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Biomedical subjects

E Takeda

Publications and source records attributed to E Takeda.

At least 73 records · Page 4Linked to original sources
PubMed1997

The predominant contribution of oligopeptide transporter PepT1 to intestinal absorption of beta-lactam antibiotics in the rat small intestine.

Although recent evidence suggests that certain beta-lactam antibiotics are absorbed via a specific transport mechanism, its nature is unclear. To confirm whether peptide transport in the rat can be largely ascribed to the intestinal oligopeptide transporter PepT1, the transporter has been functionally characterized and its significance in the intestinal absorption of beta-lactam antibiotics was evaluated. For evaluation of transport activity complementary RNA (cRNA) of rat PepT1 was synthesized in-vitro and expressed in Xenopus laevis oocytes. cRNA induced uptake of several beta-lactam antibiotics and the dipeptide [14C]glycylsarcosine; this was specifically inhibited by various dipeptides and tripeptides but not by their constituent amino acids or by tetra- or pentapeptides. The transport activity of PepT1 for beta-lactam antibiotics correlated well with their in-vivo intestinal transport and absorption. Furthermore, mutual inhibitory effects on uptake were observed between glyclsarcosine and beta-lactam antibiotics. Hybrid depletion of the functional expression of rat PepT1 in oocytes injected with rat intestinal epithelial total mRNA was studied using an antisense oligonucleotide corresponding to the 5'-coding region of PepT1. In oocytes injected with rat mRNA pre-hybridized with the antisense oligonucleotide against rat PepT1, the uptake of [14C]glycylsarcosine was almost completely abolished, whereas its uptake was not influenced by a sense oligonucleotide for the same region of PepT1. Similarly, the uptake of beta-lactam antibiotics was also reduced by the antisense oligonucleotide against rat PepT1. These results demonstrate that the intestinal proton-coupled oligopeptide transporter PepT1 plays a predominant role in the carrier-mediated intestinal absorption of beta-lactam antibiotics and native oligopeptides in the rat.

Animals↗
PubMed1997

Structural organization of the human vitamin D receptor chromosomal gene and its promoter.

The vitamin D receptor (VDR) is known to mediate the pleiotropic biological actions of 1,25-dihydroxyvitamin D3 through its ability to modulate the expression of target genes. The regulation of this ligand-activated cellular transcription factor is reported to occur at both transcriptional and posttranslational levels. To begin to address the molecular basis by which the VDR gene is regulated transcriptionally, we report here an initial characterization of the human VDR gene and its promoter. We isolated several overlapping A-phage and cosmid clones that cover more than 100 kb of human DNA and contained the entire VDR gene. The gene is comprised of 11 exons that, together with intervening introns, span approximately 75 kb. The noncoding 5'-end of the gene includes exons 1A, 1B, and 1C. Eight additional exons (exons 2-9) encode the structural portion of the VDR gene product. While primer extension and S1 nuclease-mapping studies reveal several common transcriptional start sites, three unique mRNA species are produced as a result of the differential splicing of exons 1B and 1C. The DNA sequence lying upstream of exon 1A is GC rich and does not contain an apparent TATA box. Several potential binding sites for the transcription factor SP1 and other activators are evident. Fusion of DNA fragments containing putative promoter sequences upstream of the luciferase structural gene followed by transient transfection of these plasmids into several mammalian cell lines resulted in significant reporter activity. Due to the size and complexity of the 5'-end of the VDR gene, we examined the activity of a DNA fragment surrounding exon 1C. An intron fragment 3' of exon 1C conferred retinoic acid responsivity when fused to a reporter gene plasmid, suggesting a molecular mechanism for the previously observed ability of retinoic acid to induce the VDR. The recovery of the gene for the human VDR will enable further studies on the transcriptional regulation of this gene.

Bacteriophage lambda↗
PubMed1997

A vitamin D receptor gene polymorphism in the translation initiation codon: effect on protein activity and relation to bone mineral density in Japanese women.

