21 results on '"Keulemans JL"'
Search Results
2. A new diagnostic assay for glycogen storage disease type II in mixed leukocytes.
- Author
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Okumiya T, Keulemans JL, Kroos MA, Van der Beek NM, Boer MA, Takeuchi H, Van Diggelen OP, and Reuser AJ
- Subjects
- Acarbose, Glucan 1,4-alpha-Glucosidase antagonists & inhibitors, Humans, Hydrogen-Ion Concentration, Infant, alpha-Glucosidases, Glucan 1,4-alpha-Glucosidase deficiency, Glycogen Storage Disease Type II diagnosis, Leukocytes chemistry
- Abstract
We have established a new method for the enzymatic diagnosis of glycogen storage disease type II (Pompe disease or acid maltase deficiency) using mixed leukocytes. The method employs glycogen and 4-methylumbelliferyl-alpha-D-glucopyranoside (4MU-alphaGlc) as substrates for measuring the lysosomal acid alpha-glucosidase (acid alphaGlu) activity, and incorporates acarbose to eliminate the interference of unrelated alpha-glucosidases (predominantly maltase-glucoamylase). It is shown that 3.0 micromol/L acarbose completely inhibits the maltase-glucoamylase activity at pH 4.0, but the lysosomal acid alphaGlu activity by less than 5%. With this method, we determined the acid alphaGlu activity in mixed leukocytes from 25 patients with glycogen storage disease type II (2 infantile and 23 late-onset cases), one GAA2/GAA2 homozygote and 30 healthy subjects. In the assay with glycogen as substrate, the addition of acarbose created a clear separation between the patient and the control ranges. In the assay with 4MU-alphaGlc as substrate, the two ranges were fully separated but remained very close despite the use of acarbose. The separation of the patient and normal ranges was improved considerably by taking the ratio of acarbose-inhibited over uninhibited activity. A GAA2/GAA2 homozygote was correctly diagnosed with 4MU-alphaGlc but misdiagnosed as patient when glycogen was used as substrate. We conclude that the inclusion of 3.0 micromol/L acarbose in the assays with glycogen and 4MU-alphaGlc substrates at pH 4.0 allows for the specific measurement of lysosomal acid alphaGlu activity in mixed leukocytes, thus enabling a reliable diagnosis of glycogen storage disease type II in this specimen.
- Published
- 2006
- Full Text
- View/download PDF
3. A new fluorimetric enzyme assay for the diagnosis of Niemann-Pick A/B, with specificity of natural sphingomyelinase substrate.
- Author
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van Diggelen OP, Voznyi YV, Keulemans JL, Schoonderwoerd K, Ledvinova J, Mengel E, Zschiesche M, Santer R, and Harzer K
- Subjects
- Ceramides chemistry, Clinical Enzyme Tests, Diagnosis, Differential, Dose-Response Relationship, Drug, Fibroblasts enzymology, Fibroblasts metabolism, Gene Expression Regulation, Enzymologic, Hexosaminidases chemistry, Humans, Hydrolysis, Leukocytes enzymology, Leukocytes metabolism, Mutation, Niemann-Pick Diseases enzymology, Phospholipid Ethers chemistry, Phosphorylcholine analogs & derivatives, Phosphorylcholine chemistry, Protein Binding, Reproducibility of Results, Skin metabolism, Sphingomyelins chemistry, Sphingosine analogs & derivatives, Sphingosine chemistry, Substrate Specificity, Time Factors, Blood Chemical Analysis methods, Chemistry, Clinical methods, Fluorometry methods, Niemann-Pick Diseases diagnosis, Sphingomyelin Phosphodiesterase chemistry
- Abstract
6-Hexadecanoylamino-4-methylumbelliferylphosphorylcholine (HMUPC) was shown to be a specific substrate for the determination of acid (lysosomal) sphingomyelinase (ASM; gene SMPD1). Fibroblasts (n = 27) and leukocytes (n = 8) from both the A and B types of Niemann-Pick disease showed < 6% and < 10% of mean normal ASM activity, respectively. Niemann-Pick A or B patients bearing the Q292K mutation had apparently normal ASM activity with our new artificial substrate. These patients with false-normal sphingomyelinase activity, however, could readily be detected by determining the extent of inhibition of enzymatic hydrolysis of the artificial substrate HMU-PC by an unlabelled natural substrate, in particular lysosphingomyelin. This approach is generally applicable. Our novel assay for ASM combines the ease of a rapid and robust enzyme assay using a fluorogenic substrate with the specificity of an ASM assay using a natural substrate. Such assays are obviously more convenient to the diagnostic laboratory, since radiolabelled substrates are not required.
