PubMed Health⌕ Search

Biomedical subjects

Y Ohnuki

Publications and source records attributed to Y Ohnuki.

At least 19 recordsLinked to original sources
PubMed1996

Ganglioside alterations in the central and peripheral nervous systems of patients with Creutzfeldt-Jakob disease.

We have examined the distribution and composition of gangliosides in central and peripheral nervous tissues from two patients with Creutzfeldt-Jakob disease (CJD). There were marked decreases in total ganglioside levels in CJD, with reductions in the cerebral cortex and cerebellum in the order of 20-30% and 50% of control values, respectively, though in spinal cord and sciatic nerve total gangliosides were not significantly altered. The percentage distribution of individual gangliosides was characterized by marked increases in GD3 and GD2, contrasting with severe decreases in GD1a, GD1b, GT1b and GQ1b; such changes were found throughout the patients' nervous tissues. An abnormal long-chain base composition was detected with the d20:1 component being decreased to less than 50% of control values, in the cerebrum, cerebellum and spinal cord. Changes in gangliosides occurred even in those tissues not severely affected neuropathologically. These ganglioside abnormalities are discussed in relationship to the pathogenesis of CJD.

Aged↗
PubMed1996

Chromosomal changes and progressive tumorigenesis of human bronchial epithelial cell lines.

A simian virus 40 (SV40)-transformed human bronchial epithelial cell line, BEAS-2B, underwent progressive changes, including the development of tumorigenicity, during extended in vitro passaging. Karyotypic changes occurred in parallel with the phenotypic changes. For the first 12 passages following viral transformation, there were random karyotypic changes. Immortalization occurred between passages 12 and 21, corresponding with the accumulation of four characteristic abnormal chromosomes-m-1: add(15)(p11.1); m-2: der(8;9)(q10;q10); m-3: add(16)(p13); and m-4: mar4- and the loss of one homolog of chromosomes 8, 15, 16, 21, and 22. With further passaging (from 21 to 63), the acquisition of weak tumorigenicity was observed, accompanied by an increased frequency of cells containing all four common abnormal chromosomes, m-1 through m-4, and missing one normal homolog of chromosomes 8, 15, 16, and 22. Four tumor cell lines (B39-TL, B39-TR, B61-T4 and B61-T7) were established from tumors induced by the injection of these weakly tumorigenic BEAS-2B 39th- and 61st- passage cells into athymic nude mice. One of the cell lines, B39-TL, is significantly more tumorigenic than the others. It is notable that B39-TL showed two specific abnormal chromosomes, del(3p);der(3;15) (q10;q10) and m-6; der(21)t(3;21)(p14.2;p12) inducing deletion of a short arm of chromosome 3. Fluorescence in situ hybridization analysis with a probe for protein tyrosine phosphatase-gamma demonstrated loss of heterozygosity in the 3p14 region. The development of step-wise karyotypic changes in this in vitro carcinogenesis model parallels changes documented in several common human cancers.

Animals↗
PubMed1996

[Nucleotide sequences at intron 6 and exon 7 junction of fibroblast growth factor receptor 2 and rapid mutational analysis in Apert syndrome].

Apert syndrome, acrocephalosyndactyly Type I, is an autosomal dominant craniosynostosis comprising acrocephaly, facial dysmorphism and severe syndactyly of the hands and feet. Missense mutations at codons 252 and 253 at 5'-end on exon 7 of fibroblast growth factor receptor (FGFR) 2 have been identified in a large number of patients with Apert syndrome. In this study, nucleotide sequences on the intron 6 were determined by vector ligation-PCR and direct sequencing. Five DNA samples from sporadic Apert syndrome were examined by non-RI SSCP and direct sequencing using a primer pair of intron 6 and exon 7. All cases of the syndrome showed abnormal banding pattern in the SSCP and missense mutations from Ser to Trp at codon 252 of the FGFR2 gene. The non-RI SSCP and direct sequencing of the FGFR2 exon 7 from genomic DNAs may be a useful and rapid molecular means for clinical diagnosis of Apert syndrome.

Acrocephalosyndactylia↗
PubMed1996

[Frequent missense mutations of fibroblast growth factor receptor (FGFR) gene families in craniofacial syndromes in Japanese patients].

