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T Uchida

Publications and source records attributed to T Uchida.

At least 19 recordsLinked to original sources
PubMed1992

Phosphorylation sites on tau by tau protein kinase I, a bovine derived kinase generating an epitope of paired helical filaments.

Tau protein kinase I (TPKI) isolated from bovine brain has been determined to phosphorylate tau at four distinct sites by detecting modified Ser and Thr residues with protein sequencer. Ser199, Thr231, Ser396 and Ser413 were all found to have been phosphorylated by TPKI (numbering of amino acids was done in relation to the longest human tau [Neuron, 3 (1989) 519-526]). These phosphorylations generate an epitope of PHF (paired helical filaments) and eliminate the recognition of tau by the monoclonal antibody, tau-1. These results suggested that TPKI might be responsible for at least some of the phosphorylation of tau to induce PHF formation.

Amino Acid Sequence↗
PubMed1992

Single crystal structure of a mixed-chain triacylglycerol: 1,2-dipalmitoyl-3-acetyl-sn-glycerol.

The mixed chain triacylglycerol 1,2-dipalmitoyl-3-acetyl-sn-glycerol was synthesized and its crystal structure was determined to a final reliability factor (R) of 0.11. Two molecules are present in the monoclinic unit cell: space group P2(1); a = 5.375(1), b = 8.286(2), c = 42.96(1) A; beta = 93.30(2) degrees, V = 1910 A3, rho = 1.065 g/cm3, and mu = 5.7 cm-1. The structure is a trilayer: a bilayer of palmitate chains packed in the beta mode (T parallel) and an interdigitated monolayer of acetates. The glycerol backbone and acetate extend roughly linearly from the sn-1 chain. The sn-2 chain bends around the C-2 carbon to lie next to the sn-1 chain. Analysis of the torsion angles indicate that the glycerol conformation of 1,2-dipalmitoyl-3-acetyl-sn-glycerol is markedly different from single acid triacylglycerols and from 1,2-diacyl-sn-glycerols but very similar to 1,2-dimyristoyl-sn-glycero-3-phosphocholine.

Crystallography↗
PubMed1992

Expression of two types of neurofibromatosis type 1 gene transcripts in gastric cancers and comparison of GAP activities.

To understand the molecular mechanism of gastric tumorigenesis, the status of neurofibromatosis type 1 (NF1) gene was analyzed in human gastric cancer cell lines. Although the sequencing of the GTPase activating protein (GAP)-related region of NF1 (NF1-GRD) revealed no apparent mutation, the NF1-GRD transcript (type I) and that containing an additional 63 bp insert in the center of NF1-GRD (type II) were equally expressed in most gastric cancer cells. By contrast, type II was predominantly expressed in normal stomach mucosa. When these two types of NF1-GRD were bacterially expressed and their GAP activities were tested, both types of NF1-GRD similarly stimulated ras GTPase activity. However, arachidonic acid inhibited GAP activities of two types of NF1-GRD to different extents. These results suggest that the increased expression of type I NF1 protein may modulate ras-related signal transduction and it may be related to the control of the gastric cellular proliferation.

Arachidonic Acid↗
PubMed1992

p53 gene mutations in human gastric cancer: wild-type p53 but not mutant p53 suppresses growth of human gastric cancer cells.

To further investigate the role of p53 gene inactivation in gastric tumorigenesis, the mutational status of the p53 gene in primary human gastric cancer samples was examined. Reverse transcriptase polymerase chain reaction and subsequent direct sequencing of the p53 gene from gastric cancer samples revealed frequent point mutations of the p53 gene: some of these coincided with those previously identified in gastric cancer cell lines. In addition, both allelic deletion analysis using pYNZ 22 and polymerase chain reaction-restriction fragment length polymorphism analysis demonstrated an allelic deletion of the p53 gene in cancer tissue which contained a point mutation of the p53 gene in the remaining allele. Transfection of the wild-type or mutant p53 genes into gastric cancer cells showed that the wild-type but none of the mutated p53 genes suppressed the colony formation of gastric cancer cells. Furthermore, the incorporation of thymidine into DNA was reduced in cancer cells expressing the wild-type p53 gene. The glutathione S-transferase-wild type p53 fusion protein bound to simian virus 40 large T antigen in COS-1 cell lysate. None of the p53 fusion proteins containing mutations at codons 143, 175, 248, or 273 bound to simian virus 40 large T antigen. By contrast, two different mutant p53 fusion proteins containing mutations specifically observed in gastric cancer bound to simian virus 40 large T antigen. These results indicate that inactivation of the p53 gene through mutations and the allelic deletion may play an important role in gastric tumorigenesis. These mutations may cause a conformational change in the p53 protein resulting in the loss of the suppression by p53 of the growth of gastric cells, partly through disruption of the association of p53 protein with a cellular component.