- Published
- 2005
- Full Text
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4. Prenatal diagnosis of the Hunter syndrome and the introduction of a new fluorimetric enzyme assay.
- Author
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Keulemans JL, Sinigerska I, Garritsen VH, Huijmans JG, Voznyi YV, van Diggelen OP, and Kleijer WJ
- Subjects
- Adult, Amniotic Fluid cytology, Amniotic Fluid metabolism, Cells, Cultured, Electrophoresis, Gel, Two-Dimensional, Female, Fluorometry, Glycosaminoglycans metabolism, Heterozygote, Humans, Male, Pregnancy, Pregnancy, High-Risk, Prospective Studies, Retrospective Studies, Amniocentesis, Chorionic Villi Sampling, Iduronate Sulfatase analysis, Mucopolysaccharidosis II diagnosis, Mucopolysaccharidosis II enzymology
- Abstract
Prenatal diagnosis of the Hunter syndrome (mucopolysaccharidosis type II; MPS II) is preferably achieved by the assay of iduronate-2-sulphate sulphatase (IDS) in uncultured chorionic villi (CV) as this allows early (12th week), rapid (2-3 days) and reliable results. We summarize the results of 174 prenatal analyses in the past 30 years, using various methods such as radiolabelled sulphate incorporation in amniotic fluid (AF) cells, glycosaminoglycan (GAG)-electrophoresis in AF and IDS assay in CV, CV-cells, AF and AF-cells. Twenty-seven fetuses with MPS II were diagnosed after finding clearly abnormal results in pregnancies with a male fetus; very low IDS activity has also been measured in some pregnancies with a (heterozygous) female fetus, emphasizing the need to combine enzyme assay with fetal sex determination. IDS activity has until recently been assessed by a cumbersome radioactive enzyme assay. Here we describe the use of a novel fluorigenic 4-methylumbelliferyl substrate, which allows a sensitive, rapid and convenient assay of IDS activity and reliable early prenatal diagnosis. This novel IDS assay was validated in retrospective analyses of 14 CV, CV-cell, AF and AF-cell samples from affected pregnancies in addition to prospective prenatal diagnosis in eight pregnancies at risk with one MPS II-affected fetus., (Copyright 2002 John Wiley & Sons, Ltd.)
- Published
- 2002
- Full Text
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5. A fluorimetric enzyme assay for the diagnosis of MPS II (Hunter disease).
- Author
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Voznyi YV, Keulemans JL, and van Diggelen OP
- Subjects
- Fibroblasts enzymology, Fluorometry, Humans, Hydrogen-Ion Concentration, Hymecromone analogs & derivatives, Hymecromone metabolism, Iduronate Sulfatase blood, Iduronate Sulfatase metabolism, Iduronic Acid analogs & derivatives, Iduronic Acid metabolism, Leukocytes enzymology, Lysosomes enzymology, Substrate Specificity, Mucopolysaccharidosis II diagnosis, Mucopolysaccharidosis II enzymology
- Abstract
4-Methylumbelliferyl-alpha-iduronate 2-sulphate was synthesized and shown to be a specific substrate for the lysosomal iduronate-2-sulphate sulphatase (IDS). Fibroblasts (n = 17), leukocytes (n = 3) and plasmas (n = 9) from different MPS II patients showed < 5% of mean normal IDS activity. The enzymatic liberation of the fluorochrome from 4-methylumbelliferyl-alpha-iduronate 2-sulphate requires the sequential action of IDS and alpha-iduronidase. A normal level of alpha-iduronidase activity was insufficient to complete the hydrolysis of the reaction intermediate 4-methylumbelliferyl-alpha-iduronide formed by IDS. A second incubation step in the presence of excess purified alpha-iduronidase is needed to avoid underestimation of the IDS activity.
- Published
- 2001
- Full Text
- View/download PDF
6. Adult neuronal ceroid lipofuscinosis with palmitoyl-protein thioesterase deficiency: first adult-onset patients of a childhood disease.