Craniofacial syndromes, including Crouzon syndrome, Pfeiffer syndrome, Jackson-Weiss syndrome, Apert syndrome and achondroplasia, have been indicated that syndromes were associated with mutations of fibroblast growth factor receptor (FGFR) gene families. In this report, seven Japanese patients with craniofacial syndromes, three Crouzon syndromes and four achondroplasias, were analyzed on FGFR2 and FGFR3 genes by non RI-SSCP (single strand conformation polymorphisms) and direct sequencing. Missense mutations of the FGFR3 exon 10, at codon 380 in two sporadic cases and codon 375 in two familial cases, were detected in all cases of achondroplasia. Mutations of the FGFR2 were noted in Crouzon and Apert syndromes. One of three Crouzon syndromes has a missense mutation at codon 342 on exon 9. Highly frequent mutations were clustered within some localized regions of the FGFR genes in craniofacial syndromes. Alterations in these receptors due to missense mutations would thus appear closely involved in pathogenesis of craniofacial syndrome. The non RI-SSCP and direct sequencing of the FGFR genes, shown in this report, may be an appropriate approach for diagnosis of these syndromes with extensive clinical application.

Base Sequence↗
PubMed1996

[Analysis of mutations and heteroplasmy at mitochondrial DNA 11778 using non-RI single strand conformation polymorphisms in Leber's hereditary optic neuropathy].

Leber's hereditary optic neuropathy(LHON) is a maternally inherited mitochondrial disease of an acute or subacute bilateral loss of central vision. G to A substitutions at nucleotide position 11778 in mitochondrial DNA(mt DNA) have been identified in approximately 40% to 90% of patients. In this study, regions containing mt DNA 11778 mutations were analyzed by polymerase chain reaction(PCR), non-RI single strand conformation polymorphisms(SSCP) and direct sequencing. In 26 visually affected patients, mt DNA 11778 mutations were detected in 9 patients (36.4%). In one pedigree of a LHON patient(L-6), four unaffected family members had heteroplasmy of the 11778 mutation using non-RI SSCP. Ratios of the heteroplasmy between wild type and mutant mt DNAs can be detected in non-RI SSCP and accurately quantified by video densitometric analyzer. Two types of novel polymorphisms, 11696 G to A and 11719 A to G, in the mt DNA region were also found in this non-RI SSCP analysis. Non-RI SSCP is an efficient and accurate method for diagnosis of mt DNA 11778 mutations and quantifying heteroplasmy in patients with LHON and pedigrees.

Base Sequence↗
PubMed1995

Evidence for the covalent binding of SHAP, heavy chains of inter-alpha-trypsin inhibitor, to hyaluronan.

We previously showed that serum-derived 85-kDa proteins (SHAPs, serum-derived hyaluronan associated proteins) are firmly bound to hyaluronan (HA) synthesized by cultured fibroblasts. SHAPs were then identified to be the heavy chains of inter-alpha-trypsin inhibitor (ITI) (Huang, L., Yoneda, M., and Kimata, K. (1993) J. Biol. Chem. 268, 26725-26730). In this study, the SHAP.HA complex was isolated from pathological synovial fluid from human arthritis patients. The SHAP.HA complex was digested with thermolysin, followed by CsCl gradient centrifugation. The HA-containing fragments thus obtained were further digested with chondroitinase AC II and subjected to TSK gel high performance liquid chromatography (HPLC). Peptide-HA disaccharide-containing fractions (the SHAP.HA binding regions) were further purified by reverse phase HPLC. Major peaks were analyzed by protein sequencing and mass spectrometry (electrospray ionization mass spectrometry and collision induced dissociation-MS/MS). By comparison with the reported C-terminal sequences of the human ITI family, the peptides were found to correspond to tetrapeptides derived from the C termini of heavy chains 1 of and 2 of inter-alpha-trypsin inhibitor (HC1 and HC2), and heavy chain 3 of pre-alpha-trypsin inhibitor (HC3), respectively, and a heptapeptide from HC1. Mass spectrometric analyses suggested that the C-terminal Asp of each heavy chain was esterified to the C6-hydroxyl group of an internal N-acetylglucosamine of HA chain. This report is the first demonstration to give evidence for the covalent binding of proteins to HA.

