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T Ohta

Publications and source records attributed to T Ohta.

At least 73 records · Page 4Linked to original sources
PubMed2001

An absolute requirement for STAT4 and a role for IFN-gamma as an amplifying factor in IL-12 induction of the functional IL-18 receptor complex.

IL-12 and IL-18 are both proinflammatory cytokines that contribute to promoting Th1 development and IFN-gamma expression. However, neither IL-12R nor IL-18R is expressed as a functional complex on most resting T cells. This study investigated the molecular mechanisms underlying the induction of an IL-18R complex in T cells. Resting T cells expressed IL-18Ralpha chains but did not exhibit IL-18 binding sites as detected by incubation with rIL-18 followed by anti-IL-18 Ab, suggesting a lack of IL-18Rbeta expression in resting T cells. Although they also failed to express IL-12R, stimulation with anti-CD3 plus anti-CD28 generated IL-12R. Exposure of these cells to IL-12 led not only to up-regulation of IL-18Ralpha expression but also to induction of IL-18R binding sites on both CD4(+) and CD8(+) T cells concomitant with IL-18Rbeta mRNA expression. The IL-18 binding site represented a functional IL-18R complex capable of exhibiting IL-18 responsiveness. IL-12 induction of an IL-18R complex and IL-18Rbeta mRNA expression was not observed in STAT4-deficient (STAT4(-/-)) T cells and was substantially decreased in IFN-gamma(-/-) T cells. However, the failure of STAT4(-/-) T cells to induce an IL-18R complex was not corrected by IFN-gamma. These results indicate that STAT4 and IFN-gamma play an indispensable role and a role as an amplifying factor, respectively, in IL-12 induction of the functional IL-18R complex.

Adjuvants, Immunologic↗
PubMed2001

Two-domain hemoglobin gene of the water flea Moina macrocopa: duplication in the ancestral Cladocera, diversification, and loss of a bridge intron.

Two cDNAs encoding the two-domain hemoglobin (Hb) chains of a crustacean Cladocera, Moina macrocopa, were cloned and their nucleotide (nt) sequences were determined. The amino acid (aa) sequences of both the gene products deduced from the nt sequences consisted of 348 residues and showed 98% identity with each other. These sequences together with the NH(2)-terminal aa sequences of the Hb chains determined after separation by two-dimensional gel electrophoresis showed that the Hb chains are synthesized as a secretory precursor with a signal peptide of 17 aa residues. The aa sequences of M. macrocopa Hb chains shared the following features with those of Daphnia Hb chains. Firstly, the signal peptide is followed by an NH(2)-terminal extension containing a threonine-rich sequence that might play a role in the multimerization of subunit chains. Secondly, the identity between the aa sequences of the first and second domains is exceptionally low. These facts suggest that duplication of the cladoceran Hb gene occurred before the divergence of families Moinidae and Daphniidae. Analysis of genomic DNA showed that the M. macrocopa Hb genes consist of two large repeated regions, encoding the first and second domains of Hb chains, respectively. The intron-exon organization of the first region of the M. macrocopa Hb genes was similar to that found in the Daphnia Hb genes, having the three-exon, two-intron structure characteristic of animal Hb genes. However, the intron bridging the two regions and the most downstream intron in the second region were missing in the Moina genes, providing a new example of intron loss. The following elements in the 5' flanking region were conserved in the Moina and Daphnia genes: (1) TATAAA, a typical TATA box sequence accompanied by a downstream sequence, GAAXAGCATCAGTT (the fourth residue X was G or A in Daphnia and absent in Moina); (2) CCAAT boxes, located upstream of the TATA box; (3) the binding sites for HIF-1 and GATA-1, also located upstream of the TATA box, that may be responsible for up-regulation of the cladoceran Hb genes under hypoxia.

Amino Acid Sequence↗
PubMed2001

Marked reduction of Helicobacter pylori-induced gastritis by urease inhibitors, acetohydroxamic acid and flurofamide, in Mongolian gerbils.