- Author
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van Diggelen OP, Thobois S, Tilikete C, Zabot MT, Keulemans JL, van Bunderen PA, Taschner PE, Losekoot M, and Voznyi YV
- Subjects
- Adult, Age of Onset, Female, Fluorometry methods, Humans, Middle Aged, Neuronal Ceroid-Lipofuscinoses metabolism, Thiolester Hydrolases deficiency
- Abstract
The fluorogenic enzyme assay for palmitoyl-protein thioesterase (PPT) has greatly facilitated the diagnosis of infantile neuronal ceroid lipofuscinosis (Santavuori-Haltia disease) and the search for possible new variants with atypical clinical presentation. Here, we present the first cases of adult neuronal ceroid lipofuscinosis with onset in the fourth decade of life due to a profound deficiency of PPT. The causative mutations in the CLN1 gene were the known, deleterious mutation R151X and the novel missense mutation G108R. Patients presented at onset (31 and 38 years), with psychiatric symptoms only. At present (ages 56 and 54 years), visual, verbal, and cognitive losses have progressed and both patients have cerebellar ataxia and cannot walk without support.
- Published
- 2001
- Full Text
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7. Human alpha-N-acetylgalactosaminidase (alpha-NAGA) deficiency: no association with neuroaxonal dystrophy?
- Author
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Bakker HD, de Sonnaville ML, Vreken P, Abeling NG, Groener JE, Keulemans JL, and van Diggelen OP
- Subjects
- Cells, Cultured, Child, Child, Preschool, DNA Mutational Analysis, Fibroblasts enzymology, Fibroblasts pathology, Genotype, Hexosaminidases genetics, Humans, Male, Mutation, Neuroaxonal Dystrophies genetics, Oligosaccharides analysis, Pedigree, Phenotype, Polymerase Chain Reaction, Skin enzymology, alpha-N-Acetylgalactosaminidase, Hexosaminidases deficiency, Neuroaxonal Dystrophies enzymology
- Abstract
Two new individuals with alpha-NAGA deficiency are presented. The index patient, 3 years old, has congenital cataract, slight motor retardation and secondary demyelinisation. Screening of his sibs revealed an alpha-NAGA deficiency in his 7-year-old healthy brother who had no clinical or neurological symptoms. Both sibs are homozygous for the E325K mutation, the same genotype that was found in the most severe form of alpha-NAGA deficiency presenting as infantile neuroaxonal dystrophy. Thus, at the age of 7 years the same genotype of alpha-NAGA may present as a 'non-disease' (present healthy case) and can be associated with the vegetative state (the first two patients described with alpha-NAGA deficiency). The clinical heterogeneity among the 11 known individuals with alpha-NAGA deficiency is extreme, with a 'non-disease' (two cases) and infantile neuroaxonal dystrophy (two cases) at the opposite sides of the clinical spectrum. The broad spectrum is completed by a very heterogeneous group of patients with various degrees of epilepsy/behavioural difficulties/psychomotor retardation (four patients) and a mild phenotype in adults without overt neurological manifestations who have angiokeratoma and clear vacuolisation in various cell types (three cases). These observations are difficult to reconcile with a straightforward genotype-phenotype correlation and suggest that factors or genes other than alpha-NAGA contribute to the clinical heterogeneity of the 11 patients with alpha-NAGA deficiency.
- Published
- 2001
- Full Text
- View/download PDF
8. First-trimester diagnosis of late-infantile neuronal ceroid lipofuscinosis (LINCL) by tripeptidyl peptidase I assay and CLN2 mutation analysis.
- Author
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Kleijer WJ, van Diggelen OP, Keulemans JL, Losekoot M, Garritsen VH, Stroink H, Majoor-Krakauer D, Franken PF, Eurlings MC, Taschner PE, Los FJ, and Galjaard RJ
- Subjects
- Aminopeptidases, Chorionic Villi enzymology, Chorionic Villi Sampling, Dipeptidyl-Peptidases and Tripeptidyl-Peptidases, Female, Humans, Neuronal Ceroid-Lipofuscinoses enzymology, Neuronal Ceroid-Lipofuscinoses genetics, Pregnancy, Pregnancy Trimester, First, Tripeptidyl-Peptidase 1, DNA Mutational Analysis, Endopeptidases deficiency, Endopeptidases genetics, Neuronal Ceroid-Lipofuscinoses diagnosis, Prenatal Diagnosis
- Abstract
Late-infantile neuronal ceroid lipofuscinosis (LINCL) is a progressive neurodegenerative disorder caused by the deficiency of lysosomal tripeptidyl peptidase I (TPP-I) encoded by the CLN2 gene. We report the first case of early prenatal diagnosis of LINCL by combined enzyme and mutation analysis. TPP-I activity in chorionic villi (CV) was less than 2% of the mean normal control level and g.1946A > G and g.3670C > T mutations were demonstrated, as in the two previously affected children. After termination of pregnancy, TPP-I deficiency was confirmed in cultured CV cells and in the fetal skin fibroblasts. The expression of unequivocal TPP-I deficiency in CV demonstrates that enzyme assay is a reliable option for prenatal diagnosis of LINCL., (Copyright 2001 John Wiley & Sons, Ltd.)
