villus sampling.

Two pregnancies at risk for the carbohydrate-deficient glycoprotein syndrome Type 1A (CDG1A, phosphomannomutase deficient) were monitored by enzyme and genetic linkage analyses. The index case in both families had a proven deficiency of phosphomannomutase (PMM). An unaffected fetus was predicted in family 1 following amniocentesis. Normal PMM activity was found in cultured amniotic fluid cells and there was no elevation of lysosomal enzymes in the amniotic fluid. Genetic linkage analysis using microsatellite markers closely linked to the CDG1A gene confirmed this prediction. A healthy child was born. In the second family direct assay of chorionic villi showed a profound deficiency of PMM and genetic linkage analysis showed the fetus to have the same haplotype as the proband. The pregnancy was terminated and a deficiency of PMM was confirmed in cultured fibroblasts from the fetus. Reliable prenatal diagnosis of CDG Type 1A (PMM-deficient) can be achieved by a combination of biochemical and molecular genetic tests.

Amniocentesis↗
PubMed1997

Prevention of lysosomal storage in Tay-Sachs mice treated with N-butyldeoxynojirimycin.

The glycosphingolipid (GSL) lysosomal storage diseases result from the inheritance of defects in the genes encoding the enzymes required for catabolism of GSLs within lysosomes. A strategy for the treatment of these diseases, based on an inhibitor of GSL biosynthesis N-butyldeoxynojirimycin, was evaluated in a mouse model of Tay-Sachs disease. When Tay-Sachs mice were treated with N-butyldeoxynojirimycin, the accumulation of GM2 in the brain was prevented, with the number of storage neurons and the quantity of ganglioside stored per cell markedly reduced. Thus, limiting the biosynthesis of the substrate (GM2) for the defective enzyme (beta-hexosaminidase A) prevents GSL accumulation and the neuropathology associated with its lysosomal storage.

1-Deoxynojirimycin↗
PubMed1997

Plasma chitotriosidase activity in Gaucher disease patients who have been treated either by bone marrow transplantation or by enzyme replacement therapy with alglucerase.

A marked elevation in plasma chitotriosidase (chitinase) activity has recently been observed in patients with Gaucher disease (Hollak et al 1994). It has been suggested that this phenomenon may serve as a useful marker for the efficacy of treatment with enzyme replacement therapy. We report our findings on the comparison of plasma chitotriosidase levels in 8 patients treated with the modified human placental enzyme alglucerase and 8 patients treated by allogeneic bone marrow transplantation (BMT). Two years after transplantation the activity in the BMT patients had fallen by over 93% and has continued to fall. Now, 5-12 years post-BMT, 6 patients have normal levels of plasma chitotriosidase and 2 patients have activities slightly above the reference range. Patients have been treated with alglucerase for a considerably shorter time than the BMT patients. The chitotriosidase activities have fallen and are continuing to fall in 7 patients but at a slower rate than in the BMT group. In one patient there has been no appreciable change in activity over the last 6 months, which would suggest that she may be on too low a dosage.

Adolescent↗
PubMed1997

A case of the carbohydrate-deficient glycoprotein syndrome type 1 (CDGS type 1) with normal phosphomannomutase activity.

The carbohydrate-deficient glycoprotein syndrome (CDGS) is a group of disorders characterized biochemically by abnormal glycosylation of serum and cellular glycoproteins. It has been classified into four forms on the basis of the isoelectric focusing pattern of serum transferrin and difference in clinical presentation. A deficiency of phosphomannomutase (PMM) has been reported in most patients with type 1. Seven of our eight CDGS patients, classified clinically as type 1, were shown to have a deficiency of phosphomannomutase in their fibroblast or lymphoblastoid cells (0.04-0.2 nmol/min per mg, compared with a control range of 1.0-2.1 nmol/min per mg). The eighth patient, who had many clinical features of the severe neonatal form of CDGS type 1, but lacked definite signs of CNS and ocular involvement, had a normal phosphomannomutase activity in his fibroblasts. There were approximately equal amounts of disialo- and tetrasialotransferrin and only a trace amount of asialotransferrin in the serum and ascitic fluid of this patient. The disialo- and tetrasialotransferrin isoforms were purified by ion-exchange chromatography and analysed by SDS-PAGE. The disialotransferrin had a lower molecular mass than the tetrasialotransferrin, consistent with the absence of an N-linked glycan. The N-linked glycans released enzymically from both isoforms consisted exclusively of disialylated biantennary chains, suggesting that disialotransferrin results from underglycosylation, as in the PMM-deficient CDGS type 1 patients. It is concluded that the clinical and biochemical phenotype in CDGS type 1 can result from more than one basic defect.

