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M Nomoto

Publications and source records attributed to M Nomoto.

At least 37 records · Page 2Linked to original sources
PubMed2000

Expression of UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferase isozymes T1 and T2 in human colorectal cancer.

Uridine diphosphate (UDP)-GalNAc: polypeptide N-acetylgalactosaminyltransferase (GalNAc transferase) catalyzes the initial step in mucin type O-glycosylation, and its expression has been assumed to be altered between normal epithelial cells and cancer cells. We studied the alteration of GalNAc transferase expression during the carcinogenesis of human colorectal epithelial cells. We produced polyclonal antibodies against synthetic polypeptides with specific sequence to two GalNAc transferase isozymes, T1 and T2. Surgically resected specimens from 50 patients with colorectal cancer were immunohistochemically stained, and the staining grade (percentage of positively stained cells) was compared between cancer and its normal counterpart in the same specimen. Significant signals for both T1 and T2 expression were seen in the supranuclear region of normal and cancer cells, indicating the subcellular localization of the enzymes in the Golgi apparatus. The prevalence of positive staining for T1 and T2 expression in colorectal cancer was significantly higher than that in normal epithelium (P < 0.05). However, the difference in staining grades between cancer and normal tissues varied in each patient. These results indicate that there is variability in the expression patterns of GalNAc transferase isozymes in normal and cancerous cells colorectal among individuals.

Adenocarcinoma↗
PubMed2000

Improvement of intestinal absorption of peptides: adsorption of B1-Phe monoglucosylated insulin to rat intestinal brush-border membrane vesicles.

In a previous study we glycosylated insulin to improve its intestinal absorption. When the glycosylated product, p-(succinylamido)-phenyl-alpha-D-glucopyranoside (SAPG)-substituted insulin (SAPG-INS), was administered intra-intestinally to rats, it showed a greater hypoglycemic effect than native bovine insulin. The enhanced hypoglycemic effect of SAPG-INS was considered to be due to an increase in membrane permeability as well as an increase in resistance to enzymatic degradation. In particular, membrane permeability may be related to an interaction with the Na(+)-dependent D-glucose transporter (SGLT-1) which is located in the brush-border membrane of epithelial cells. The insulin product used in the previous study, however, comprised a mixture of mono-, di- and tri-SAPG-substituted insulin. In this study SAPG-INS with a defined substitution number and position was synthesized to examine the interaction between the transporter and glycosylated insulin in more detail. The new product was mono-SAPG-substituted insulin substituted at the B1-phenylalanine position (B1-SAPG-INS) and was selectively synthesized after protection of the A1-glycine and varepsilonB29-lysine amino acids. The hypoglycemic effect of B1-SAPG-INS in rats after an intravenous dose of 71 microg/kg was almost the same as that of native bovine insulin at a dose of 1 U/kg and B1-SAPG-INS retained about 60% of the immunoreactivity of native bovine insulin. The interaction of B1-SAPG-INS with the intestinal transporter was examined by a rapid filtration technique using (125)I-labeled B1-SAPG-INS and brush-border membrane vesicles (BBMVs) which were prepared from rat small intestine by the Mg-precipitation method. The amount of B1-SAPG-INS adsorbed or absorbed by BBMVs in the presence of an inward Na(+)-gradient into BBMVs was greater than that of native bovine insulin. This adsorption/absorption was significantly inhibited by the presence of 1 mM phloridzin. A similar inhibition was observed when Na(+) was replaced with K(+) and when B1-SAPG-INS was incubated with BBMVs at 4 degrees C. From the effect of osmolarity on the extent of adsorption/absorption, it was considered that B1-SAPG-INS was not taken up into the intravesicular space but adsorbed onto the external membrane surface of BBMVs. These findings suggested that B1-SAPG-INS was adsorbed specifically onto the transporter. The hypoglycemic effect of insulin was enhanced by glycosylation at the B1 position in in situ experiments using normal and diabetic rats. Consequently, it is suggested that B1-SAPG-INS was adsorbed specifically onto the glucose transporter of intestinal BBM. This specific adsorption may be involved in the mechanism of the enhanced hypoglycemic effect of B1-SAPG-INS both in normal and diabetic rats.

Adsorption↗
PubMed2000

Effect of shitei-to, a traditional Chinese medicine formulation, on pentylenetetrazol-induced kindling in mice.

This study measured the effects of Shitei-To (STT), a traditional Chinese Medicine, which is a mixture of extracts from three medicinal herbs, Shitei (SI, Kaki Calyx; calyx of Diospyros kaki L. f.), Shokyo (SK, Zingiberis Rhizoma; rhizome of Zingiber officinale Roscoe) and Choji (CJ, Caryophylli flos; flowerbud of Syzygium aromaticum [L.] Merrill et. Perry), has long been used for the treatment of hiccups in Japan and China, against fully pentylenetetrazol-kindled seizures and on the development of pentylenetetrazol kindling in mice. Repeated administration of STT (3.0 g/kg p.o.) mildly retards the development of pentylenetetrazol-induced kindling in mice. STT also decreased the number of tonic-clonic convulsions resulting from progression kindling. On the other hand, STT had no effect on convulsions in fully pentylenetetrazol-kindled mice. These findings suggest that STT protects against the development of convulsions, and that STT may have therapeutic effects in the prevention of secondarily generalized seizures.

Animals↗
PubMed2000

A novel PTEN mutation in a Japanese patient with Cowden disease.