Alpha-Globulins↗
PubMed1995

[Molecular diagnosis of a kindred with novel mutation of methylmalonyl-CoA mutase gene using non-RI SSCP].

A deficiency of methylmalonyl-CoA mutase (MCM) results in methylmalonic acidemia, which is inherited as an autosomal recessive disease and is characterized by accumulation of precursors and abnormal derivatives of methylmalonyl-CoA in body fluids. Abnormal splicing with 13 base pairs (bp) insertion at MCM exons 2 and 3 junction in MCM transcripts and a homozygous point mutation, g to a transition, on 5 bp downstream exon 2 were detected in a proband with methylmalonic acidemia. The parents in the kindred were heterozygous carriers of the g to a transition in MCM intron 2. Non-RI single strand conformation polymorphisms (SSCP) was conducted to devise for analysis of this MCM mutation. This non-RI SSCP is considered to be useful diagnostic means with high potential for extended clinical application.

Adult↗
PubMed1993

Development of tumorigenicity in simian virus 40-immortalized human bronchial epithelial cell lines.

Of five SV40-transformed clonal human bronchial epithelial cell lines previously shown to be nontumorigenic at early passages (R. R. Reddel et al., Cancer Res., 48: 1904-1909, 1988), two lines (BES-1A1 and BEAS-2B) from different donors have become weakly tumorigenic with further passaging. BES-1A1 passage 26 cells formed tumors in 3 of 9 athymic nude mice given s.c. injections, whereas BEAS-2B cells of > or = 32 passages formed highly cystic tumors at 8 of 58 injection sites after long latency periods [17 +/- 7 (SD) weeks]. These tumors took a total of 36 +/- 8 weeks to reach a diameter of 1.0 cm. Tumor cell lines were established from four BEAS-2B tumors, and these are resistant to the growth-inhibitory effects of serum, an inducer of squamous differentiation in BEAS-2B and normal bronchial epithelial cells. This finding supports the hypothesis that development of resistance to inducers of terminal squamous differentiation may be a step in the process of bronchial carcinogenesis. One of these tumor cell lines, B39-TL, is significantly more tumorigenic than the others and has a deletion from the short arm of chromosome 3 as has been described previously for some naturally occurring human bronchial carcinomas. Thus, from the clonally derived BEAS-2B cell line, cell populations with various degrees of tumorigenicity have developed. Analysis of the changes in these cells may yield insights into the multiple events involved in acquisition of the tumorigenic phenotype.

Animals↗
PubMed1992

Portal vein thrombosis after splenectomy successfully treated by an enormous dosage of fibrinolytic agent in a short period: report of two cases.

Portal vein thrombosis (PVT) after splenectomy in a patient with portal hypertension occurs with unusually high frequency. Recently, two patients with PVT following splenectomy were treated by fibrinolytic therapy with an enormous dosage of urokinase (UK) in a short period. PVT was quickly dissolved without side effects and the patients are now doing well without any recurrence of PVT. Therefore, when there is no evidence of bowel infarction, fibrinolytic therapy with an enormous dosage of UK over a short period is deemed to be both effective and essential as a conservative therapy for PVT.

Adult↗
PubMed1988

Biochemical and cytogenetic studies of human lung cancers.

In ongoing studies, we have tested resected lung cancers from 41 men and 49 women; of those with primary lung cancer, 46 patients are free of disease and 35 have died of cancer or have persistent disease. Measurements and studies were as follows: total cellular deoxyribonucleic acid content by image analysis (n = 77); total genomic deoxyribonucleic acid methylation state and banding patterns from probed Southern blots (n = 36); radioimmunoassay for motilin, bombesin, gastrin, vasoactive intestinal peptide, and cholecystokinin (n = 18); and cytogenetic analysis (n = 39). All lung cancers were hyperploid. Adenocarcinomas and epidermoid carcinomas were generally hexaploid to nearly septaploid; comparisons by stage and histologic features suggested potential prognostic correlations. There was general hypomethylation of deoxyribonucleic acid (p less than 0.001). Deoxyribonucleic acid digests from restriction endonuclease Hpa II, when probed with deoxyribonucleic acid homologous to KPN, showed banding patterns that separated histologically indistinguishable primary adenocarcinomas and metastatic adenocarcinomas from one another. Cancers studied with radioimmunoassay were all negative for polypeptide hormones. Five cancers grew adequately in vitro to permit study of 190 detailed karyotypes (20 to 50 per tumor). Chromosome modal numbers ranged from 49 to 109. There were from 4 to 20 clearly abnormal marker chromosomes per tumor; abnormality derived from chromosome 1 was prevalent. Ten of 19 tumors xenotransplanted to nude mice were carried through two to five transplant generations without a change in histologic patterns.