- Published
- 2001
- Full Text
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9. Pre- and postnatal enzyme analysis for infantile, late infantile and adult neuronal ceroid lipofuscinosis (CLN1 and CLN2).
- Author
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Van Diggelen OP, Keulemans JL, Kleijer WJ, Thobois S, Tilikete C, and Voznyi YV
- Subjects
- Adult, Aminopeptidases, Cells, Cultured, Chorionic Villi Sampling, Dipeptidyl-Peptidases and Tripeptidyl-Peptidases, Endopeptidases metabolism, Female, Fibroblasts cytology, Fibroblasts enzymology, Humans, Leukocytes enzymology, Middle Aged, Pregnancy, Serine Proteases, Thiolester Hydrolases metabolism, Tripeptidyl-Peptidase 1, Endopeptidases deficiency, Neuronal Ceroid-Lipofuscinoses diagnosis, Neuronal Ceroid-Lipofuscinoses enzymology, Thiolester Hydrolases deficiency
- Abstract
The recent development of simple, fluorogenic enzyme assays for infantile and late infantile neuronal ceroid lipofuscinosis (INCL and LINCL; CLN1 and CLN2) has greatly facilitated the diagnostic process for these diseases. In leucocytes and fibroblasts from INCL (n = 38) patients we found profound deficiencies of palmitoyl-protein thioesterase I (PPT1), the residual activity was < 5% of mean control activity. In fibroblasts from LINCL patients we found a similar deficiency of tripeptidyl-peptidase I activity (TPP-I), with < 2% activity in 16 patients. The residual TPP-I activity in leucocytes from LINCL patients seemed substantially higher. We also showed the feasibility of reliable prenatal enzyme analysis. In five first-trimester and two second-trimester prenatal analyses for INCL, four affected foetuses were detected (PPT activity 3-6%). Two first trimester pregnancies at risk for LINCL were analysed and a clear TPP-I deficiency was detected in both cases (TPP-I activity 3-4%). The first patient with adult neuronal ceroid lipofuscinosis (ANCL) due to a deficiency of PPT is presented; her present age is 53 years and the onset of the disease was at 38 years with psychiatric symptoms.
- Published
- 2001
- Full Text
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10. A new simple enzyme assay for pre- and postnatal diagnosis of infantile neuronal ceroid lipofuscinosis (INCL) and its variants.
- Author
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Voznyi YV, Keulemans JL, Mancini GM, Catsman-Berrevoets CE, Young E, Winchester B, Kleijer WJ, and van Diggelen OP
- Subjects
- Female, Fluorescent Dyes, Humans, Hymecromone, Infant, Infant, Newborn, Pregnancy, Substrate Specificity, Fluorometry methods, Neuronal Ceroid-Lipofuscinoses diagnosis, Neuronal Ceroid-Lipofuscinoses enzymology, Prenatal Diagnosis, Thiolester Hydrolases analysis, Thiolester Hydrolases deficiency
- Abstract
Palmitoyl-protein thioesterase (PPT) deficiency was recently shown to be the primary defect in infantile neuronal ceroid lipofuscinosis (INCL). The available enzyme assay is complicated and impractical for diagnostic use and is, in practice, unavailable. We have developed a new fluorimetric assay for PPT based on the sensitive fluorochrome 4-methylumbelliferone. This PPT assay is simple, sensitive, and robust and will facilitate the definition of the full clinical spectrum associated with a deficiency of PPT. PPT activity was readily detectable in fibroblasts, leucocytes, lymphoblasts, amniotic fluid cells, and chorionic villi, but was profoundly deficient in these tissues from INCL patients. Similarly, a deficiency of PPT was shown in patients with the variant juvenile NCL with GROD. These results show that rapid pre- and postnatal diagnosis can be performed with this new enzyme assay for PPT.
- Published
- 1999
11. First-trimester diagnosis of infantile neuronal ceroid lipofuscinosis (INCL) using PPT enzyme assay and CLN1 mutation analysis.
