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Biomedical subjects

S Tomatsu

Publications and source records attributed to S Tomatsu.

At least 73 records · Page 4Linked to original sources
PubMed1995

Four novel mutations in mucopolysaccharidosis type VII including a unique base substitution in exon 10 of the beta-glucuronidase gene that creates a novel 5'-splice site.

Mucopolysaccharidosis type VII (MPS VII), or Sly syndrome, is a lysosomal storage disorder caused by a deficiency in the enzyme beta-glucuronidase. Various clinical phenotypes of this autosomal recessively inherited disease have been described. Recent isolation and characterization of human beta-glucuronidase cDNA and the genomic sequences facilitate analysis of molecular defects underlying the different phenotypes, and eight mutations in the beta-glucuronidase gene have been described. This report summarizes studies characterizing four new mutations in two Caucasian patients with a severe form of MPS VII. Three are point mutations, resulting in two missense and one nonsense change, and one is a 38 bp deletion. The first patient was a compound heterozygote having P148S and Y495C alleles. The second patient was a compound heterozygote of W507X and a 38 bp deletion at position 1642-1679 in exon 10(1642 delta 38nt). The 38 bp deletion was caused by a single base change mutation in exon 10 that generates a new, premature 5' splice site. Expression of mutant cDNAs encoding each of the four mutations showed that all four resulted in a severe reduction of beta-glucuronidase activity, indicating that these mutations are responsible for the reduced enzyme activity in patient cells. These four previously undescribed mutations provide further evidence for the broad molecular heterogeneity in Sly syndrome.

Base Sequence↗
PubMed1995

Genetic analysis of IgE and the IGHE, IGHEP1 and IGHEP2 genes in atopic families.

The familial occurrence of allergic diseases was studied in 478 individuals and their family members. The results showed that there was pronounced familial clustering. Total serum IgE concentrations of atopic patients and their parents were well correlated. Moreover, the concentration of specific IgE of the patients and their parents was also well correlated, suggesting that IgE production was genetically determined. To determine if major structural abnormalities of IGHE, IGHEP1 and IGHEP2 genes might lead to aberrant control and subsequent increase in IgE concentration, genomic DNAs from 55 individuals, i.e., 31 atopic patients and their family members, were examined. We detected the IGHE, IGHEP1 and IGHEP2 genes in all 55 leukocyte DNAs. We could not find any large deletions or duplications in the IGHE, IGHEP1 and IGHEP2 genes of atopic patients with high serum IgE concentrations.

Adolescent↗
PubMed1995

Mucopolysaccharidosis IVA: identification of a common missense mutation I113F in the N-Acetylgalactosamine-6-sulfate sulfatase gene.

Mucopolysaccharidosis IVA is an autosomal recessive lysosomal storage disorder caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase. The recent isolation and characterization of cDNA and genomic sequences encoding GALNS has facilitated identification of the molecular lesions that cause MPS IVA. We identified a common missense mutation among Caucasian MPS IVA patients. The mutation was originally detected by SSCP, and successive sequencing revealed an A-->T transversion at nt 393. This substitution altered the isoleucine at position 113 to phenylalanine (I113F) in the 622 amino acid GALNS protein and was associated with a severe phenotype in a homozygote. Compound heterozygotes with one I113F-allele mutation have a wide range of clinical phenotypes. Transfection experiments in GALNS-deficient fibroblasts revealed that the mutation drastically reduces the enzyme activity of GALNS. Allele-specific oligonucleotide or SSCP analysis indicated that this mutation accounted for 22.5% (9/40) of unrelated MPS IVA chromosomes from 23 Caucasian patients, including 6 consanguineous cases. Of interest, the I1e 113-->Phe substitution occurred in only Caucasian MPS IVA patients and in none of the GALNS alleles of 20 Japanese patients. These findings identify a frequent missense mutation among MPS IVA patients of Caucasian ancestry, that results in severe MPS IVA when homoallelic, and will facilitate molecular diagnosis of most such patients and identification of heterozygous carriers. In addition to this common mutation, 10 different point mutations and 2 small deletions were detected, suggesting allelic heterogeneity in GALNS gene.

Base Sequence↗
PubMed1995

[Mucopolysaccharidoses].

Mucopolysaccharidoses (MPS) are a lysosomal storage disorders caused by deficiency of several enzymes needed for degradation of mucopolysaccharides (glycosaminoglycans). Undegraded glycosaminoglycans accumulate in the cell, part of which are excreted into the urine. There are 10 known enzyme deficiencies that give rise to six distinct MPS. Despite the different enzyme defect, there is clinical similarity between different deficiencies and conversely, wide phenotypic variety among even one enzyme deficiency. Most of the cDNAs corresponding the defect enzyme as well as genomic DNA have been cloned. Mutational analyses of the patient gene have revealed the molecular basis of the disease, correlation of the genotype and phenotype and made the accurate heterozygote diagnosis feasible. Supportive management can greatly improve the quality of life and moreover, development of therapies, such as BMT, administration of recombinant enzyme and gene transfer are in progress for the patients suffering from MPS.