The effect of a T-C transition polymorphism at the translation initiation codon of the human vitamin D receptor (VDR) gene on the biological function of the encoded protein was investigated. Of 239 Japanese women volunteers subjected to genotype analysis for this polymorphism, 32 (13%) were genotype MM (the M allele is ATG at the putative translation start site), 75 (31%) were genotype mm (the m allele is ACG at the putative translation start site), and 132 (55%) were genotype Mm. The bone mineral density (BMD) in the lumbar spine (L2-L4) was determined for 110 healthy premenopausal women from the volunteers and was shown to be 12.0% greater (p < 0.05) for mm homozygotes than for MM homozygotes. Synthesis of the proteins by the M and m alleles from the cloned cDNAs in vitro and in transfected COS-7 cells revealed them to have a size of 50 and 49.5 kD, respectively, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. This size difference is consistent with initiation of translation of the M allele-encoded protein from an ATG codon located at nucleotides +10 to +12 in the conventional open reading frame. The extent of vitamin D-dependent transcriptional activation of a reporter construct under the control of a vitamin D response element in transfected HeLa cells was approximately 1.7-fold greater for the m type VDR than for the M type protein. These results suggest that the polymorphism at the translation start site of the VDR gene may modulate BMD in premenopausal Japanese women.

Adult↗
PubMed1997

[Reflexions on the functional method of nursing].

The main target of the present essay is to carry out a reflexion on the adoption of the functional method in nursing work, as well as on the criticism to this method. A secondary aim will be to show how the nursing team has been "using" this method in their daily work routine in the search for a more humanized nursing care.

Humanism↗
PubMed1997

[Structural polymorphism of ACTBP2 STR region].

We investigated structural polymorphisms of the ACTBP2-STR region by sequence analysis instead of using PCR-amplified fragment length polymorphisms. A gel spot extraction reamplification method using 2% agarose gel was effective for preparing the individual alleles from zygotes with a length difference of more than 16bp. In a survey of polymorphisms in 116 unrelated Japanese subjects, alleles from 73 zygotes were prepared by this method. The STR structure of the allele was determined from the forward sequence image. The allele designation was made according to the number of the repeated AAAG tetranucleotide and also the position of the inserted AA or AG dinucleotide. Forty-three zygotes impossible to separate were first sequenced in a forward direction. Among them, 28 zygotes were typed directly by analysing the sequence image which exhibited a peculiar overlap of two sequences. For the remaining 15 zygotes, it was possible to type by analysing the forward and reverse sequence images together. This investigation revealed the presence of 79 kinds of alleles in 116 zygotes of ACTBP2. The most frequent allele was type 19, which accounted for 9.5% although this was still a small portion. Genotypes 19/20, 19/21, 18/21, 18/10AA12, 19/13AA11 and 20/13AA15 were detected from two subjects, respectively, and others including the type 19 homozygyote were unique.

Alleles↗
PubMed1997

Comparison of energy metabolism in insulin-dependent and non-insulin-dependent diabetes mellitus.

To compare the metabolic consequences of insulin-dependent diabetes mellitus (IDDM) and non-insulin-dependent diabetes mellitus (NIDDM), glycemic control and energy metabolism were evaluated in 18 children displaying IDDM and 19 NIDDM adult patients. With rising concentrations of fasting blood glucose (FBG), hemoglobin A1C and free fatty acid, the percentage of the ratio of resting energy expenditure (REE) to predicted REE expressed as %REE increased and the respiratory quotient (RQ) decreased. The linear regression between RQ and FBG showed the same gradient in IDDM and NIDDM although the RQ in IDDM was always 0.07 lower than that in NIDDM given various FBG concentrations. Those patients whose RQ values were less than 0.7, indicating ketone body production, included 8 (44%) IDDM and 2 (11%) NIDDM patients. These results may explain the relatively greater manifestation of ketoacidosis in IDDM.

Adult↗
PubMed1997

Repair of a cheek defect with the tissue expander method: a case report.

A 28-year-old female had a severe depressed deformity of the left cheek due to false local injection of ZnCl2 into her buccal mucosa instead of an anesthetic. After several attempts at plastic surgery in another hospital with unsatisfactory results, she was treated surgically in our clinic using cheek skin expanded by a tissue expander. The outcome was satisfactory.

Adult↗
PubMed1997

Mechanism of substrate recognition by the ribotoxin, alpha-sarcin.