- Author
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de Vries BB, Kleijer WJ, Keulemans JL, Voznyi YV, Franken PF, Eurlings MC, Galjaard RJ, Losekoot M, Catsman-Berrevoets CE, Breuning MH, Taschner PE, and van Diggelen OP
- Subjects
- Adult, Cells, Cultured, Child, Preschool, Chorionic Villi Sampling, Female, Fluorometry methods, Humans, Infant, Male, Neuronal Ceroid-Lipofuscinoses enzymology, Neuronal Ceroid-Lipofuscinoses genetics, Pregnancy, Pregnancy Trimester, First, DNA Mutational Analysis, Neuronal Ceroid-Lipofuscinoses diagnosis, Palmitoyl-CoA Hydrolase analysis, Prenatal Diagnosis methods
- Abstract
Infantile neuronal ceroid lipofuscinosis (INCL) is a progressive neurodegenerative disorder in childhood which is caused by the deficiency of the lysosomal palmitoyl-protein thioesterase (PPT) encoded by the CLN1 gene. In a pregnancy at risk for INCL, chorionic villi (CV) were studied using a novel fluorometric PPT enzyme assay in combination with mutation-analysis of the CLN1 gene. The PPT activity in chorionic villi was found to be deficient and homozygosity for the C451T mutation in CLN1 was found. The pregnancy was terminated and the PPT deficiency was confirmed in cultured CV cells as well as in the cultured fetal skin fibroblasts. This report shows the first early prenatal diagnosis of INCL performed by fluorometric enzyme analysis and mutation analysis of the CLN1 gene.
- Published
- 1999
12. A rapid fluorogenic palmitoyl-protein thioesterase assay: pre- and postnatal diagnosis of INCL.
- Author
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van Diggelen OP, Keulemans JL, Winchester B, Hofman IL, Vanhanen SL, Santavuori P, and Voznyi YV
- Subjects
- Dose-Response Relationship, Drug, Fluorometry, Humans, Hydrogen-Ion Concentration, Thiolester Hydrolases blood, Thiolester Hydrolases cerebrospinal fluid, Time Factors, Clinical Enzyme Tests, Neuronal Ceroid-Lipofuscinoses diagnosis, Prenatal Diagnosis methods, Thiolester Hydrolases analysis
- Abstract
A deficiency of palmitoyl-protein thioesterase (PPT) was recently shown to be the primary defect in infantile neuronal ceroid lipofuscinosis (INCL). The available enzyme assays are complicated and impractical for diagnostic use. We have recently developed a new, fluorometric assay for PPT based on the sensitive fluorochrome 4-methylumbelliferone, requiring an overnight incubation to measure PPT. Now we have synthesized an analogue of this substrate which allows PPT determinations in 1 h. This improved PPT assay is simple, sensitive, and robust and will facilitate the definition of the full clinical spectrum associated with a deficiency of PPT. PPT activity was readily detectable in fibroblasts, leukocytes, amniotic fluid cells, chorionic villi, plasma, and cerebrospinal fluid from controls. PPT activity was profoundly deficient in these tissues and fluids from INCL patients. Similarly, a deficiency of PPT activity was demonstrated in patients with the variant juvenile NCL with GROD. These results show the feasibility of rapid pre- and postnatal diagnosis of INCL and its variants., (Copyright 1999 Academic Press.)
- Published
- 1999
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13. An Asn > Lys substitution in saposin B involving a conserved amino acidic residue and leading to the loss of the single N-glycosylation site in a patient with metachromatic leukodystrophy and normal arylsulphatase A activity.
- Author
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Regis S, Filocamo M, Corsolini F, Caroli F, Keulemans JL, van Diggelen OP, and Gatti R
- Subjects
- Amino Acid Sequence, Binding Sites, Child, Preschool, Glycosylation, Humans, Leukodystrophy, Metachromatic, Male, Molecular Sequence Data, Saposins, Sphingolipid Activator Proteins, Amino Acid Substitution, Arylsulfatases metabolism, Asparagine genetics, Conserved Sequence, Glycoproteins genetics, Lysine genetics
- Abstract
Sphingolipid activator proteins are small glycoproteins required for the degradation of sphingolipids by specific lysosomal hydrolases. Four of them, called saposins, are encoded by the prosaposin gene, the product of which is proteolytically cleaved into the four mature saposin proteins (saposins A, B, C, D). One of these, saposin B, is necessary in the hydrolysis of sulphatide by arylsulphatase A where it presents the solubilised substrate to the enzyme. As an alternative to arylsulphatase A deficiency, deficiency of saposin B causes metachromatic leukodystrophy. We identified a previously undescribed mutation (N215K) in the prosaposin gene of a patient with metachromatic leukodystrophy but with normal arylsulphatase A activity and elevated sulphatide in urine. The mutation involves a highly conserved amino acidic residue and abolishes the only N-glycosylation site of saposin B.
- Published
- 1999
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14. Prevalent mutations in the GALC gene of patients with Krabbe disease of Dutch and other European origin.