Urease has been suggested to be essential for colonization and pathogenesis of Helicobacter pylori infection. In the present study, we evaluated the effects of urease inhibitors [acetohydroxamic acid (AHA) and flurofamide (FFA)] on H. pylori-induced gastritis in Mongolian gerbils. Animals were orally inoculated with H. pylori, and given urease inhibitors in their diet throughout the experimental period of six weeks or four weeks, starting from two weeks after H. pylori inoculation. With the administration of AHA at doses of 100, 500, and 2500 ppm throughout the experimental period, H. pylori-induced gastritis in animals was decreased in a dose-dependent manner, significantly so at 2500 ppm. Suppression of gastric lesions was also evident in animals administered 2500 ppm AHA after the H. pylori infection. Bacterial infection rates were reduced to 40-50% of the control value of 100%, by the highest dose of AHA. The potent urease inhibitor, FFA, also caused marked amelioration of H. pylori-associated gastritis on administration at 100 ppm throughout the six-week experimental period or for four weeks after H. pylori infection. Animals treated with FFA had few visible gastric lesions, and the proportion infected with H. pylori was reduced to less than 10%. Since antibiotic-resistant strains of H. pylori have become a serious problem, nonantibiotic urease inhibitors may be very useful to control H. pylori-associated gastroduodenal disease.

Administration, Oral↗
PubMed2001

Optical anisotropy of oxidized Si(001) surfaces and its oscillation in the layer-by-layer oxidation process.

Reflectance-difference (RD) measurements for the oxidation of single-domain (2x1)-reconstructed Si(001) surfaces show that the polarity of the interface-induced optical anisotropy is reversed repeatedly with increasing oxide thickness. The oscillation of the RD amplitude, which we show is due to layer-by-layer progression of the oxidation, has allowed us to count the number of oxidized Si layers in situ during oxidation. The origins of the observed spectral line shape are discussed.

Journal Article↗
PubMed2001

A theoretical study of alcohol oxidation by ferrate.

The conversion of methanol to formaldehyde mediated by ferrate (FeO(4)2-), monoprotonated ferrate (HFeO4-), and diprotonated ferrate (H2FeO4) is discussed with the hybrid B3LYP density functional theory (DFT) method. Diprotonated ferrate is the best mediator for the activation of the O-H and C-H bonds of methanol via two entrance reaction channels: (1) an addition-elimination mechanism that involves coordination of methanol to diprotonated ferrate; (2) a direct abstraction mechanism that involves H atom abstraction from the O-H or C-H bond of methanol. Within the framework of the polarizable continuum model (PCM), the energetic profiles of these reaction mechanisms in aqueous solution are calculated and investigated. In the addition-elimination mechanism, the O-H and C-H bonds of ligating methanol are cleaved by an oxo or hydroxo ligand, and therefore the way to the formation of formaldehyde is branched into four reaction pathways. The most favorable reaction pathway in the addition-elimination mechanism is initiated by an O-H cleavage via a four-centered transition state that leads to intermediate containing an Fe-O bond, followed by a C-H cleavage via a five-centered transition state to lead to formaldehyde complex. In the direct abstraction mechanism, the oxidation reaction can be initiated by a direct H atom abstraction from either the O-H or C-H bond, and it is branched into three pathways for the formation of formaldehyde. The most favorable reaction pathway in the direct abstraction mechanism is initiated by C-H activation that leads to organometallic intermediate containing an Fe-C bond, followed by a concerted H atom transfer from the OH group of methanol to an oxo ligand of ferrate. The first steps in both mechanisms are all competitive in energy, but due to the significant energetical stability of the organometallic intermediate, the most likely initial reaction in methanol oxidation by ferrate is the direct C-H bond cleavage.

Journal Article↗
PubMed2001

Ethidium bromide and SYBR Green I enhance the genotoxicity of UV-irradiation and chemical mutagens in E. coli.

Ethidium bromide (EtBr) and SYBR Green I are nucleic acid gel stains and used commonly in combination with UV-illumination. EtBr preferentially induces frameshift mutations but only in the presence of an exogenous metabolic activation system, while SYBR Green I is a very weak mutagen that induces frameshift mutations. We found that EtBr and SYBR Green I, without an added metabolic activation system, strongly potentiated the base-substitution mutations induced by UV-irradiation in E. coli B/r WP2 cells. Each DNA stain alone showed no mutagenicity to the strain. Base-substitutions induced by 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX) and 4-nitroquinoline-1-oxide were similarly potentiated by EtBr and SYBR Green I. SYBR Green I had a much greater effect. No enhancing effects were observed on mutations induced by mitomycin C, cisplatin, transplatin, cumene hydroperoxide, base analogs, and alkylating agents. Another DNA stain, acridine orange, showed similar enhancing effects on UV- and MX-mutagenicity, but no effect was observed for 4',6-diamidino-2-phenylindole (DAPI). UV- and MX-induced mutations in E. coli WP2s (uvrA), which is defective in nucleotide excision repair activity, were not potentiated by the addition of EtBr, SYBR Green I, or acridine orange. Those nucleic acid stains might inhibit the nucleotide excision repair of DNA damaged by UV and MX treatment.

