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Biomedical subjects

A Ono

Publications and source records attributed to A Ono.

At least 91 records · Page 5Linked to original sources
PubMed1997

Immunologic and virologic characterization of the primary infiltrating cells in the aqueous humor of human T-cell leukemia virus type-1 uveitis. Accumulation of the human T-cell leukemia virus type-1-infected cells and constitutive expression of viral and interleukin-6 messenger ribonucleic acids.

PURPOSE: To characterize immunologically and virologically the infiltrating cells in the aqueous humor of patients with human T-cell leukemia virus type-1 (HTLV-1) uveitis (HU). METHODS: With their informed consent, patients had 0.1 ml of the aqueous humor in the anterior chamber collected with a needle under an operating microscope. An aliquot of the collected sample from patients without steroid therapy was examined by May-Giemsa staining and immunocytochemically. The presence of the HTLV-1-infected cells was investigated by polymerase chain reaction (PCR) using the gag and pol regions of the provirus genome. The population of the infected cells was compared by PCR testing the amplification of the virus genome from 60 cells, or determining the endpoint of successful amplification of the twofold dilution series of the samples, collected from the aqueous humor and peripheral blood mononuclear cells (PBMCs), which were obtained at the same time. Expression of viral and cytokine genes was studied by reverse transcriptase-PCR (RT-PCR). The interleukin-6 (IL-6) level in the aqueous humor of patients with HU and control subjects was measured by a high-sensitivity enzyme-linked immunosorbent assay kit. RESULTS: The number of the infiltrating cells ranged from 475 to 3563 (mean = 2111) per 0.1 ml of aqueous humor, and all the identifiable cells were lymphocytes. Most of them were CD3-positive T cells (mean = 78%), whereas CD4-positive cells constituted less than half (mean = 35.3%). HTLV-1 provirus was detected by PCR in the infiltrating cells of 36 of 38 patients with HU tested, whereas it was detected in 1 of 4 seropositive patients with other entities of uveitis. A higher population of the infected cells in the aqueous humor than in the PBMC was found in seven of nine patients with HU by two independent approaches. Expression of HTLV-1 env or pX genes or both was shown in all 12 patients with HU tested by RT-PCR. IL-6 messenger ribonucleic acid (mRNA) was detected by RT-PCR in 10 of these 12 patients, whereas those of interleukin-1 alpha, interleukin-2, interleukin-4, and tumor-necrosis factor-alpha were not, and that of interferon-gamma was detected in only 1 patient. The IL-6 level was elevated significantly in the aqueous humor of nine patients with HU compared with that of five control subjects (520.2 +/- 841 pg/ml versus 2.77 +/- 1.59 pg/ml, P < 0.01 by Mann-Whitney test). CONCLUSIONS: HU is characterized by lymphocytic infiltration with a predominance of T cells and by the presence and probable accumulation of HTLV-1-infected lymphocytes in the affected eye. Production of viral antigens and IL-6 by the infiltrating cells could be responsible for the development of HU.

Adult↗
PubMed1997

[Human T-lymphotropic virus type 1 uveitis in children].

We report here five pediatric patients with human T-lymphotropic virus type 1 (HTLV-I) uveitis. The patients were one boy and four girls aged between 3 and 14 years. The transmission route was considered to be breast feeding from their mothers. All patients had unilateral uveitis and the ocular symptoms were similar to those in HTLV-I uveitis in adults. The ocular inflammation responded to therapy with topical or systemic corticosteroids, but recurred in three patients. HTLV-I provirus DNA was detected by polymerase chain reaction (PCR) from infiltrating cells in the anterior chamber in one patient. The percentage of HTLV-I-infected cells in the peripheral blood mononuclear cells was measured by quantitative PCR, and the values were high (2.9 approximately 7.3%) in three cases tested as compared with an asymptomatic carrier. These five cases show that HTLV-I uveitis can be induced in a relatively short period (3 approximately 10 years) after the viral infection, and that HTLV-I uveitis should be considered as one possible etiology of uveitis in children, particularly in a viral endemic area.

Adolescent↗
PubMed1997

Synthesis of stereoselectively 5'-monodeuterated nucleoside with defined S/R-ratios. An application to the assignment of 5'-methylene signals of DNA oligomers.

A method to prepare 5'-monodeuterated nucleosides with various S/R-ratios is described. 5-Oxopentose derivatives synthesized from glucose were converted into 5-monodeuterated pentose derivatives by LiAID4 in the presence of various ligands. The stereoselectivities of the deuteration reactions were investigated under a variety of conditions, and the S/R-ratios of the 5-monodeuterated pentoses varied from 4 : 1 to 1 : 7.4. By mixing these 5-monodeuterated pentose derivatives, we have successfully synthesized thymidine with a defined S/R-ratio at C5'.