The substrate recognition and catalytic mechanisms of alpha-sarcin were explored with kinetic method by using synthetic 25-mer RNA mimicking the alpha-sarcin/ricin loop in 23S rRNA of E. coli ribosomes. The oligomer containing deoxy-G at the site of alpha-sarcin (G14) was a potent competitive inhibitor. The RNA having deoxy-G8 however, increases the Kcat value by about five times but without significant alteration on Km. Surprisingly, the deletion of G8 makes the oligomer become a strong noncompetitive inhibitor of the enzyme. These results suggested that there are at least two sites in the RNA substrate which are recognized by alpha-sarcin, one is the G8 bulge or at around its neighbor and the other is the GAGA in the sarcin/ricin loop of the rRNA.

Base Sequence↗
PubMed1996

Vitamin D3 elicits calcium response and activates blood monocyte-derived macrophages from patients with vitamin D dependent rickets type II.

We studied the effects of vitamin D3 metabolites on intracellular free Ca2+ concentration ([Ca2+]i) and the respiratory burst of monocyte-derived macrophages (MDM) from patients with vitamin D dependent rickets type II. Treatment of MDM from the patients and healthy donors with 1 nM 1,25(OH)2D3 produced a rapid elevation of [Ca2+]i and similarly primed both types of cells for enhanced capacity for O2- release with phorbol diester. These results suggest that macrophages may have distinct non-genomic pathways of vitamin D3, which partly explain the absence of immunodeficiency and the disappearance of rickets after treatment with vitamin D3 in the patients.

24,25-Dihydroxyvitamin D 3↗
PubMed1996

Effects of truncation of the COOH-terminal region of a Na+-independent neutral and basic amino acid transporter on amino acid transport in Xenopus oocytes.

To determine the role of a neutral and basic amino acid transporter (NBAT) in amino acid transport, we microinjected several COOH-terminal deletion mutants of NBAT cRNA into Xenopus oocytes and measured transport activity for arginine, leucine, and cystine in the presence and absence of sodium. Wild-type NBAT significantly stimulated the uptake of all three amino acids 10-20-fold compared with controls. On the other hand, no mutant, except a Delta511-685 mutant, stimulated the uptake of these amino acids. The Delta511-685 mutant significantly increased the uptake of arginine. In the presence of sodium, the Delta511-685 mutant also increased the uptake of leucine. The Delta511-685 mutant did not stimulate cystine uptake in the presence or absence of sodium. The stimulation of arginine uptake by the Delta511-685 mutant was inhibited by a 100-fold excess of unlabeled leucine in the presence of sodium. Inhibition of L-arginine uptake by L-homoserine was seen only in the presence of sodium, and an increase in the inhibition of L-arginine uptake by L-histidine was seen when the extracellular pH was decreased. Furthermore, an inward current in oocytes injected with the Delta511-685 mutant was recorded electrophysiologically when basic amino acids were applied. Homoserine was also taken up, but sodium was necessary for their transport. These properties of the Delta511-685 mutant correspond to those of the y+ amino acid transporter. If NBAT is a component of the b0,+-like amino acid transport system, it is unlikely that a mutant protein (Delta511-685) is able to stimulate an endogenous y+-like transport system. These results suggest that NBAT functions as a activator of the amino acid transport system in Xenopus oocytes.

Amino Acid Sequence↗
PubMed1996

The basic amino acid transporter (rBAT)-like immunoreactivity in paraventricular and supraoptic magnocellular neurons of the rat hypothalamus.

In the rat hypothalamus, the basic amino acid transporter (rBAT)-like immunoreactivity was analyzed by immunohistochemistry using an antibody against the 15-amino acid sequence of the deduced rat rBAT protein. In the supraoptic and the paraventricular nuclei, magnocellular neurons exhibited the marked rBAT-like immunoreactivity in intracellular structures but not in the plasma membrane. The results suggest that the rBAT serves as an intracellular amino acid transport system in magnocellular neurons.

Amino Acid Sequence↗
PubMed1996

Sequence, tissue distribution and developmental changes in rat intestinal oligopeptide transporter.

Complementary DNA clones encoding the rat PepT1 small-intestinal oligopeptide transporter were isolated from a jejunal library by cross-hybridization with a rabbit PepT1 cDNA probe. The cDNA sequence indicates that rat PepT1 is composed of 710 amino acids and shows 77% and 83% amino acid sequence identity with rabbit and human PepT1, respectively. Northern blot analysis detected rat PepT1 mRNA in the small intestine and kidney. Intestinal PepT1 mRNA levels were highest in 4-day old rats, and then decreased reaching the adult level by day 28 after birth. These results indicate that the expressions of PepT1 gene change markedly during development.