- Author
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Kleijer WJ, Keulemans JL, van der Kraan M, Geilen GG, van der Helm RM, Rafi MA, Luzi P, Wenger DA, Halley DJ, and van Diggelen OP
- Subjects
- Alleles, Cell Line, DNA analysis, DNA Mutational Analysis, Europe epidemiology, Fibroblasts metabolism, Gene Deletion, Haplotypes, Heterozygote, Humans, Netherlands epidemiology, Polymorphism, Genetic, Skin cytology, Leukodystrophy, Globoid Cell genetics, Mutation
- Abstract
Sixty-four unrelated patients with infantile Krabbe disease (globoid cell leukodystrophy, GLD) of Dutch (n = 41) or other European origin (n = 23) were screened for the presence of a large 30 kb deletion starting in intron 10 (IVS10del30 kb), a base substitution 1538T(T513M) and a polymorphism, 502T. The deletion and the T513M mutation were present in 52% and 8.5%, respectively, of the 82 GALC alleles of the Dutch patients. The 502T polymorphism, which had an allele frequency of 5.3% in a Dutch control panel, occurred in 65% of the GLD alleles. Analysis of patients and both parents in 26 of the families showed that del30 kb was invariably associated with 502T. However, 502T was also present on 40% of the GLD alleles with an as yet unidentified mutation, which is 7.5 times higher than its frequency in controls. This suggests that besides del30 kb at least one other relatively frequent mutation has arisen on the 502T GALC allele. A relatively high incidence of del30 kb was also found in 23 other European (non-Dutch) patients (allele frequency 35%), but T513M did not occur in this group. Practical examples described in this report illustrate the potential usefulness of mutation analysis in many families with Krabbe disease for heterozygote detection and prenatal diagnosis.
- Published
- 1997
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15. Asymptomatic and late-onset ornithine transcarbamylase deficiency caused by a A208T mutation: clinical, biochemical and DNA analyses in a four-generation family.
- Author
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Ausems MG, Bakker E, Berger R, Duran M, van Diggelen OP, Keulemans JL, de Valk HW, Kneppers AL, Dorland L, Eskes PF, Beemer FA, Poll-The BT, and Smeitink JA
- Subjects
- Adult, Aged, Allopurinol metabolism, Autopsy, Biopsy, Child, Child, Preschool, Female, Glutamine analysis, Glutamine blood, Heterozygote, Humans, Liver metabolism, Male, Mutation, X Chromosome, Ornithine Carbamoyltransferase genetics, Ornithine Carbamoyltransferase Deficiency Disease, Pedigree
- Abstract
We describe a 4-generation family in which a previously healthy 10-year-old boy died of late-onset ornithine transcarbamylase (OTC) deficiency. Pedigree analysis and allopurinol loading tests in female relatives were not informative. A missense mutation (A208T) in the OTC gene was detected in the deceased patient and in several clinically healthy male and female relatives, the oldest male being 97 years old. OTC deficiency was established in autopsy liver tissue of the propositus and liver biopsy samples of his sister, mother, and a maternal uncle. The males had 4% and 6% residual activity, respectively, the females 58% and 67%, respectively. The observed relation between the mutation and the decreased OTC activity in liver tissue of these subjects suggests that the mutation is a deleterious one. Late-onset, "mild" OTC deficiency can have a fatal or a favorable outcome. The disease can segregate undetected in families.
- Published
- 1997
16. Asymptomatic and late-onset ornithine transcarbamylase (OTC) deficiency in males of a five-generation family, caused by an A208T mutation.
- Author
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van Diggelen OP, Zaremba J, He W, Keulemans JL, Boer AM, Reuser AJ, Ausems MG, Smeitink JA, Kowalczyk J, Pronicka E, Rokicki D, Tarnowska-Dziduszko E, Kneppers AL, and Bakker E
- Subjects
- Age of Onset, Female, Humans, Male, Mutagenesis, Pedigree, Ornithine Carbamoyltransferase genetics, Ornithine Carbamoyltransferase Deficiency Disease
- Abstract
In a large five-generation Polish family, late-onset ornithine transcarbamylase (OTC) deficiency in males segregated with the missense mutation Ala208Thr (A208T), and all heterozygous females were asymptomatic. No other mutations were found in the coding sequences and intron-exon boundaries of the OTC gene. Surprisingly, the mutation originated from the great-grandfather of the index patient who died at age 59 of liver carcinoma. He never had dietary restrictions or hyperammonemic spells throughout life and appears to be the oldest male reported with OTC deficiency. The index patient had a severe OTC deficiency (3% of normal). Eight males died suddenly at ages 4 months to 23 years (average 14 years) after a foudroyant episode triggered by a common infection. The patients remained undiagnosed for 28 years because a metabolic defect was not considered to be the cause of the acute episodes. Recognition of the familial pattern of inheritance was initially unnoticed since the patients were admitted to eight different hospitals. DNA analysis predicted that two 'healthy' boys also had OTC deficiency, which was confirmed by abnormal results of allopurinol challenge tests. Initial suspicion of OTC deficiency in such families is complicated, since symptoms can develop at any age, or even remain absent. This obscures the typical pattern of X-linked inheritance in small families.