Acridine Orange↗
PubMed2001

Structure of the human acyl-CoA:cholesterol acyltransferase-2 (ACAT-2) gene and its relation to dyslipidemia.

Acyl-CoA:cholesterol acyltransferase (ACAT) catalyzes cholesterol esterification in mammalian cells. Two isoforms of ACAT have been reported to date (ACAT-1 and ACAT-2). ACAT-1 is ubiquitously expressed in tissues except the intestine. In contrast, ACAT-2 is expressed mainly in the intestine in humans. To investigate the relationship between ACAT-2 and dyslipidemia, we determined the structure of the human ACAT-2 gene and then studied the relationship between mutations of the ACAT-2 gene and dyslipidemia. To isolate human ACAT-2 genomic DNA, we designed primers based on the human ACAT-2 cDNA sequence: forward primer 5'-ACACCTCGATCTTGGTCCTGCCATA-3' and reverse primer 5'-GGAATGCAGACAGGGAGTCCT-3'. Using these primers, a human P1-derived artificial chromosome (PAC) library was screened by PCR-based procedures. Isolated PAC clones were completely digested with BamHI and subcloned into plasmid vector. Subclones that contained exons were screened by dot-blot hybridization using partial ACAT-2 cDNA fragments. The coding region of the ACAT-2 gene was encoded in 15 exons from 51 to 265 base pairs on a 21 kilobase span of genomic DNA. The exonic sequences coincided completely with that of ACAT-2 cDNA, and each exon-intron junction conserved splicing consensus sequences. Next, 187 (91 dyslipidemic and 96 normolipidemic) subjects were screened by PCR single-strand conformational polymorphism analysis of the ACAT-2 gene. Three mutations were identified by DNA sequencing: two missense mutations (E14G in exon 1 and T254I in exon 7) and a point mutation in intron 7 (-35G-->A). Mutations in exon 1 and intron 7 were not associated with plasma concentrations of lipids and apolipoproteins (apo). However, plasma apoC-III levels in T254I heterozygotes were significantly higher than those in subjects without mutation. Plasma triglyceride (TG) levels in T254I heterozygotes were similar to those in subjects without mutation. Although further studies are needed, our data suggest that ACAT-2 may contribute to apoC-III gene expression and the assembly of apoC-III and TG, possibly in the intestine.

Apolipoproteins↗
PubMed2001

Whole genome sequencing of meticillin-resistant Staphylococcus aureus.

BACKGROUND: Staphylococcus aureus is one of the major causes of community-acquired and hospital-acquired infections. It produces numerous toxins including superantigens that cause unique disease entities such as toxic-shock syndrome and staphylococcal scarlet fever, and has acquired resistance to practically all antibiotics. Whole genome analysis is a necessary step towards future development of countermeasures against this organism. METHODS: Whole genome sequences of two related S aureus strains (N315 and Mu50) were determined by shot-gun random sequencing. N315 is a meticillin-resistant S aureus (MRSA) strain isolated in 1982, and Mu50 is an MRSA strain with vancomycin resistance isolated in 1997. The open reading frames were identified by use of GAMBLER and GLIMMER programs, and annotation of each was done with a BLAST homology search, motif analysis, and protein localisation prediction. FINDINGS: The Staphylococcus genome was composed of a complex mixture of genes, many of which seem to have been acquired by lateral gene transfer. Most of the antibiotic resistance genes were carried either by plasmids or by mobile genetic elements including a unique resistance island. Three classes of new pathogenicity islands were identified in the genome: a toxic-shock-syndrome toxin island family, exotoxin islands, and enterotoxin islands. In the latter two pathogenicity islands, clusters of exotoxin and enterotoxin genes were found closely linked with other gene clusters encoding putative pathogenic factors. The analysis also identified 70 candidates for new virulence factors. INTERPRETATION: The remarkable ability of S aureus to acquire useful genes from various organisms was revealed through the observation of genome complexity and evidence of lateral gene transfer. Repeated duplication of genes encoding superantigens explains why S aureus is capable of infecting humans of diverse genetic backgrounds, eliciting severe immune reactions. Investigation of many newly identified gene products, including the 70 putative virulence factors, will greatly improve our understanding of the biology of staphylococci and the processes of infectious diseases caused by S aureus.