Animals↗
PubMed1994

Overexpression rescues the mutant phenotype of L176F mutation causing beta-glucuronidase deficiency mucopolysaccharidosis in two Mennonite siblings.

Two beta-glucuronidase-deficient Mennonite siblings were found to be homozygous for a mutation in exon 3 of the beta-glucuronidase gene that produces a Leu-->Phe substitution (L176F). The siblings also have the previously described benign polymorphism, P649L. Although their cultured fibroblasts contained 1.5-2.2% of normal beta-glucuronidase activity, transient expression of the L176F/P649L cDNA in COS cells produced nearly as much enzyme activity as the wild-type control cDNA. The L176F/P649L enzyme was as stable as wild-type enzyme following endocytosis by fibroblasts and delivery to lysosomes, but was more labile to heat inactivation at 65 degrees C. To study the mutant enzyme at lower levels of expression, we stably transfected mouse mucopolysaccharidosis type VII cells with the L176F/P649L cDNA and selected single-copy cell lines. Metabolic labeling with [35S]methionine revealed that cell lines expressing the mutant enzyme activity at low levels (7-10% of the wild type) actually produced the same amount of enzyme protein as the cell lines expressing the more active wild-type enzyme. However, the cell lines expressing four times this much mutant enzyme protein produced 150-200% as much enzyme activity as the cell line expressing the single-copy wild-type cDNA. These data suggest that overexpression can drive the folding reaction or the self-association of mutant monomers to form active tetramers and, at least partially, correct the beta-glucuronidase deficiency seen at low levels of expression with certain missense mutations.

Alleles↗
PubMed1994

Mucopolysaccharidosis IV A: molecular cloning of the human N-acetylgalactosamine-6-sulfatase gene (GALNS) and analysis of the 5'-flanking region.

Mucopolysaccharidosis IV A (MPS IV A) is the result of a genetic deficiency in a lysosomal hydrolase, N-acetylgalactosamine-6-sulfatase (GALNS). To investigate MPS IV A patients at the level of the genome, we analyzed the structure of the human GALNS-encoding gene. From the genomic library of a normal subject in lambda EMBL3, we isolated five overlapping clones covering the coding region of the GALNS cDNA and determined the structural organization. The gene is about 50 kb long and contains 14 exons. The 5'-flanking region lacks a canonical TATA box and CCAAT sequences, but is G+C-rich (70.5%), with four GC boxes, characteristic of a housekeeping gene promoter. The transcription initiation site was determined by primer extension analysis, using RNA from human liver and HeLa cells. Transcription was found to initiate at a few sites, the major ones being 58 and 22 bp upstream of the translation initiation codon. The 5'-flanking region had promoter activity by transient expression, determined using a CAT assay. In addition, this region retained promoter activity, even in reverse orientation. The region -98 to -1 upstream of the ATG codon was defined by deletion analysis to be a minimal promoter. One GC box in this region is likely to be a binding site of a regulatory element.

Base Sequence↗
PubMed1994

Identification of a nonsense mutation in ALD protein cDNA from a patient with adrenoleukodystrophy.

The molecular basis of X-linked adrenoleukodystrophy (ALD) was investigated. Six (A to 50) fragments of cDNA for ALD protein (Mosser et al. Nature 361: 726-730, 1993) from an adult patient with adrenomyeloneuropathy were amplified by PCR and mutations were screened by Mutation Detection Enhancement gel electrophoresis. A single base substitution (2154 C-->T, which resulted in the formation of a termination codon for glutamine (Q590STOP) and deletes Pst I site (CTGCAG-->CTGTAG), was detected. Eight other ALD patients did not have this mutation. A family study revealed the presence of both the mutant and normal alleles in the mother, a sister and a niece, indicating that these individuals were carriers. A nephew with childhood ALD who died 10 years earlier had the same mutant allele as detected by Pst I restriction assay. This report is the first description of a mutant allele for ALD, at the cDNA level, and presents confirmatory evidence of ALD protein as the primary etiology of ALD.

ATP Binding Cassette Transporter, Subfamily D, Mem↗
PubMed1994

Proliferative responses towards native, heat-denatured and pepsin-treated ovalbumin by peripheral blood mononuclear cells from patients with hen's egg-sensitive atopic dermatitis.