Ascitic Fluid↗
PubMed1997

A novel acid alpha-glucosidase mutation identified in a Pakistani family with glycogen storage disease type II.

A novel mutation, C118T, in exon 2 of the acid alpha-glucosidase gene has been found in an infant with glycogen storage disease type II. This mutation is predicted to result in protein truncation. The phenotype was that of the severe infantile form of the disorder with lack of motor development, but with eye regard, social smile and vocalization. The parents were heterozygous for C118T and belong to an Islamic community opposed to termination of pregnancy. As the C118T mutation results in the loss of one of two AvaI sites present in an informative PCR product, reliable premarriage carrier detection became possible and was acceptable to the members of this extended family.

Bronchopneumonia↗
PubMed1997

The effects of calystegines isolated from edible fruits and vegetables on mammalian liver glycosidases.

The polyhydroxylated nortropane alkaloids called calystegines occur in many plants of the Convolvulaceae, Solanaceae, and Moraceae families. Certain of these alkaloids exhibit potent inhibitory activities against glycosidases and the recently demonstrated occurrence of calystegines in the leaves, skins, and sprouts of potatoes (Solanum tuberosum), and in the leaves of the eggplant (S. melongena), has raised concerns regarding the safety of these vegetables in the human diet. We have surveyed the occurrence of calystegines in edible fruits and vegetables of the families Convolvulaceae, Solanaceae, and Moraceae by GC-MS. Calystegines A3, B1, B2, and C1 were detected in all the edible fruits and vegetables tested; sweet and chili peppers, potatoes, eggplants, tomatoes, Physalis fruits, sweet potatoes, and mulberries. Calystegines B1 and C1 were potent competitive inhibitors of the bovine, human, and rat beta-glucosidase activities, with Ki values of 150, 10, and 1.9 microM, respectively for B1 and 15, 1.5, and 1 microM, respectively, for C1. Calystegine B2 was a strong competitive inhibitor of the alpha-galactosidase activity in all the livers. Human beta-xylosidase was inhibited by all four nortropanes, with calystegine C1 having a Ki of 0.13 microM. Calystegines A3 and B2 selectively inhibited the rat liver beta-glucosidase activity. The potent inhibition of mammalian beta-glucosidase and alpha-galactosidase activities in vitro raises the possibility of toxicity in humans consuming large amounts of plants that contain these compounds.

Animals↗
PubMed1997

Bone marrow transplantation for mucopolysaccharidosis type I: experience of two British centres.

Bone marrow transplantation was carried out on 38 patients with mucopolysaccharidosis type I over a period of 15 years. The donor was an HLA identical relative in 10 cases, an HLA non-identical relative in 16 cases, and an HLA identical unrelated volunteer donor in 12 cases. Ten patients received a second transplant. One patient received three transplants. Thirteen engrafted patients have survived five years or more. Most patients have shown an arrest or slowing down of psychomotor regression. However, dysostosis multiplex has progressed. Careful selection of patients may be necessary to ensure optimum results.

Bone Marrow Transplantation↗
PubMed1997

Fucosidosis: genetic and biochemical analysis of eight cases.

The molecular basis of the deficiency of alpha-L-fucosidase has been investigated in eight patients who had been diagnosed clinically and enzymatically as suffering from the autosomal recessive lysosomal storage disease fucosidosis. None of the patients had a deletion or gross alteration of the alpha-L-fucosidase gene (FUCA1). Single strand conformation polymorphism (SSCP) analysis followed by direct sequencing of amplified exons and flanking regions identified putative disease causing mutations in six of the patients, who had severe forms of the disease and very low residual alpha-L-fucosidase activity and protein. They were a 10 bp deletion in exon 1 (E113fs), a 1 bp deletion at position -2 of intron 2 (S216fs), a g-->a transition at IVS5+1, point mutations W183X and N329Y in exons 3 and 6, respectively, and a compound allele consisting of a point mutation in the signal peptide in exon 1, P5R, and a 1 bp insertion in exon 6 (Y330fs). One patient in whom an SSCP change was not detected had residual alpha-L-fucosidase activity and cross reacting protein in the heterozygous range and normal metabolism of metabolites containing fucose in his fibroblasts, consistent with the low activity polymorphism. The eighth patient, who had a partial deficiency of alpha-L-fucosidase in her fibroblasts and leucocytes at a young age but normal alpha-L-fucosidase activity and protein at a later age, was homozygous for the common Q281R polymorphism in exon 5. She had no other sequence changes and Kivlin (Peters plus) syndrome has subsequently been diagnosed. The basis of her transient deficiency of alpha-L-fucosidase is not known. The detection of five novel mutations in six severely affected patients confirms the genetic heterogeneity in fucosidosis.