Antigens, Viral, Tumor↗
PubMed1992

Tau protein kinase I converts normal tau protein into A68-like component of paired helical filaments.

From bovine brain microtubules we purified tau protein kinase I (TPKI, Mr 45,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and tau protein kinase II (TPKII) whose activity was attributed to a 30-kDa protein on SDS-PAGE by affinity-labeling using an ATP analog. Both kinases were activated by tubulin. TPKII, but not TPKI, phosphorylated tau fragment peptides previously used for detection of a Ser/ThrPro kinase activity. Therefore, TPKII was considered to be the Ser/ThrPro kinase. TPKI was more effective than TPKII for producing the decrease of tau-1 immunoreactivity and mobility shift of tau on SDS-PAGE. Moreover, TPKI, but not TPKII nor other well-known protein kinases, generated an epitope present on paired helical filaments. These findings suggested that tau phosphorylated by TPKI resembled A-68, a component of paired helical filaments.

Affinity Labels↗
PubMed1992

Molecular cloning, characterization, and expression in Escherichia coli of a cDNA encoding mammalian choline kinase.

A rat liver lambda gt11 cDNA library was screened with antibody against rat liver choline kinase. One of the putative phage clones obtained was found to express choline kinase activity in infected Escherichia coli cells and used for the isolation of fully extended cDNA clones by hybridization. The obtained clones had identical overlapping nucleotide sequences and gave a composite cDNA sequence of 2540 bases. Poly(A)+ tails were found at three different sites of the cDNA sequence. Within the sequence there was a large open reading frame encoding 435 amino acids with a molecular mass of 49,743 Da. When this open reading frame was placed after the trc promoter and introduced into E. coli cells, fully active choline kinase was produced. This enzyme was partially purified using a published choline kinase purification method and was found to mediate the phosphorylation of choline, N,N-dimethylethanolamine,N-monomethylethanolamine, and ethanolamine. Similar to the case with choline kinases from various sources, Km decreased as the number of N-methyl groups increased. The deduced amino acid sequence significantly resembled the yeast choline kinase sequence. It also showed local sequence similarity to protein kinases and some bacterial phosphotransferases. Northern blot analysis revealed that the cDNA probe hybridized to 2.8- and 1.8-kilobase RNA in various rat tissues. The sizes of the transcripts were fairly consistent with that of cDNA. The order of mRNA abundance was testis, brain, lung, kidney, and liver, but did not coincide with the order of the activity levels in these tissues. These results show that the cloned cDNA encodes one of the choline kinase isoforms present in mammalian tissues.

Amino Acid Sequence↗
PubMed1992

High-performance affinity chromatography system for the rapid, efficient assay of glycated albumin.

A high-performance affinity chromatographic system was constructed and shown to permit highly reproducible, rapid, automatic assays of serum glycated albumin (GA) by separation of albumin (Alb) on an anion-exchange column (Asahipak ES-502N) packed with a vinyl alcohol copolymer bearing diethylamino groups and consecutive separation of GA on a column packed with a vinyl alcohol copolymer bearing boronate groups. The first column selectively retained Alb free of other serum proteins and permitted at least a 95% recovery of sample Alb. The purity of the Alb peak was confirmed by two-dimensional electrophoresis. Chromatographic analyses of human serum Alb incubated with glucose on the second column showed that the peak area for GA increased in accordance with the incubation time and suggested selective adsorption of GA on the second column. Optimization of the conditions for the two-column system reduced the analysis time to 10 min. Analyses of human sera with the present system showed GA to be 16.1 +/- 1.1% (mean +/- S.D.) of total Alb in non-diabetic children and 39.9 +/- 9.1% (mean +/- S.D.) in diabetic children (0-17 years old).