- Published
- 1996
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17. Prenatal diagnosis of Sanfilippo A syndrome: experience in 35 pregnancies at risk and the use of a new fluorogenic substrate for the heparin sulphamidase assay.
- Author
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Kleijer WJ, Karpova EA, Geilen GC, Keulemans JL, Huijmans JG, Tsvetkova IV, Voznyi YaV, and van Diggelen OP
- Subjects
- Chorionic Villi Sampling, Female, Humans, Hydrolases deficiency, Pregnancy, Reference Values, Substrate Specificity, Amniocentesis methods, Chorionic Villi enzymology, Fetal Diseases diagnosis, Hydrolases metabolism, Mucopolysaccharidosis III diagnosis, Pregnancy, High-Risk
- Abstract
We have investigated the use of a 4-methylumbelliferone (MU)-derived artificial substrate, MU-alpha-D-N-sulphoglucosaminide, for the sulphamidase assay in chorionic villi and amniotic fluid cells. In the new two-step enzyme assay, fluorescent MU is released by the successive action of endogenous sulphamidase and an added yeast enzyme preparation which hydrolyses the MU-alpha-glucosaminide intermediate. Optimal conditions for a sensitive, accurate, and convenient procedure for use in the prenatal diagnosis of Sanfilippo A syndrome are described. Previously, prenatal diagnosis of Sanfilippo A syndrome has been achieved by a radioactive sulphamidase assay in chorionic villi or in cultured amniocytes and by two-dimensional electrophoresis of glycosaminoglycans in amniotic fluid. Our experience using these methods in 35 pregnancies at risk is reported. The feasibility of the new fluorogenic assay was evaluated by retrospective testing of stored homogenates of chorionic villi and amniotic fluid cells from 22 pregnancies at risk. Unequivocal assignment of the fetal status in five affected pregnancies and 17 pregnancies with a normal outcome confirms the reliability of the new sulphamidase assay, which is in every respect more convenient than the conventional method using 35S-radiolabelled heparin.
- Published
- 1996
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18. Human alpha-N-acetylgalactosaminidase (alpha-NAGA) deficiency: new mutations and the paradox between genotype and phenotype.
- Author
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Keulemans JL, Reuser AJ, Kroos MA, Willemsen R, Hermans MM, van den Ouweland AM, de Jong JG, Wevers RA, Renier WO, Schindler D, Coll MJ, Chabas A, Sakuraba H, Suzuki Y, and van Diggelen OP
- Subjects
- Animals, Cells, Cultured, Child, Child, Preschool, Female, Genotype, Humans, Infant, Male, Mutation, Pedigree, Phenotype, Rabbits, Skin cytology, alpha-N-Acetylgalactosaminidase, Hexosaminidases deficiency, Hexosaminidases genetics, Skin enzymology
- Abstract
Up to now eight patients with alpha-NAGA deficiency have been described. This includes the newly identified patient reported here who died unexpectedly aged 1 1/2 years of hypoxia during convulsions; necropsy was not performed. Three patients have been genotyped previously and here we report the mutations in the other five patients, including two new mutations (S160C and E193X). The newly identified patient is consanguineous with the first patients reported with alpha-NAGA deficiency and neuroaxonal dystrophy and they all had the alpha-NAGA genotype E325K/E325K. Clinical heterogeneity among patients with alpha-NAGA deficiency is extreme. Two affected sibs, homozygotes for E325K, are severely affected and have the signs and symptoms of infantile neuroaxonal dystrophy, but prominent vacuolisation is lacking. The mildly affected patients (two families, three patients) at the opposite end of the clinical spectrum have clear vacuolisation and angiokeratoma but no overt neurological manifestations. Two of them are homozygous for the stop mutation E193X, leading to complete loss of alpha-NAGA protein. These observations are difficult to reconcile with a simple genotype-phenotype correlation and we suggest that factors or genes other than alpha-NAGA contribute to the clinical heterogeneity of the eight patients with alpha-NAGA deficiency. At the metabolic level, the patients with alpha-NAGA deficiency are similar. The major abnormal urinary oligosaccharides are sialylglycopeptides of the O linked type. Our enzymatic studies indicated that these compounds are not the primary lysosomal storage products.