Adult↗
PubMed1996

The identification of abnormal glycoforms of serum transferrin in carbohydrate deficient glycoprotein syndrome type I by capillary zone electrophoresis.

One of the biochemical characteristics of carbohydrate deficient glycoprotein syndromes is the presence of abnormal glycoforms in serum transferrin. Both glycoform heterogeneity and variable site occupancy may, in principle, lead to the generation of a range of glycoforms which contain different numbers of sialic acid residues, and therefore variable amounts of negative charge. Capillary zone electrophoresis was used to resolve the glycoforms of normal human serum transferrin and also of a set of glycoforms which were prepared by digesting the sugars on the intact glycoprotein with sialidase. The sugars on the intact glycoprotein were also modified by a series of exoglycosidase enzymes to produce a series of neutral glycoforms which were-also analysed by capillary zone electrophoresis. The oligosaccharide population of human serum transferrin was analysed by a series of mixed exoglycosidase digests on the released glycan pool and quantified using a novel HPLC strategy. Transferrin was isolated from carbohydrate deficient glycoprotein syndromes type I serum and both the intact glycoforms and released sugars were resolved and quantified. The data presented here confirm the presence of a hexa-, penta- and tetra-sialoforms of human serum transferrin in both normal and carbohydrate deficient glycoprotein syndrome type I serum samples. Consistent with previous reports carbohydrate deficient glycoprotein syndrome type I transferrin also contained a di-sialoform, representing a glycoform in which one of the two N-glycosylation sites is unoccupied, and a non-glycosylated form where both remain unoccupied. This study demonstrates that capillary zone electrophoresis can be used to resolve quantitatively both sialylated and neutral complex type glycoforms, suggesting a rapid diagnostic test for the carbohydrate deficient glycoprotein syndromes group of diseases.

Carbohydrates↗
PubMed1996

A fluorimetric enzyme assay for the diagnosis of Sanfilippo disease type A (MPS IIIA).

4-Methylumbelliferyl-alpha-D-N-sulphoglucosaminide (MU-alpha-GlcNS) was synthesized and shown to be a substrate for the lysosomal heparin sulphamidase. Sanfilippo A patients' fibroblasts (n = 42) and lymphocytes (n = 1) showed 0-3% of mean normal heparin sulphamidase activity; in total leukocytes from patients (n = 8) sulphamidase activity was clearly deficient. In fibroblasts from obligate heterozygotes for Sanfilippo A, the sulphamidase activity was reduced in 9 out of 10 cases. Heparin sulphamidase desulphates MU-alpha GlcNS to MU-alpha GlcNH2 and further hydrolysis during a second incubation is required to liberate 4-methylumbelliferone, which can be measured. Yeast alpha-glucosidase, which has low but sufficient alpha-glucosaminidase activity, was used to hydrolyse the reaction intermediate MU-alpha GlcNH2 to release 4-methylumbelliferone and free glucosamine.

Clinical Enzyme Tests↗
PubMed1996

Single base deletion in exon 7 of the glycosylasparaginase gene causes a mild form of aspartylglycosaminuria in a patient of Mauritian origin.

Aspartylglycosaminuria (AGU) is a lysosomal storage disorder of glycoprotein degradation caused by deficiency of glycosylasparaginase (GA). A deletion mutation was found in a mildly affected AGU patient whose parents are first-cousins of Mauritian origin. One bp deletion at position 787 or 788 (delta T788) in exon 7 of the GA gene resulted in a frameshift and produced an immediate stop codon. The resulting truncated polypeptide was defective in its post-translational proteolytic processing and remained as a single chain (36 kDa) with no GA activity.

Acetylglucosamine↗
PubMed1996

Deoxyiminoalditols from aldonolactones--V. Preparation of the four stereoisomers of 1,5-dideoxy-1,5-iminopentitols. Evaluation of these iminopentitols and three 1,5-dideoxy-1,5-iminoheptitols as glycosidase inhibitors.