Animals↗
PubMed2001

In vitro ubiquitination of cyclin D1 by ROC1-CUL1 and ROC1-CUL3.

Overexpression of cyclin D1 has been implicated in a variety of tumors, such as breast cancers, gastrointestinal cancers and lymphomas. Both gene amplification and protein degradation mediated by ubiquitin (Ub)-dependent proteolysis regulate the abundance of cyclin D1. Here we report that ROC1 interacted with all three D type cyclins in vivo but did not bind to other cyclins tested. The ROC1-CUL1 and ROC1-CUL3, but not ROC1-CUL2, -CUL3 and -CUL4, immunocomplexes promoted polyubiquitination of bacterially purified cyclin D1 in vitro. RING finger mutations of ROC1 eliminated the Ub ligase activity toward cyclin D1. In all cases the ubiquitination of cyclin D1 was accompanied by autoubiquitination of the cullins. The results suggest the involvement of ROC1-cullin ligases in cyclin D1 ubiquitination and a potential mechanism whereby the cullin subunit is ubiquitinated itself while ubiquitinating a substrate.

Anaphase-Promoting Complex-Cyclosome↗
PubMed2001

Vasopressin-induced contraction in the rat basilar artery in vitro.

Vasopressin ([Arg(8)]vasopressin)-induced contraction was characterized using receptor agonists and antagonists for vasopressin and channel blockers in the rat basilar artery ring preparations. Vasopressin induced rhythmic contractions superimposed on a contraction in endothelium-intact preparations but not in denuded ones. Endothelium removal shifted the concentration-response curve for vasopressin leftward and upward. In endothelium-denuded preparations, vasopressin V(1) receptor antagonist shifted the concentration-response curve for vasopressin downward and rightward. Vasopressin V(1) receptor agonist caused contraction but V(2) receptor agonist did not. The contractile response to vasopressin was partly inhibited by nifedipine, SK&F 96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole) and niflumic acid. In the absence of extracellular Ca(2+), vasopressin produced a transient contraction. Charybdotoxin produced an upward and leftward shift of the concentration-response curve for vasopressin. These results suggest that vasopressin elicits contraction due to Ca(2+) influx through voltage-dependent and receptor-operated Ca(2+) channels and to Ca(2+) release from Ca(2+) stores by activating vasopressin V(1) receptors in the rat basilar artery.

Animals↗
PubMed2001

Novel adhesion prevention membrane based on a bioresorbable copoly(ester-ether) comprised of poly-L-lactide and Pluronic: in vitro and in vivo evaluations.

Block copolymers consisting of poly(L-lactide) (PLLA) and poly(oxyethylene-co-oxypropylene), with various compositions, were synthesized and characterized in vitro and in vivo for their application as postoperative adhesion prevention membranes. It was found that the flexibility and degradability of the cast films of the block copolymers grew with increasing Pluronic F68 [PN; poly(oxyethylene-co-oxypropylene] composition. The receding contact angle of the copolymer films against water became lower than that of the PLLA film, because the surface was predominantly covered with more hydrophilic PN segments in a wet state. This surface property significantly affects the cell attachment property of the copolymer films, and the fibroblasts cultured on the films exhibit a spheroid-like morphology. The copolymer films subcutaneously implanted in the back of rats induced milder tissue responses compared with PLLA homopolymers, because of the increased surface hydrophilicity in the former. In vivo evaluation using a uterus horn model in rats revealed that the performance of these copolymer films as an adhesion-prevention membrane is comparable to that of a conventionally utilized membrane of oxidized regenerated cellulose. These results indicate that the copolymer films are biocompatible materials with controllable mechanical properties and biodegradability as adhesion-prevention membranes.

