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Biomedical subjects

K Miki

Publications and source records attributed to K Miki.

At least 163 records · Page 9Linked to original sources
PubMed1997

[Present status and advances in detecting Shiga toxin-producing Escherichia coli O157].

Currently, detection of Shiga toxin-producing Escherichia coli(STEC) in stool samples is based on the isolation method in most clinical laboratories. The procedures are as follows: i) isolation with selective agar plates, ii) biological test with differential media, iii) serological test of anti-O antisera, iv) detection of toxin or toxin gene. These procedures take 4 days, therefore more rapid method is required. In the near future, a rapid detection method that detects STEC directly from stool samples will be introduced. Polymerase-chain reaction (PCR), enzyme-linked immuno-sorbent assay (ELISA), detection of serum anti-O157 antibodies are now available in clinical laboratories. Result of PCR for detection Shiga toxin gene and serum anti-O157 antibodies are described. Fifteen stool and serum samples from patients suspected of STEC infection were examined. With the culture and PCR method, 2 patients were positive by both methods and the results were confirmed in both cases. Six patients were positive by the antibodies detection method. From these results, the PCR method using stool samples was useful as a rapid detection method in clinical laboratories. Detection of serum antibodies has been simplified and is not an expensive method. Therefore, the method is useful for clinical diagnosis of STEC infection, especially, for diagnosing HUS or after antimicrobial agents have been administered to patients.

Adolescent↗
PubMed1997

Asialoglycoprotein receptor and hepatic blood flow using technetium-99m-DTPA-galactosyl human serum albumin.

UNLABELLED: Asialoglycoprotein receptor (ASGP-R) amount and hepatic blood flow were quantitatively measured by using a newly developed kinetic model of 99mTc-labeled diethylenetriaminepentaacetic acid-galactosyl human serum albumin (99mTc-GSA) in which receptor-mediated endocytosis and receptor recycling were considered. METHODS: Five healthy volunteers were intravenously injected 3-mg and 9-mg 99mTc-GSA doses. The absolute amounts of 99mTc-GSA in the liver and extrahepatic blood were estimated from the time-activity curves for the liver, heart and lung. The metabolic process was represented by five differential equations with 10 parameters as variables. To estimate total receptor amount (Rtotal), hepatic plasma flow (Q) and hepatic plasma volume (Vh), other parameters were fixed and estimated by analyzing their data with the least-squares method. Nineteen patients with liver diseases were given a 3-mg dose, and the data were analyzed to estimate Rtotal, Q and Vh. RESULTS: The values of the fixed parameters were estimated as follows: dissociation constant, 0.032 microM; rate constant for internalization, 0.604 min(-1); and ratio of surface receptors to total receptors, 6.1%. The fitted liver uptake curve corresponded well to the measured data. The simulated liver uptake curve was significantly influenced by Rtotal and Q in cases with normal receptor amounts. Analysis in patients with normal livers, chronic hepatitis and liver cirrhosis showed statistically significant differences in their Rtotal values, but not in their Q or Vh values. The s.e. values of Rtotal, Q and Vh for normal livers were small, and the s.e. values of Q and Vh were high for cirrhotic livers. CONCLUSION: This method is useful for measuring ASGP-R amount and hepatic blood flow simultaneously based on dynamic images, without the need for blood sampling, and reflects the cellular transport of asialoglycoproteins and the ASGP-R recycling mechanism.

Adult↗
PubMed1996

Early induction of the NGFI-B/Nur77 family genes in nephritis induced by anti-glomerular basement membrane antibody.

We recently isolated a novel nuclear receptor NOR-1, which is a member of the steroid/thyroid receptor superfamily, and belongs to the NGFI-B/Nur77 family. In the present study, we examined gene expression of NOR-1 and its closely related members in nephritis induced by anti-glomerular basement membrane (GBM) antibody. The mRNA levels for NOR-1, NGFI-B and RNR-1 increased 24 h after injection of anti-GBM antibody (day 1). Gene expression of NOR-1 and NGFI-B reached maximum levels on day 3, gradually decreased thereafter and returned to control levels on day 28. RNR-1 reached a peak on day 7, and then decreased. Renal injuries were most prominent on day 7 and persisted until day 28, indicating that NOR-1, NGFI-B and RNR-1 genes are induced during the early stage of glomerulonephritis and may be associated with the progression of glomerulonephritis. The induction of the NGFI-B gene was less remarkable than that of NOR-1 and RNR-1. In addition, administration of glucocorticoid hormone suppressed NOR-1 and RNR-1 gene expression to almost normal levels, whereas NGFI-B gene expression was not significantly repressed. These findings also suggest that the NGFI-B/Nur77 family may possess different biological roles and NGFI-B might act as a general transcription factor in cell function.

Animals↗
PubMed1996

Structure, mapping and expression of a human NOR-1 gene, the third member of the Nur77/NGFI-B family.