Carbon Isotopes↗
PubMed1997

Selective multiple labeling strategy to obtain accurate NMR parameters for nucleic acids. Conformational analysis around the glycosidic bond.

A systematic synthesis of pyrimidine nucleosides with 13C-labels only at specific atoms in both the base and sugar moieties has been developed in order to obtain conformational information about the glycosidic bond through measurement of the accurate vicinal coupling constants between H1' and C2/C6. For this purpose, 13C labels at three positions, namely C1', C2, and C6, are essential. We have synthesized selectively multiply labeled nucleosides, such as [2, 1'-13C2]-thymidine, [2, 1'-13C2]-2'-deoxycytidine, [6, 1'-13C2]-thymidine, and [6, 1'-13C2]-2'-deoxycytidine. These nucleosides will be useful to determine the relative orientation of the base and sugar moieties.

Carbon Isotopes↗
PubMed1997

X-ray analysis of DNA dodecamer containing 2'-deoxy-N6-methoxyadenosine.

Oxyamines have been known as a mutagen which attacks amino groups of DNA bases. It is expected that the modified adenine derivative has a tautomer which can form a stable base pair not only with thymine but also with cytosine. For establishing such tautomerization mutagenesis, we have solved a crystal structure of DNA dodecamer containing 2'-deoxy-N6-methoxyadenosine. It is shown that the N6-methoxyadenine takes a imino form to form a Watson-Crick type pairing with cytosine in the DNA duplex.

Base Composition↗
PubMed1997

Structure model and physicochemical properties of the C-U mismatch pair in the double stranded RNA in solution.

Structure of the C-U mismatch pair was reported only in crystal but none in solution. Here we have studied the structure and physicochemical properties of the C-U mismatch pair in a double stranded RNA in solution. RNA oligomers r(CGACUCAGG) and r(CCUGCGUCG) form a double stranded structure with the C-U pair. The arrangement of the C-U pair derived from the model building based on nOe gives a similar structural feature to that in crystal. The modeling reveals that the amino-proton of cytidine and the keto-oxygen of uridine are located within hydrogen bonding distance, and the imino proton of uridine is exposed to bulk water. From the melting experiment which monitors chemical shifts of non-exchangeable protons, the melting of all the base-pairs including the C-U pair occurs simultaneously. This suggests the possibility where the C-U mismatch pair is stacked on the neighboring base-pairs in the double helix until the duplex is denatured to single strands.

Base Composition↗
PubMed1996

Base analog N6-hydroxylaminopurine mutagenesis in Escherichia coli: genetic control and molecular specificity.

We have studied the molecular specificity of the base analog N6-hydroxylaminopurine (HAP) in the E. coli lacI gene, as well as the effects of mutations in DNA repair and replication genes on HAP mutagenesis. HAP induced base substitutions of the two transition types (A . T-->G . C and G . C-->A . T) at equal frequency. This bi-directional transition specificity is consistent with in vitro primer extension experiments with the Klenow fragment of DNA polymerase I in which we observed that either dTTP or dCTP were incorporated opposite HAP in an oligonucleotide template. The spectrum of HAP-induced transitions was different from the spontaneous transitions in either a wild-type or a mismatch-repair-defective (mutL) strain. Mutations in genes controlling excision repair, exonucleolytic proofreading, mismatch correction, error-prone (SOS) repair and 8-oxo-guanine repair did not affect HAP-induced mutagenesis substantially. However, an extensive deletion of several genes in the uvrB-bio region conferred supersensitivity to the lethal and mutagenic effects of HAP, perhaps due to an effect on HAP metabolism. dnaE antimutator alleles reduced HAP-forward mutagenicity in allele-specific manner: dnaE911 reduced it several fold, while dnaE915 abolished it almost completely. The results obtained are consistent with the idea that HAP is mutagenic in E. coli via a pathway generating replication errors.

Adenine↗
PubMed1996

Carnitine transport defect in fibroblasts of juvenile visceral steatosis (JVS) mouse.

Juvenile visceral steatosis (JVS) mice are associated with systemic carnitine deficiency (Kuwajima, et al., 1991). In order to investigate the cause of this deficiency, we compared fibroblast carnitine transport activities in normal mice and JVS mice. The kinetic analysis showed that in formal fibroblasts, the Km and Vmax values for saturable uptake was 15.6 microM and 2.56 pmol/min/mg protein, respectively. In JVS fibroblasts, however, saturable uptake was not observed. There was no great difference in the linear component of uptake between normal and JVS fibroblasts. At the physiological concentration (50 microM) of carnitine, the fibroblast carnitine transport activity in JVS mice was decreased to 18% of that in normal mice. Thus there is hardly any carnitine transport activity in the fibroblasts of JVS mice, indicating that the JVS mouse can be regarded as an animal model of primary carnitine deficiency.