Adolescent↗
PubMed1992

Missense mutations and a deletion of the p53 gene in human gastric cancer.

To investigate the molecular pathogenesis of human gastric cancers the p53 gene, a suppressor oncogene, was analyzed in 12 human gastric cell lines. Southern blot and Northern blot analysis revealed a total deletion of p53 gene in KATO-III cells but no major abnormality of p53 gene in other cell lines. By the use of the reverse-transcriptase polymerase chain reaction and direct sequencing 7 cell lines showed point mutations of p53 gene resulting in amino-acid substitutions. Most of them were rare mutations which had not been observed in other types of cancers. One of these mutations was also detected through the use of PCR and oligomer-specific hybridization. Six out of 7 cell lines with mutations of p53 gene also lost one allele of chromosome 17p. Immunoblotting of cell lysates with an antibody specific to p53 demonstrated the absence of p53 protein in KATO-III cell. By contrast, the high levels of the p53 protein were observed in 5 cell lines all of which contained mutations of p53 gene. These results further suggest that the inactivation of p53 gene may play an important role in the transformation of gastric cells to the malignant phenotype. KATO-III cells might be a good model for studying the significance of the loss of p53 gene in cellular transformation.

Alleles↗
PubMed1992

Zinc iodide-osmium tetroxide impregnation of the "tubulo-vesicular system" in Tomes' process of the rat incisor ameloblast.

Zinc iodide-osmium tetroxide (ZIO) is a nonspecific but selective impregnation method that visualizes a tubulo-vesicular system in cells. The detailed structure and three-dimensional distribution of this ZIO-impregnated system was studied in the Tomes' process of secretory ameloblasts in the rat incisor. The ZIO-impregnated system consisted of an extensive array of smooth membrane-bound thick and thin tubules and vesicles. The interconnected thick and thin tubules formed a complex "core network" in the central cytoplasm of Tomes' process that enmeshed and often surrounded individual secretory granules. From the core network, radial branches extended toward the smooth cell membrane of the interdigitating portion of Tomes' process. Although the core network and branches frequently appeared connected to the secretory granules and the cell membrane, stereo-pair electron microscopy failed to show conclusive evidence of such continuity. However, many coated vesiclelike structures were attached to the core network and its branches. No special relationship was found between interrod and rod secretory sites and the tubulo-vesicular network. In thick sections, the ZIO-impregnated tubulo-vesicular network occupied a considerable volume of cytoplasm. The vinblastine-labile nature of this network as demonstrated previously (Nanci et al., 1987) indicated that the system undergoes rapid and extensive turnover. Considering the dynamic nature and sheer volume of the tubulo-vesicular system, we propose that it be regarded as a major cell organelle.

Ameloblasts↗
PubMed1992

Preparation and evaluation of Eudragit gels. III: Rectal gel preparations for sustained release of pentoxifylline.

A novel method was established for the preparation of Eudragit S, Eudragit L, and Eudispert high-viscosity (hv) hydrogel and xerogel preparations containing a medicinal component such as pentoxifylline (an agent that improves cerebral microcirculation). A significant correlation was found between the in vitro mean dissolution time and the in vivo mean residence time of pentoxifylline after rectal administration of these preparations in rabbits. Staying properties of the hydrogel and xerogel preparations in the lower part of the rectum were compared with those of polyethylene glycol 2000 and Witepsol S-55 suppositories in rats, with water-insoluble dye. Eudispert hv hydrogel and xerogel preparations have excellent staying properties in the rectum. The efficacy of Eudispert hv xerogel preparations may be reduced by first passage through the liver after oral administration.

Acrylic Resins↗
PubMed1992

Preparation and evaluation of Eudragit gels. IV: Rectal gel preparations for sustained release and avoidance of first-pass metabolism of propentofylline.