We identified a human homologue of NOR-1 (neuron-derived orphan receptor) from the fetal brain. There are two transcripts for human NOR-1, encoding 626 amino acid residues with a calculated molecular mass of 68 kDa. The high homology between hNOR-1, mNur77/rNGFI-B/hTR3, and mNurr1/rRNR-1/hNOT indicated that these three orphan receptors form a distinct subfamily within the steroid/thyroid receptor superfamily. Human NOR-1 mRNA was detected in the adult heart and skeletal muscle as well as in the fetal brain, indicating that its expression is not restricted to events that occur during neural development. The hNOR-1 gene is more than 35 kilobases long and interrupted by seven introns. The exon-intron structure of the gene is generally conserved when compared with the steroid/thyroid receptor superfamily and is remarkably similar to that of the Nur77/NGFI-B genes. This suggests that the Nur77/NGFI-B family has evolved from a common ancestral gene. Fluorescence in situ hybridization (FISH) revealed that the gene is located on chromosome 9q.

Amino Acid Sequence↗
PubMed1996

Ventral tegmental injection of nicotine induces locomotor activity and L-DOPA release from nucleus accumbens.

Effects of nicotine systemically or locally on locomotor activity and L-3,4-dihydroxyphenylalanine (L-DOPA) release were studied using microdialysis in the nucleus accumbens of freely moving rats. The basal L-DOPA release was Ca2(+)-dependent and tetrodotoxin-sensitive. Systemic nicotine (1 mg/kg s.c.) increased locomotor activity and L-DOPA release preferentially in the nucleus accumbens as compared with the striatum. Injection of nicotine (30 micrograms) into the ventral tegmental area increased locomotor activity and L-DOPA release from the nucleus accumbens. These increases were antagonized by prior injection of mecamylamine into the ventral tegmental area. Nicotine induces locomotor activity and L-DOPA release from the nucleus accumbens via nicotinic receptors in the ventral tegmental area. The release may be relevant to behavioral actions of nicotine.

Animals↗
PubMed1996

Potential sites for processing of the human invariant chain by cathepsins D and E.

Seven peptides of 15-30 amino acid residues were synthesized that covered almost the entire sequence of the lumenal domain of the human invariant chain (Ii), and their hydrolysis by cathepsins D and E was investigated. Two sites were identified that were very susceptible to such cleavage. One site, the Leu174-Phe175 bond, was cleaved by both cathepsins, and the other site, the Met99-Gln100 bond, was specifically cleaved by cathepsin E. These two sites could be the sites at which native Ii is cleaved by aspartic proteinases. The cleavage of the Met99-Gln100 bond by cathepsin E might be important in the inactivation of Ii and its functional derivatives.

Amino Acid Sequence↗
PubMed1996

Stage-specific elevated expression of the genes for hepatocyte growth factor, keratinocyte growth factor, and their receptors during the morphogenesis and differentiation of rat stomach mucosa.

Hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) are two factors considered to be involved in the morphogenesis of several organs. To understand the role of HGF and KGF in the stomach development, we analyzed changes in the levels of expression of the genes for the two growth factors and their receptors in the fetal rat stomach by competitive RT-PCR. Resembling our previous results for HGF, the expression of the genes for KGF and its receptor was observed in the mesenchyme and epithelium of 16.5 day fetal stomach, respectively, indicating the possibility that KGF mediates the epithelial-mesenchymal interaction in the early stage of stomach development. The developmental profile of the expression of the genes for the two growth factors and their receptors were different, indicating a difference in their roles; the genes for HGF and c-met, the receptor for HGF, are expressed mainly during the morphogenetic period, while the genes for KGF and its receptor mainly after the morphogenetic period. Thus, it is probable that HGF controls the proliferation of epithelial cells during the morphogenetic process. The expression of the genes for KGF and its receptor is not correlated with epithelial proliferation during morphogenesis, but it does appear to be linked with epithelial differentiation. These results, together with the absence of significant mitogenic effect of KGF on the epithelial cells of the fetal rat glandular stomach in vitro, suggest a role for KGF as a differentiation factor. In addition, the expression profile of the genes for KGF and its receptor towards the end of fetal period appears to be closely correlated with that of mesenchymal cell proliferation, suggesting another role for the growth factor that is not regulated by the epithelial-mesenchymal interaction.

Animals↗
PubMed1996

A water channel closely related to rat brain aquaporin 4 is expressed in acid- and pepsinogen-secretory cells of human stomach.