- Published
- 1996
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19. A fluorimetric enzyme assay for the diagnosis of Sanfilippo disease type A (MPS IIIA).
- Author
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Karpova EA, Voznyi YaV, Keulemans JL, Hoogeveen AT, Winchester B, Tsvetkova IV, and van Diggelen OP
- Subjects
- Fluorometry, Humans, Hydrolases metabolism, Clinical Enzyme Tests, Mucopolysaccharidosis III diagnosis
- Abstract
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.
- Published
- 1996
- Full Text
- View/download PDF
20. Applications of a new fluorimetric enzyme assay for the diagnosis of aspartylglucosaminuria.
- Author
-
Voznyi YaV, Keulemans JL, Kleijer WJ, Aula P, Gray GR, and van Diggelen OP
- Subjects
- Acetylglucosamine urine, Amino Acid Metabolism, Inborn Errors diagnosis, Amino Acid Metabolism, Inborn Errors enzymology, Amniotic Fluid enzymology, Chorionic Villi enzymology, Female, Fibroblasts enzymology, Fluorometry, Genetic Carrier Screening, Humans, Leukocytes enzymology, Pregnancy, Prenatal Diagnosis methods, Substrate Specificity, Acetylglucosamine analogs & derivatives, Amino Acid Metabolism, Inborn Errors urine, Aspartylglucosylaminase analysis
- Abstract
L-Aspartic acid-beta-7-amido-4-methylcoumarin is a sensitive and specific fluorogenic substrate for lysosomal glycoasparaginase (aspartylglucosaminidase). Fibroblasts and leukocytes from 8 patients with aspartylglucosaminuria, showed 1-7% of the mean normal glycoasparaginase activity. Heterozygotes showed intermediate activities. Glycoasparaginase activity in chorionic villi, cultured trophoblasts, cultured amniotic fluid cells and amniotic fluid was readily detectable, indicating that prenatal analysis of aspartylglucosaminuria should be possible with this assay. beta-Aspartyl-4-methylumbelliferone was synthesized but this potential substrate can not be used to assay glycoasparaginase since it hydrolyses spontaneously.
- Published
- 1993
- Full Text
- View/download PDF
21. X-ray micro-analysis of cultured respiratory epithelial cells from patients with cystic fibrosis.
- Author
-
Sagström S, Roomans GM, Wroblewski R, Keulemans JL, Hoogeveen AT, and Bijman J
- Subjects
- 8-Bromo Cyclic Adenosine Monophosphate pharmacology, Amiloride pharmacology, Calcimycin pharmacology, Cells, Cultured, Chlorides metabolism, Cyclic AMP biosynthesis, Cystic Fibrosis metabolism, Electron Probe Microanalysis, Epithelium metabolism, Humans, Ouabain pharmacology, Potassium metabolism, Respiratory System metabolism, Sodium metabolism, Cystic Fibrosis pathology, Respiratory System pathology
- Abstract
X-ray micro-analysis was carried out on cultured respiratory cells from polyps removed from individuals with and without cystic fibrosis (CF). In a first set of experiments, proper experimental conditions were established. Washing the cells with 300 mmol l-1 mannitol in distilled water was found to give the best removal of the culture medium. The elemental concentrations stabilized in about 10 min after the start of the preincubation. Intracellular [Na] and [Cl] increased slightly with increasing passage number, whereas intracellular [K] decreased. Under resting conditions there were no significant differences in elemental content between CF and control cells, and there were no indications for abnormally high total [Ca] in CF cells. In normal cells, stimulation with a cAMP-analogue resulted in a decrease of cellular [Cl], whereas in CF cells an increase was measured. Exposure of both normal and CF cells to ouabain resulted in decreased [K] and increased [Na] and [Cl] level. The calcium ionophore A23187 had a similar effect on normal cells but did not affect CF cells markedly. Application of amiloride to the apical side of the cells resulted in a decrease of cellular [Na] in CF cells, whereas [Na] in control cells was not affected. The results correspond with what is known about the defective cAMP-regulated transepithelial Cl-transport in CF cells. The effect of the calcium ionophore on cellular electrolyte content is more complicated and may be the result of two separate effects: efflux of Cl- via a Ca(2+)-dependent mechanism and inhibition of the Na(+)-K(+)-ATPase by intracellular Ca2+ ions causing an influx of Na+ and Cl- ions.
- Published
- 1992
- Full Text
- View/download PDF
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