Formulations of a high-viscosity acrylic resin gel (Eudispert hv) containing propentofylline, a new cerebral microcirculation-improving agent, were prepared and tested for avoidance of the first-pass metabolism of propentofylline through the liver and for sustained release of propentofylline. The absolute bioavailability of propentofylline after oral administration was only 4% in rabbits. The relative bioavailabilities of propentofylline from polyethylene glycol (M(r), 2000) and Witepsol H-15 suppositories were approximately 8- and 16-fold, respectively, compared with oral administration. Furthermore, the absolute bioavailability of propentofylline from Eudispert hv hydrogel and xerogel preparations was approximately 100%. The results indicate that, in principle, drug loss caused by first-pass metabolism may be avoided completely by placing Eudispert hv hydrogel and xerogel formulations in the lower part of the rectum for long periods.

Acrylic Resins↗
PubMed1992

A case of multiple leiomyomatous lesions of the lung: an analysis of flow cytometry and hormone receptors.

A 36 year old woman was admitted to our department because of a chest X-ray which showed multiple developing shadows. She underwent bilateral exploratory thoracotomies and a total 5 tumors were resected and pathologically diagnosed as benign metastasizing leiomyoma, the largest of which was positive for the progesterone receptor and negative for the estrogen receptor. A histogram of this tumor using a flow cytometer showed a diploid pattern and 4.6 percent of the S phase which was not more than that of a leiomyoma of the uterus from another patient. Two months later, she underwent a hysterectomy and bilateral salpingo-oophorectomy for treatment of the positive progesterone receptor in the pulmonary lesions. The resected uterine myoma and normal myometrium showed positive estrogen and progesterone receptors. For the subsequent 28 months she has been free of any further symptoms. Benign metastasizing leiomyoma of the uterus is a rare disease and very interesting because of its histological benignity and hormonal dependency. However, according to the literature, it is often confused in entity due to the fact that normal lung tissue also possesses hormone receptors. Considering our data on hormone receptors, it is rational to think that multiple leiomyomatous lesions in the lung should only be diagnosed as benign metastasizing leiomyomas when they possess positive estrogen and progesterone receptors.

Adult↗
PubMed1992

The localization and characterization of proteinases for the initial cleavage of porcine amelogenin.

In the outermost layer of porcine-developing enamel adjacent to the ameloblasts in the secretory stage, the activities of two proteinases having molecular masses of 76 and 78kDa were detected by enzymography using gelatin as a substrate. On the other hand, high activities of known 30 and 34kDa proteinases were localized in the inner layer of the enamel. The 76kDa proteinase cleaved the carboxyl-terminal peptide of porcine 25kDa amelogenin to convert it to 20kDa amelogenin. The 78kDa proteinase also acted on the 25kDa amelogenin similarly, but its activity was weak. The results indicate that the 25kDa amelogenin synthesized and secreted by ameloblasts is converted to 20kDa amelogenin by the action of proteinase localized in the outermost layer of the secretory enamel, and then further degraded by the proteinases in the inner layer of the enamel associated with the increase of mineralization.

Amelogenin↗
PubMed1992

Neurotensin prevents intestinal mucosal hypoplasia in rats fed an elemental diet.

Liquid elemental diets are associated with mucosal hypoplasia of both the small intestine and colon. Neurotensin, a tridecapeptide widely distributed in the gut, is trophic for the small intestine of rats fed a normal chow diet. The purpose of this study was to determine whether neurotensin could reverse the hypoplasia of intestinal mucosa that is associated with feeding a liquid elemental diet. Forty male Sprague-Dawley rats were randomized into five groups. Four groups were fed (for seven days) a glutamine-free liquid elemental diet. Subcutaneous injection of saline (control) or neurotensin (33, 100 or 300 micrograms/kg) were given to the groups of rats every 8 hr for seven days. Group five (Chow) received rat chow ad libitum for seven days. Rats were killed on day 8, and the proximal jejunum, distal ileum, and proximal colon removed. Mucosal weight, DNA, RNA, and protein contents were determined. Neurotensin (300 micrograms/kg) increased the cellularity of the small intestinal mucosa and reversed mucosal hypoplasia due to an elemental diet; a more pronounced effect was noted in the jejunum compared to the ileum. Neurotensin (33 and 100 micrograms/kg) increased mucosal DNA content in the jejunum but was not effective in reversing the hypoplasia. Neurotensin had no effect on growth of colonic mucosa. These results suggest that neurotensin may be an important trophic hormone for the small intestine. Administration of neurotensin may alleviate hypoplasia of the small bowel mucosa and maintain functional integrity of the gut during prolonged feeding of an elemental diet.