We isolated a cDNA clone encoding a water channel protein, aquaporin ( AQP), from human stomach. The encoded protein consisted of 323 amino acid residues, containing six putative transmembrane domains. The protein was designated human aquaporin 4 (hAQP4) because of its 94% sequence similarity to rat brain AQP4. Expression of hAQP4 cRNA in Xenopus oocytes resulted in a significant increase in osmotic water permeability, indicating that this protein functions as a water channel. Northern blot analysis demonstrated a strong signal of hAQP4 mRNA in brain, lung, and skeletal muscle as well as in stomach. Immunohistochemical experiments with human stomach tissues showed that hAQP4 as a protein is expressed mainly in cells located in the glandular portion of the fundic mucosa. These include chief cells which secrete pepsinogen and parietal cells which secrete hydrochloric acid. These results strongly indicate that hAQP4 is a principal factor involved in the osmotic regulation of pepsinogen and acid secretion in the stomach.

Amino Acid Sequence↗
PubMed1996

Crystallization and preliminary X-ray diffraction study of aldehyde reductase from a red yeast, Sporobolomyces salmonicolor.

Crystals of aldehyde reductase from a red yeast, Sporobolomyces salmonicolor, have been grown from an ammonium sulfate solution, pH 7.0, by means of the vapor-diffusion procedure. The crystals belong to the hexagonal system, space group P6(1)22 or its enantiomorph, P6(5)22, with unit-cell dimensions of a = 72.2 and c = 320 A. The X-ray diffraction patterns extend to at least 2 A resolution with the use of synchrotron radiation. The crystals are stable on exposure to X-rays and suitable for high-resolution X-ray structure determination.

Journal Article↗
PubMed1996

Complementary DNA cloning and sequencing of rat enteropeptidase and tissue distribution of its mRNA.

A cDNA clone encoding enteropeptidase (EC 3.4.21.9), a key enzyme for the conversion of trypsinogen to trypsin, was isolated from a rat duodenal mucosa cDNA library. Sequences of the 3585 base pair clone predicted that enteropeptidase is synthesized as a single-chain precursor form, proenteropeptidase, consisting of 1058 amino acid residues with an internal signal sequence (51 residues) and is then processed into the mature enzyme consisting of three different peptide chains, i.e., mini, light and heavy chains, not the previously reported two-chain enzyme. The structure of enteropeptidase is relatively conserved among different species and the rat enteropeptidase is 24 and 39 amino acids longer than the porcine and human ones, respectively. Northern blot analysis of rNAs from normal rat tissues revealed that the enteropeptidase mRNA of around 4.4 kb in size was expressed only in the duodenal mucosa, and high proteolytic activity of the enzyme was detected in the proximal small intestine. Additional analysis of the RNAs by RT-PCR revealed that a low level of the mRNA was also expressed in the other parts of the small intestine, i.e., jejunum and ileum. These results indicate that the biosynthesis of enteropeptidase takes place mainly in the proximal small intestine, the duodenum, and the importance of the region in the physiology of intestinal protein digestion regulated by the enzyme is suggested. Furthermore a faint signal of the mRNA was also detected in the stomach, colon and brain in which the existence of trypsin-like serine proteases were reported. The significance of the low level expression of the gene is unclear, but the potential peptide-processing function of the enzyme in these tissues is also suggested.

Amino Acid Sequence↗
PubMed1996

Measurements of oxidoreductase-like activity of intact bacterial cells by an amperometric method using a membrane-coated electrode.

The oxidation of D-glucose and nicotinic acid by intact cells of Gluconobacter industrious and Pseudomonas fluorescens, respectively, is successfully measured by an amperometric method using such compounds as Fe(CN)6(3-), p-benzoquinone, and dichlorophenolindophenol as electron acceptors. Analysis of the experimental results reveals that the intact cells behave like oxidoreductases whose kinetics follows a Michaelis-Mententype equation. The catalytic behavior is explained by a model which treats the bacterial cells as bags of enzymes and assumes distribution equilibrium in the concentrations of both the substrate and the electron acceptor between the test solution and the medium within the cells. The catalytic activity can be characterized by three quantities: the maximum reaction rate (vB) and the ratios of the Michaelis constant to the distribution constant for the substrate (Ks,cell/Ks,p) and to that for the electron acceptor (KM,cell/KM,p). Advanced modification of the model to involve the membrane permeability reveals that the three quantities are effective for explaining the catalytic behavior even when the permeability effect is significant. Thus, the three quantities should be regarded as the parameters which can reflect the permeability effect.

Electrochemistry↗
PubMed1996

Structure of flavoprotein FP390 from a luminescent bacterium Photobacterium phosphoreum refined at 2.7 A resolution.