In order to clarify the mechanism of food-antigen recognition, the proliferative responses of peripheral blood mononuclear cells (PBMCs) to native, heat-denatured or pepsin-treated ovalbumin (OA) were investigated in 16 hen's egg-sensitive patients with atopic dermatitis (AD). Seven of them had hypersensitivity to boiled hen's egg and others had not. The responses of PBMCs to heat-denatured OA were lower than those to native OA in the patients without hypersensitivity to boiled hen's egg. However, there were no differences of the responses of PBMCs between heat-denatured OA and native OA in the patients with hypersensitivity to boiled hen's egg. Moreover, the reduction of the responses of PBMCs to pepsin-treated OA was recognized in six out of seven patients. The primary structure of OA did not change by heating or pepsin treatment according to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). These results suggested that the secondary structure of OA changed in connection with the reduction of the responses of PBMCs to denatured OA. In addition, we demonstrated the suppressive effect of anti-HLA-DR and anti-HLA-DQ monoclonal antibodies on the proliferative response of PBMCs to OA. The results suggested that the proliferative responses of PBMCs to OA were restricted by HLA-DR or HLA-DQ in hen's egg-sensitive patients with AD.

Antibodies, Monoclonal↗
PubMed1994

Intravenous immunoglobulins suppress immunoglobulin productions by suppressing Ca(2+)-dependent signal transduction through Fc gamma receptors in B lymphocytes.

A high dose intravenous immunoglobulin (IVIG) therapy is used in the treatment of a wide range of autoimmune disorders. However, the mechanisms of the action of IVIGs remain poorly understood. To analyse the mechanisms of effects of IVIGs on immunoglobulin (Ig) production of B cells, the effects of IVIGs on B lymphoblastoid cell lines transformed by Epstein-Barr virus (LCLs) were investigated. The productions of IgG or IgM of LCLs were dose-dependently suppressed by polyethylene glycol (PEG)-treated IVIG or pH 4-treated IVIG though the productions were not or only slightly suppressed by pepsin-treated IVIG. The suppression by IVIGs was blocked by anti-human IgG Fc or anti-Fc gamma RII. C mu gene expression and mu s C terminal gene expression of LCLs were suppressed by PEG-treated IVIG, whereas neither C mu gene expression nor mu s C terminal gene expression of LCLs were suppressed by pepsin-treated IVIG. Although the increase in intracellular calcium concentration in LCLs was not suppressed by pepsin-treated IVIG, the increase was suppressed by PEG-treated IVIG. This suppressing effect of PEG-treated IVIG on intracellular calcium concentration of LCLs was blocked by anti-human IgG Fc or anti- Fc gamma RII. Our results suggest that IVIGs suppressed the Ca(2+)-dependent signal transduction through Fc gamma R on B-cell membrane, consequently, the transcription of C mu mRNA, especially secreted mu mRNA was suppressed in the B cells.

B-Lymphocytes↗
PubMed1994

Comparison of allergic diseases and specific IgE antibodies in different parts of Japan.

We investigated the differences of allergic diseases and specific IgE antibodies to inhaled or ingested allergens between two cities in a Japanese pediatric population. One of the two cities is Itoman, which is a subtropical city in Okinawa prefecture; the other is Gifu, which is a Temperate Zone city in Gifu prefecture. The prevalence of asthma or wheezy bronchitis was significantly higher in Itoman; however, the prevalence of atopic dermatitis was significantly higher in Gifu. Furthermore, the positive percentages of specific IgE antibodies to inhaled allergens such as house dust, and Dermatophagoides farinae were higher in Itoman. The positive percentages of specific IgE antibodies to ingested allergens such as egg white, cow milk, and soybean were higher in Gifu. These results suggest that the differences in prevalence of allergic diseases and allergens may depend on differences in the triggering environmental factors between different parts of Japan.

Administration, Inhalation↗
PubMed1994

Novel subtype of peroxisomal acyl-CoA oxidase deficiency and bifunctional enzyme deficiency with detectable enzyme protein: identification by means of complementation analysis.

We describe four infants with a novel subtype of an isolated deficiency of one of the peroxisomal beta-oxidation enzymes with detectable enzyme protein. The patients showed characteristic clinical and biochemical abnormalities, including hypotonia, psychomotor retardation, hepatomegaly, typical facial appearance, accumulation of very-long-chain fatty acids, and decreased lignoceric acid oxidation. However, beta-oxidation enzyme proteins were detected by immunoblot analyses, and large peroxisomes were identified by immunofluorescence staining. In order to identify the underlying defect in these patients, complementation analysis was introduced using fibroblasts from these patients and patients with an established deficiency of either acyl-CoA oxidase or bifunctional enzyme, as identified by immunoblotting. In the complementing combinations, fused cells showed increased lignoceric acid oxidation, resistance against 1-pyrene dodecanoic acid/UV selection, and normalization of the size and the distribution of peroxisomes. The results indicate that two patients with a more severe clinical course were suffering from bifunctional enzyme deficiency and that the other two infants, who were siblings and had a less severe clinical presentation, were the first patients with acyl-CoA oxidase deficiency with detectable enzyme protein.

3-Hydroxyacyl CoA Dehydrogenases↗