Animals↗
PubMed1996

Isolation of cDNA and genomic clones of a human Ras-related GTP-binding protein gene and its chromosomal localization to the long arm of chromosome 7, 7q36.

A Ras-related GTP-binding protein cDNA has been isolated from a human skin fibroblast cDNA library using a genomic subclone derived from a YAC clone as a probe. The polypeptide, consisting of 184 amino acids deduced from nucleotide sequences, contains five repeats of the Ras-related GTP-binding region and is highly homologous to the rat RHEB (Ras homologue enriched in brain) gene, which encodes a Ras-related growth factor- and synaptic activity-regulated protein, with 98.9% amino acid identity. Therefore, it is suggested to be a human homologue of the rat RHEB protein, and we have designated it human RHEB. Using fluorescence in situ hybridization, we concluded that this human RHEB gene was localized to band q36 on chromosome 7. Considering the chromosomal localization as well as the potential function of this protein, it will be very important to investigate whether it may play a role in the etiopathogenesis of holoprosencephaly type 3 or hereditary sacral agenesis, in which the disease susceptible locus is linked to the microsatellite marker, D7S22, in this chromosomal region, 7q36.

Amino Acid Sequence↗
PubMed1996

Definition of the locus responsible for systemic carnitine deficiency within a 1.6-cM region of mouse chromosome 11 by detailed linkage analysis.

Carnitine is an essential cofactor for oxidation of mitochondrial fatty acids. Carnitine deficiency results in failure of energy production by mitochondria and leads to metabolic encephalopathy, lipid-storage myopathy, and cardiomyopathy. The juvenile visceral steatosis (JVS) mouse, an animal model of systematic carnitine deficiency, inherits the JVS phenotype in autosomal recessive fashion, through a mutant allele mapped to mouse chromosome 11. As a step toward identifying the gene responsible for JVS by positional cloning, we attempted to refine the jvs locus in the mouse by detailed linkage analysis with 13 microsatellite markers, using 190 backcross progeny. Among the 13 loci tested, 5 (defined by markers D11Mit24, D11Mit111, D11Nds9, D11Mit86, and D11Mit23) showed no recombination, with a maximum lod score of 52.38. Our results implied that the jvs gene can be sought on mouse chromosome 11 within a genetic distance no greater than about 1.6 cM.

Animals↗
PubMed1996

Altered expression of carnitine palmitoyltransferase II in liver, muscle, and heart of mouse strain with juvenile visceral steatosis.

We conducted a quantitative study of the effect of carnitine deficiency on the mRNA level of carnitine palmitoyltransferase II in the liver, muscle and heart of mice with juvenile visceral steatosis, a strain that is systematically deficient in carnitine. The amount of carnitine palmitoyltransferase II mRNA was increased in liver and muscle of homozygotes, as compared with heterozygotes and normal controls, at 2, 4, and 8 wk of age. The mRNA levels of this enzyme were normalized after carnitine administration. The mRNA level of carnitine palmitoyltransferase II in the heart was increased only at 8 wk, and was not affected by carnitine administration. These results suggest that carnitine displays some effect on the mRNA level of the carnitine palmitoyltransferase II gene in liver and muscle, probably through fatty acid metabolic change.

Ammonia↗
PubMed1996

Disordered expression of hepatic glycolytic and gluconeogenic enzymes in Otsuka Long-Evans Tokushima fatty rats with spontanteous long-term hyperglycemia.

Expression of key regulatory enzymes involved in glucose metabolism was studied in the livers of Otsuka Long-Evans Tokushima fatty (OLETF) rats, a model of non-insulin dependent diabetes mellitus. The activity and mRNA levels of glucokinase and L-type pyruvate kinase was increased in the liver of OLETF rats compared with control rats. There was no such remarkable change in liver-type phosphofructokinase. The activities of glucose-6-phosphatase and fructose-1,6-biphosphatase also increase despite high plasma levels of glucose and insulin. The activity of phosphoenolpyruvate carboxykinase did not show any significant change. The mRNA levels for fructose-1,6-biphosphatase, and phosphoenolpyruvate carboxykinase exhibited no marked changes. These results suggest that the expression of glucose-6-phosphatase and fructose-1,6-biphosphatase is disordered in OLETF rats.

Animals↗
PubMed1996

Sequencing of a 23 kb fragment from Saccharomyces cerevisiae chromosome VI.