Animals↗
PubMed1988

[Clinicopathological study of early multicentric gastric carcinomas].

During a two-year period, from August, 1983 to July, 1985, 144 gastric cancer patients have been treated at the National Tohsei Hospital. Among them, 57 patients were determined as having an early gastric carcinoma. Four patients (7.0%) of this number were found to have a multicentric type, 3 manifesting a double II c type, and 1 patient having both a I type and II a type. Pathologically these two types carcinomas were found in only one patient, in a so-called kissing position.

Adenocarcinoma↗
PubMed1985

High performance liquid chromatographic separation of stereoisomeric bile acids as their UV-sensitive esters.

High performance liquid chromatographic separation of a series of mono-, di- and trihydroxylated 5 beta-cholanic acids, which differ only in position and configuration of hydroxyl groups at positions C-3, C-7 and/or C-12, is reported. The C-24 free acids were derivatized to four different classes of UV-sensitive esters, i.e., p-bromophenacyl (BP), m-methoxyphenacyl (MP), 4-nitrophthalimidemethyl (NPM) and 9-anthrylmethyl (AM) esters, and chromatographed on two variants of C18 reversed-phase columns (Nova-Pak C18 and Zorbax ODS) with methanol-water systems as mobile phase. Separation efficiency and elution order of some isomeric pairs were influenced by both the structure of the C-24 ester groups and the nature of the columns used. Excellent chromatographic properties were found for those derivatives, particularly for the NPM esters.

Bile Acids and Salts↗
PubMed1985

Selective growth of normal adult human urothelial cells in serum-free medium.

A serum-free medium (HMRI-2) has been developed for the outgrowth and subculture of epithelial cells from normal adult human ureter and bladder. Medium HMRI-2 consists of Ham's MCDB 152 with double the amounts of the essential amino acids in Stock 1, low Ca2+ (0.06 mM) and is supplemented with epithelial growth factor, 5 ng/ml; transferrin, 5 micrograms/ml; insulin, 5 micrograms/ml; ethanolamine and phosphoethanolamine, 0.1 mM each; hydrocortisone, 2.8 X 10(-6) M; and bovine pituitary extract, 126 micrograms protein/ml. The cultured cells showed ultrastructural markers of epithelial cells (prekeratin fibers, tonofilaments, surface microvilli with glycocalyx), exhibited ABO antigens, and had a normal human diploid karyotype. Primary cultures could be subcultured and also cryopreserved in HMRI-2 in liquid nitrogen. Cells in mass cultures showed a population doubling time of 40.5 +/- 4.5 h and had a maximum in vitro life span of 20 to 25 population doublings. It was observed that primary outgrowths, secondary cultures, and even cryopreserved cells all retained the capacity to respond to high Ca2+ and serum by differentiation and desquamation. This study has resulted in the availability of easily obtainable serum-free epithelial cultures from normal adult human ureter and bladder. The useful in vitro life span of these cultures may be important in future studies of carcinogenesis.

Adolescent↗
PubMed1985

Novel antigen system for tracking epithelial cell migration in serum-free culture.

We report here a unique system for tracking normal human urothelial cell migration in serum-free culture medium (HMRI-1). The key observation was that urothelial cells deposited red blood cell surface antigen on the culture dish in a remarkable pattern. Scrutiny of this pattern showed that each migrating cell left behind antigen imprints which formed parallel tracks the width of the cell. Hence the previous migratory history of the cells was instantly mapped by simply visualizing the antigen tracks deposited by the cells on the dish. Apart from providing a simple method for tracking urothelial cells, this observation has wider implications for mechanistic studies of epithelial cell movement in general. It also highlights the complicating effects associated with the addition of serum as a traditional culture supplement, since the inclusion of serum in the HMRI-1 medium abolished the above effect by inhibiting cell migration.

ABO Blood-Group System↗