3T3 Cells↗
PubMed2001

Effect of pre-and postoperative plasmapheresis on posttransplant recurrence of focal segmental glomerulosclerosis in children.

BACKGROUND: Posttransplant recurrence is frequent in patients who received renal transplantation for focal segmental glomerulosclerosis (FSGS). The recurrence has been ascribed to a circulating permeability factor or factors. We have used plasmapheresis (PP) to treat recurrent FSGS and also studied whether preoperative PP is effective in preventing recurrence of FSGS. METHODS: We retrospectively analyzed 21 allografts of 20 patients with nephrotic syndrome and biopsyproven FSGS. They were divided into two groups depending on whether they had prophylactic PP; a prophylactic (n=15) and a nonprophylactic group (n=6). PP was performed two to three times prophylactically and therapeutically until proteinuria was markedly reduced. In each session, 50-75 ml/kg of the patient's plasma was exchanged with 5-8% albumin. RESULTS: FSGS recurred in 9 of 21 allografts, 4 of 6 in the nonprophylactic group, and 5 of 15 in the prophylactic group. Therapeutic PP was performed in seven of nine recurrent patients without definite adverse effect, with satisfactory results except in one patient. Children lost proteinuria after 6 to > 100 sessions of PP and the number correlated with the pretreatment level of proteinuria. The mean follow-up periods were 62.7 and 41.6 months for the prophylactic and nonprophylactic groups, respectively. At the last follow-up, 66.7% of relapsing and 81.8% of nonrelapsing patients had a functioning graft. CONCLUSION: PP appears to be effective for the prevention and treatment of posttransplant recurrence of FSGS, although further consideration of cost/benefit and risks is required before a conclusive judgement can be made.

Child↗
PubMed2001

Addressing the stability of C-capped dipeptide efflux pump inhibitors that potentiate the activity of levofloxacin in Pseudomonas aeruginosa.

Synthetic optimization of a biologically labile class of dipeptides that function as efflux pump inhibitors to potentiate the antibacterial agent levofloxacin in Pseudomonas aeruginosa has led to the discovery of a related series of compounds that are completely stable in a variety of biological matrices. Other than the stability profile, the in vitro profile of the new series is essentially identical to that observed with the original one. A prototypical compound from the new series demonstrates potentiation in an in vivo model of infection.

Animals↗
PubMed2001

The RING heterodimer BRCA1-BARD1 is a ubiquitin ligase inactivated by a breast cancer-derived mutation.

BRCA1-BARD1 constitutes a heterodimeric RING finger complex associated through its N-terminal regions. Here we demonstrate that the BRCA1-BARD1 heterodimeric RING finger complex contains significant ubiquitin ligase activity that can be disrupted by a breast cancer-derived RING finger mutation in BRCA1. Whereas individually BRCA1 and BARD1 have very low ubiquitin ligase activities in vitro, BRCA1 combined with BARD1 exhibits dramatically higher activity. Bacterially purified RING finger domains comprising residues 1-304 of BRCA1 and residues 25-189 of BARD1 are capable of polymerizing ubiquitin. The steady-state level of transfected BRCA1 in vivo was increased by co-transfection of BARD1, and reciprocally that of transfected BARD1 was increased by BRCA1 in a dose-dependent manner. The breast cancer-derived BARD1-interaction-deficient mutant, BRCA1(C61G), does not exhibit ubiquitin ligase activity in vitro. These results suggest that the BRCA1-BARD1 complex contains a ubiquitin ligase activity that is important in prevention of breast and ovarian cancer development.

BRCA1 Protein↗
PubMed2001

Characterization of ruthenium-dinitrogen tetraamine complexes by XAFS spectroscopy.

A reaction between N2 and trans-Ru(II)Cl2L (L=2,5,9,12-tetramethyl-2.5,9,12-tetraazatridecane) in aqueous solutions has been investigated by XAFS spectroscopy. Ru K-edge EXAFS clearly revealed that there exists an N2-bridged binuclear complex and that the Ru-Ru distance is 4.96 A with the coordination number of 0.5. The Ru K-edge energies of the N2 complexes are closer to that of Ru(III)Cl2L+ rather than that of Ru(II)Cl2L, indicating back donation from the Ru dpi to the N2 pi* orbitals.

Journal Article↗