Animals↗
PubMed1992

Degradation of methyl and ethyl mercury into inorganic mercury by various phagocytic cells.

In connection with the dealkylation of methyl mercury (MeHg) and ethyl Hg (EtHg) with reactive oxygen-producing systems, we examined the ability of phagocytic cells to degrade MeHg or EtHg into inorganic mercury in vitro by collecting them from blood or peritoneal cavity of several species of animal. EtHg was readily degraded by human polymorphonuclear leukocytes (PMN), rat PMN, guinea-pig PMN, rabbit PMN, guinea-pig macrophages (M phi), human monocytes and guinea-pig eosinophils. In contrast, rat hepatocytes and the M phi hybridoma clone 39 cells were weaker in their degrading ability. Degradation of MeHg by these cells was always much weaker than EtHg, under identical conditions; however, by increasing the cell numbers, MeHg was appreciably degraded by human PMN, rat PMN and rabbit PMN. The reactive oxygen species mainly responsible for alkyl Hg degradation seemed to be hydroxyl radicals produced by M phi, and hypochlorous acid produced by PMN, monocytes and eosinophils. It was also suggested that the degradation of alkyl Hg by these cells might be an intraphagosomal event.

Animals↗
PubMed1992

Hepatitis E: review.

Hepatitis E is endemic, often provoking epidemics in many developing countries. It resembles hepatitis A clinically and epidemiologically but show a higher mortality rate and less infectiousness. Several lines of evidence strongly support the assumption that humans become immunized once they contract hepatitis E. Because of the low infectiousness, most of the adult population of endemic areas are susceptible to hepatitis E until an epidemic occurs, although they are almost always infected with hepatitis A during infancy. Epidemics are caused by accidental contamination by the hepatitis E virus (HEV) in feces of water provided to these people. The liver change reveals necroinflammation related to the immune-mediated mechanism. The HEV is molecularly cloned and sequenced and has a single-stranded, positive-sense RNA genome, 7,194 nucleotides followed by a poly (A) tail. There are three open reading frames. The non-structural gene, approximately 5 kb is located at the 5' end, while the structural gene, approximately 2 kb is located at the 3' end of the genome. There is a low level of nucleotide variations among HEV strains isolated from Myanmar and China and a single serotype appears to exist. The HEV may be a new RNA virus or belong to Caliciviridae family. Further investigation include in vitro propagation, elucidation of the gene replication, global seroepidemiology and vaccination of the HEV.

Animals↗
PubMed1992

Escherichia coli RecA protein modified with a nuclear location signal binds to chromosomes in living mammalian cells.

We tried to make a well-characterized bacterial protein function in mammalian cell nuclei. For this purpose we chose Escherichia coli RecA protein and fused its carboxy terminus to the nuclear location signal of SV40 large T-antigen by oligonucleotide-dependent modification of the gene. When injected into the cytoplasm, the modified RecA protein (T-RecA for the T-antigen signal) accumulated efficiently in the nuclei, whereas the wild-type RecA protein remained in the cytoplasm. The T-RecA protein retained its original in vivo activity, judging from the finding that uv-sensitive bacteria (recA- E. coli) became uv-resistant on transformation with the T-recA plasmid as well as the recA plasmid. For expression of the T-recA gene in mammalian cells, the 5' region was replaced by the chicken beta-actin promoter and Kozak's initiation signal. A high level of expression was observed when Chinese hamster ovary (CHO-K1) cells were transfected with this plasmid. Indirect immunofluorescence examination revealed that the T-RecA protein in nuclei of mammalian cells bound to chromatin.

Amino Acid Sequence↗