The four stereoisomeric 1,5-dideoxy-1,5-iminopentitols with D-arabino-(D-lyxo-) (3), ribo- (9), L-lyxo (L-arabino-) (13) and xylo-(18) configurations were synthesized. The corresponding aldonolactones (1, 7 and 11) or aldonic acid ester (15b) having a leaving group at C-5 gave by reaction with aqueous ammonia, the 5-amino-5-deoxy-1,5-lactams, 2, 8, 12 and 17, respectively. Reduction of the lactam function using sodium borohydride/acetic or trifluoroacetic acid, or borane dimethyl sulfide complex yielded the iminopentitols. The compounds 3, 9, 13 and 18, together with the three 1,5-dideoxy-1,5-iminoheptitols 19,20 and 21 were tested for inhibition of the glycosidase activities present in an extract from human liver. Compound 18 was a potent and 19 a moderately good inhibitor of beta-glucosidase. Compound 3 together with 19, 20 and 21, all having D-arabino-configuration at the hydroxy-substituted carbon atoms, were good inhibitors of alpha-L-fucosidase.

Enzyme Inhibitors↗
PubMed1996

Fabry disease: fourteen alpha-galactosidase A mutations in unrelated families from the United Kingdom and other European countries.

The nature of the molecular lesions in the alpha-galactosidase A gene causing Fabry disease in 12 unrelated families from the United Kingdom and 4 from other European countries was determined in order to provide precise heterozygote detection and prenatal diagnosis for these families. The entire alpha-galactosidase A coding region and flanking intronic sequences were analyzed by amplification of genomic DNA and solid-phase direct sequencing or by SSCP analysis followed by solid-phase direct sequencing. Fourteen new mutations were identified including 10 missense mutations (M42V, R49S, C56Y, D92H, D93G, P205T, W236C, W287G, N298H, and W340R), 2 nonsense mutations (Q107X and Q119X) and 2 small deletions (257del18 and 1087del1). Together with the previously reported mutations, a total of 33 lesions in the alpha-galactosidase A gene have been identified in unrelated British families, further documenting the molecular genetic heterogeneity of this disease.

Fabry Disease↗
PubMed1995

Pseudodeficiency of arylsulphatase A: strategy for clarification of genotype in families of subjects with low ASA activity and neurological symptoms.

A benign deficiency (pseudodeficiency) of the lysosomal enzyme arylsulphatase A (ASA) (EC 3.1.6.8) towards synthetic substrates complicates the diagnosis of metachromatic leukodystrophy (MLD). The pseudodeficiency is due to a single base substitution in the 3'-untranslated region of the ASA gene (1524+95 A-->G) and it has been reported that this mutation (PD2) always occurs on a chromosome carrying a second mutation in the ASA gene (PD1), which abolishes an N-glycosylation site (N350S). Analysis of the two PD mutations in the ASA gene separately was carried out in a large group of subjects with neurological symptoms and low ASA activity, including close relatives and MLD patients. The relationship between ASA enzyme activity and the different genotypes identified is presented. Evidence for the existence of an allele containing the PD2 mutation alone is presented. A strategy for cases with low ASA activity and neurological symptoms in families carrying a PD allele or both PD and MLD alleles is proposed.

Cerebroside-Sulfatase↗
PubMed1995

Allogeneic bone marrow transplantation for fucosidosis.

Bone marrow transplantation was performed on an 8-month-old boy who was diagnosed as having fucosidosis following the diagnosis of the disease in his older brother. Although he was asymptomatic and his development was normal, abnomalities were found on an MRI scan prior to transplant. In the absence of a suitable related donor, an unrelated volunteer donor was used. Conditioning for the transplant consisted of busulphan and cyclophosphamide. Graft-versus-host disease prophylaxis consisted of in vitro T cell-depletion of the bone marrow and in vivo administration of cyclosporin. The post-transplant period was complicated by moderately severe graft-versus-host disease. Engraftment was documented by the presence of donor levels of alpha-fucosidase, donor blood group and tissue type (difference in the DQ antigen), and chromosomal polymorphism pattern of donor origin. Eighteen months after transplant, there is evidence of mild neurodevelopmental delay. By contrast, his elder sibling showed far greater developmental delay at the same age. The patient's MRI scan shows improvement. We believe this to be the first case of human fucosidosis treated by bone marrow transplantation.

Bone Marrow Transplantation↗
PubMed1995

A specific protein found in human senescent fibroblasts.

The limited potential for proliferation of human fibroblasts in culture represents cell level senescence. Aging is a programmed process under genetic control, and at certain stage of the life-span of animal cells, some genes start to express. Studying the biomarkers of senescent cells is important to understanding the basic mechanism of aging which may be relevant to the normal cell growth control and tumor biology. A protein with 72,000 Dalton molecular weight was detected by the hybridoma method in our laboratory. The protein shows specificity to senescent or presenescent cells of several cell lines, including WI-38, K.D., U2OS, etc.

Animals↗