Plasmid clone gapB and lambda phage clone 4682, which contain fragments of Saccharomyces cerevisiae chromosome VI, were analysed. A 23 kb sequence was determined and ten open reading frames (ORFs) were revealed. Among them, five ORFs were identical to five yeast genes (SEC4, MSH4, SPB4, DEG1 and NIC96), two were identical to transposable elements (TYA and TYB), one (gapBorfF003) was highly homologous to a yeast expressed sequence tag, and another (4682orfF002) was predicted to be a nuclear protein. Sequence data have been submitted to DDBJ/EMBL/GenBank data library under Accession Number D44604 (clone gapB) and D44600 (clone 4682), respectively.

Amino Acid Sequence↗
PubMed1996

Disordered expression of glycolytic and gluconeogenic liver enzymes of juvenile visceral steatosis mice with systemic carnitine deficiency.

A quantitative study of the effect of carnitine deficiency on expression of glycolytic and gluconeogenic enzymes was performed using juvenile visceral steatosis mice which are systemically deficient in carnitine. The amounts of glucokinase and L-type pyruvate kinase mRNA were reduced in homozygotes, compared to heterozygotes and normal controls at 2 and 8 weeks. Liver-type phosphofructokinase, however, did not differ significantly. The abundance of fructose 1,6-bisphosphatase mRNA was unchanged at 2 and 8 weeks. The level of phosphoenolpyruvate carboxykinase mRNA was increased slightly at 2 weeks, but not at 8 weeks. A part of these changes could not be explained by the plasma glucose or insulin level. Carnitine administration restored the mRNA of these enzymes to normal levels. These results suggest that carnitine deficiency affects the expression of these liver enzymes.

Age Factors↗
PubMed1996

Increased expression of carnitine palmitoyltransferase I gene is repressed by administering L-carnitine in the hearts of carnitine-deficient juvenile visceral steatosis mice.

The juvenile visceral steatosis (JVS) mouse is a novel mutant animal for studying systemic carnitine deficiency. The importance of the model has been pointed out in carnitine-deficient cardiac hypertrophy, since cardiomyopathy has been often improved after oral carnitine therapy in human systemic carnitine deficiency. To understand the effects of carnitine deficiency on gene expression in the heart, we tried to find the genes regulated by carnitine by means of a modified differential display procedure. Carnitine palmitoyltransferase I (CPT I) was one of the isolated genes. The level of CPT I gene expression in the ventricles of the JVS mice was at least three- to sixfold that of normal mice as judged by reverse transcription-polymerase chain reaction (RT-PCR). When the JVS mice were treated with carnitine, CPT I gene expression was repressed to the level of normal mice. Therefore, the increased expression of the CPT I gene was associated with carnitine deficiency.

Amino Acid Sequence↗
PubMed1996

HTLV-I uveitis.

Human T-cell lymphotropic virus type I (HTLV-I) is known to cause adult T-cell leukemia/T-cell lymphoma and tropical spastic paraparesis/HTLV-I-associated myelopathy. Recent seroepidemiologic, clinical, and virologic studies indicate that the virus is also related to a certain type of uveitis, which has been classified as uveitis without defined etiologies or idiopathic uveitis. According to the seroepidemiologic survey, the seroprevalence of HTLV-I in patients with idiopathic uveitis was significantly higher than that of two control groups, that is, patients with uveitis with defined etiologies and patients with nonuveitic ocular diseases. Clinically, the uveitis seen in HTLV-I carriers is characterized by moderate to severe cellular infiltration in the eye and by moderate retinal vasculitis, and the intraocular inflammation responds well to corticosteroid therapy. Interestingly, 25% of female patients with the disease had a previous history of Graves disease with hyperthyroidisms. The following virologic, molecular biologic findings suggest that cytokines produced by HTLV-I-infected T cells in the eye play the central role in the pathogenic mechanisms of the uveitis: (a) the virus load in the peripheral blood monocytes analyzed by the quantitative polymerase chain reaction methods was significantly greater in patients with the uveitis than in asymptomatic carriers, (b) the proviral DNA of HTLV-I and the gene expression of the virus at the mRNA level was detected in the infiltrating cells from the eyes of the patients, (c) the virus particles were detected by electron-microscopic examination in the T-cell clones established from the intraocular fluid of the patients, and (d) the HTLV-I-infected T cells produced a variety of cytokines without any stimuli, such as interleukin (IL)-1 alpha, IL-2, IL-3, IL-6, IL-8, IL-10, tumor necrosis factor alpha, interferon-gamma, and granulocyte-macrophage colony-stimulating factor. Based on the seroepidemiologic, clinical, and virologic data, the uveitis seen in HTLV-I carriers is considered to be a distinct clinical entity related to HTLV-I infection, and the disease is designated as HTLV-I uveitis.

Adolescent↗