The three-dimensional structure of a flavoprotein, FP(390), from a luminescent bacterium, Photobacterium phosphoreum, solved by the molecular-replacement method, was refined to an R factor of 24.0% for 17 433 independent reflections, from 6.0 to 2.7 A resolution, collected by synchrotron radiation. The asymmetric unit of the crystal (space group P4(3)22, a = b = 76.8 and c = 242 A) contains two monomer molecules related by a non-crystallographic twofold axis to form a dimer. There are two Q-flavin [flavin mononucleotide (FMN) with myristic acid] molecules in FP(390) monomer. One of them is located at the interface of dimer which is bound to both monomer and the another is at the molecular surface. The electron density of myristic acids of Q-flavins at the dimer interface in both monomer are weak and unclear, showing the possibility that the Q-flavins bound in this site are not a single species but a mixture of two components, 6-(3"-myristic acid)-FMN and 6-(4"- myristic acid)-FMN.

Journal Article↗
PubMed1996

Electrostatic properties deduced from refined structures of NADH-cytochrome b5 reductase and the other flavin-dependent reductases: pyridine nucleotide-binding and interaction with an electron-transfer partner.

Electrostatic properties on the protein surface were examined on the basis of the crystal structure of NADH-cytochrome b5 reductase refined to a crystallographic R factor of 0.223 at 2.1 A resolution and of the other three flavin-dependent reductases. A structural comparison of NADH-cytochrome b5 reductase with the other flavin-dependent reductases, ferredoxin-NADP+ reductase, phthalate dioxygenase reductase, and nitrate reductase, showed that the alpha/beta structure is the common motif for binding pyridine nucleotide. Although the amino acid residues associated with pyridine nucleotide-binding are not conserved, the electrostatic properties and the location of the pyridine nucleotide-binding pockets of NADH-requiring reductases were similar to each other. The electrostatic potential of the surface near the flavin-protruding side (dimethylbenzene end of the flavin ring) of NADH-cytochrome b5 reductase was positive over a wide area while that of the surface near the heme-binding site of cytochrome b5 was negative. This implied that the flavin-protruding side of NADH-cytochrome b5 reductase is suitable for interacting with its electron-transfer partner, cytochrome b5. This positive potential area is conserved among four flavin-dependent reductases. A comparison of the electron-transfer partners of four flavin-dependent reductases showed that there are significant differences in the distribution of electrostatic potential between inter-molecular and inter-domain electron-transfer reactions.

Binding Sites↗
PubMed1996

Immunohistochemically demonstrated variation in expression of cathepsin E between uracil-induced papillomatosis and N-butyl-N-(4-hydroxybutyl)nitrosamine-induced preneoplastic and neoplastic changes in rat urinary bladder.

Expression of rat urinary bladder cathepsin E in benign papillomatosis induced by uracil and various stages of N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN)-induced carcinogenesis was investigated immunohistochemically. Seven-week-old, male F344/DuCrj rats were used. In the normal urothelium of control rats, cathepsin E stained in all layers of cells, although in umbrella cells and some basal cells the reaction was relatively weak. In rats given a diet containing 3% uracil for 5 weeks immunoreactivity of cathepsin E in uracil-induced papillomatosis was consistently homogeneous in all layers, but weaker than in normal urothelium. In rats given 0.05% BBN in drinking water for 12 weeks and subsequently maintained without treatment for 48 weeks cells with little cathepsin E, never observed in normal urothelium, appeared at 5 weeks above the basement membrane in the earliest stage of BBN-induced urinary bladder cancer (simple hyperplasia). Throughout the neoplastic process, groups of cells with a little cathepsin E were randomly distributed, with expression in the urothelium being markedly unstable. Almost all areas of squamous cell proliferation in TCC were negative for cathepsin E. Instability of cathepsin E expression in rat urothelium therefore appears characteristic for carcinogenesis and offers the possibility of using this feature as an early biomarker for urinary bladder carcinogenesis.

Animals↗
PubMed1996

Enterobacter kobei sp. nov., a new species of the family Enterobacteriaceae resembling Enterobacter cloacae.

The name Enterobacter kobei sp. nov. is proposed for a group of organisms referred to as NIH Group 21 at the National Institute of Health, Tokyo. The members of this species are Gram-negative, motile rods conforming to the definition of the family Enterobacteriaceae. The DNA relatedness of 23 strains of NIH Group 21 to the representative proposed as the type strain of this species averaged 82% at 70 degrees C, whereas the relatedness to other species within the family Enterobacteriaceae was less than 42%. Because the phenotypic resemblance to Enterobacter cloacae is very close and the DNA relatedness (12-42%) is closer to species of the genus Enterobacter than to other species of the family, the members of NIH Group 21 were placed in the genus Enterobacter. Close phenotypic and genetic relationships were also found between NIH Group 21 and a member of a group of organisms referred to as Enteric Group 69 at the Centers for Disease Control and Prevention (CDC), Atlanta, Georgia, USA. It is suggested that the latter could be regarded as a subspecific rank of E. kobei, though this is subject to study of further strains. The majority of strains of E. kobei were isolated from clinical specimens. A culture of the type strain (NIH 1485-79) has been deposited in the Japan Collection of Microorganisms as JCM 8580.

Bacteriological Techniques↗