Age Factors↗
PubMed1995

Cloning and functional expression of a Na(+)-dependent phosphate co-transporter from human kidney: cDNA cloning and functional expression.

A cDNA clone encoding a protein 69% identical in amino acid sequence with that of the Na/P(i) co-transporter NaP(i)-1 was isolated from a human kidney cDNA library. The DNA sequence was identical with that of NPT-1 cDNA published by Chong, Kristjansson, Zoghbi and Hughe (1993) (Genomics, 18, 355-359). In the present study, we have characterized the function of the encoded protein and the tissue distribution of its mRNA. Injection of RNA transcribed from NPT-1 into Xenopus oocytes resulted in expression of Na/P(i) co-transport activity showing a high affinity for P(i) transport (Km 0.29 mM). Kinetic characterization ([P(i)], [Na+]) demonstrated that the expressed transport activity has properties similar to those displayed by oocytes injected with human kidney poly(A)+ RNA. Northern blotting demonstrated that NPT-1 mRNA is expressed in renal cortex, liver and brain but not in other tissues. Hybrid depletion with antisense oligonucleotides to NaP(i)-3 and NPT-1 completely inhibited poly(A)+ RNA-induced Na(+)-dependent P(i) uptake in oocytes. These findings indicate that two high-affinity Na/P(i) cotransporters (NaP(i)-3 and NPT-1) are present in human kidney cortex.

Animals↗
PubMed1995

Molecular genetic analysis of a female patient with pyruvate dehydrogenase deficiency: detection of a new mutation and differential expression of mutant gene product in cultured cells.

A new 18 bp insertion mutation in the gene for the alpha subunit of pyruvate dehydrogenase (E1 alpha) was found in a female patient with congenital lactic acidaemia. Cultured skin fibroblasts and Epstein-Barr virus-transformed lymphoblastoid cells from this patient showed decreased and normal pyruvate dehydrogenase complex (PDHC) activity, respectively. This 18 bp insertion was a de novo mutation, because it was not present in her parents. Although this female patient was heterozygous for the normal and the mutant alleles, 97% of cultured skin fibroblasts expressed the mutant allele, while 100% of cultured lymphoblastoid cells, 94% of peripheral blood lymphocytes and 98% of IL-2-activated T-cells expressed the normal allele. These results suggest that in this patient the X chromosome containing the normal allele was predominantly inactivated in fibroblasts and the X chromosome containing the mutant allele was predominantly inactivated in lymphocytes. The diagnosis of E1 alpha deficiency is usually established by measurement of PDHC activity and the level of immunoreactive proteins. However, these methods are not sufficient to diagnose the disorder in female patients with E1 alpha deficiency due to differential inactivation of the X chromosome. Therefore, it is necessary to develop a new method to firmly establish the diagnosis of E1 alpha deficiency.

Alleles↗
PubMed1995

[Sequence polymorphism of mitochondrial DNA and its forensic application].

We have investigated polymorphism of mitochondrial (mt) DNAs in the Japanese population. In order to compare 288-bp sequences (nucleotide positions 16,111 to 16,398) in the noncoding region, a 452-bp segment elongated by 82 bp at both sides of the target was amplified by PCR and analyzed directly by Taq cycle sequencing with FITC-labelled primers. A survey of 100 Japanese individuals revealed the existence of 66 types of mtDNAs. Among them, 10 types including two frequent types were shared by more than one individual. During this investigation, it was found that 19 types (from 25 individuals) possessed a common nucleotide replacement of T by C at position 16,189, by which a "C-continuous stretch" was formed, and that clear reading of their sequences was restricted in the region lying ahead of the C stretch on either forward or reverse sequencing. Cloning analysis of PCR product possessing the C stretch demonstrated that the length of the C stretch was heterogeneous. Consequently, we propose the following rules for mtDNA sequencing and its forensic application: 1) the replacement of T by C at position 16,189 is acceptable as a variation factor. 2) The number of Cs in the C stretch is tentatively fixed at 10.

Asian People↗