Cowden disease (CD) is an autosomal dominant syndrome characterized by multiple hamartomatous lesions and an increased risk for malignancies. Recent evidence has indicated that the PTEN gene, encoding a protein tyrosine phosphatase, is the CD susceptibility gene. However, another line of evidence has suggested that CD might be genetically heterogeneous. Clinical features of CD are variable, and there are interfamilial differences in the expression of skin lesions. Therefore, information on PTEN mutations in CD patients should be accumulated to clarify the genotype-phenotype correlation. In the present study, we found heterozygous germline mutations of PTEN in all of three Japanese patients with CD examined, indicating no genetic heterogeneity among our patients. The mutations included two non-sense mutations of R335X and R130X, and a mis-sense mutation of C136R. To the best of our knowledge, the C136R mutation has not previously been reported in CD patients. This novel mutation was located outside the core motif of the phosphatase domain of PTEN protein, where most of the missense mutations previously reported in CD patients were clustered. Mucocutaneous manifestations were far fewer in the patient with this mutation than in the patients with nonsense mutations. Whether the phenotypic difference in mucocutaneous features was due to the different mutations remains unclear.

Adult↗
PubMed2000

Quantitative evaluation of genomic instability as a possible predictor for development of hepatocellular carcinoma: comparison of loss of heterozygosity and replication error.

Both loss of heterozygosity (LOH) and replication error (RER) are considered to be phenotypes of genomic instability. To unveil the role of the genomic instability in hepatocarcinogenesis, frequencies of LOH and RER were simultaneously determined in 15 hepatocellular carcinomas (HCCs), surrounding nontumorous liver tissues (SL), and 13 liver tissues with chronic viral hepatitis void of cancer (NC) by referencing peripheral blood leukocytes (PBLs) from the corresponding donor using 18 microsatellite markers spread throughout the genome. LOH was significantly frequent in HCC compared with that in SL or NC (P =.005, P =.0003, respectively) and observed preferentially at particular microsatellite loci, D1S204, D2S123, D8S1106, D9S266, D16S748, and D19S601. Although the higher prevalence of RER was also significant in HCC compared with that in NC (P =.03), in most cases the errors were detected at very low frequencies and random loci. Both LOH and RER tended to appear more prevalently in SL than in NC. The occurrence rate of LOH was higher in the tissues associated with hepatitis B virus (HBV) than with hepatitis C virus (HCV) infection especially in HCC (P =.03). When referencing SL instead of PBLs, the prevalence of LOH and RER in HCC significantly decreased (P =.02 and P =.03, respectively). These results suggest that LOH is closely associated with multistep hepatocarcinogenesis especially under HBV infection, but RER is imperceptibly associated. The quantitative evaluation of the frequency of LOH by referencing PBLs may be a useful predictor for HCC development in chronic liver diseases.

Blood Cells↗
PubMed2000

Alcoholic fatty liver differentially induces a neutrophil-chemokine and hepatic necrosis after ischemia-reperfusion in rat.

Primary graft nonfunction of steatotic liver allograft is one of the factors causing shortage of donor livers. Ischemia/reperfusion (I/R) injury is an important contributory factor to primary graft nonfunction. In this study, we investigated the complex chain of events from transcription factor activation to necrosis through cytokine induction and apoptosis in steatotic rat liver after warm I/R. Rats with alcoholic or nonalcoholic fatty liver were subjected to hepatic warm I/R and compared with control rats. Rats fed an ethanol diet for 6 to 8 weeks developed severe hepatic necrosis accompanied by increased neutrophil recruitment after I/R, compared with rats with nonalcoholic fatty liver or control. Hepatic apoptosis as assessed by DNA fragmentation at 4 hours after I/R, however, increased to a similar degree in each of the 2 fatty liver models compared with the control. Alcoholic fatty liver exposed to I/R showed a rapid increase in nuclear factor-kappaB (NF-kappaB) binding activity at 1 hour after I/R, which preceded an increased expression of tumor necrosis factor alpha (TNF-alpha) and cytokine-induced neutrophil chemoattractant-1 (CINC-1). In contrast, nonalcoholic fatty liver did not show such potentiation of either NF-kappaB activation or cytokine induction after I/R. Our results have indicated that alcoholic fatty liver may differentially induce CINC-1 production and hepatic necrosis after I/R. Furthermore, our results suggest that apoptosis per se does not always lead to necrosis in the liver following I/R.

Animals↗
PubMed2000

[Functions and metabolic rates of transmitters in the basal ganglia].

The caudate nucleus and putamen showed different neuronal circuits with the cerebral cortex and different functions in motor control. The dopamine content was same in the caudate nucleus and putamen, however, the turnover ratio was higher in the putamen than in the caudate nucleus, when it was studied in the control or MPTP-treated marmosets. The dopaminergic neurons projecting to the putamen seemed to show higher activity than those projecting to the caudate nucleus. The caudate nucleus is long in the shape and the dopamine turnover ratio was higher in the former part than in the latter part in normal marmosets. Administration of fluvoxamine increased akinesia in MPTP-treated common marmosets, and systemic administration of 8-OHDPAT, a 5-HT1A receptor or autoreceptor agonist reversed parkinsonism in MPTP-treated common marmosets. These results indicated that serotonergic neurons potentiated hypoactivity in parkinsonism and suppression of serotonergic function may contribute to the remission of parkinsonism.

Acetylcholine↗
PubMed1999

Structural basis for the regulation of UDP-N-acetyl-alpha-D-galactosamine: polypeptide N-acetylgalactosaminyl transferase-3 gene expression in adenocarcinoma cells.

The UDP-N-acetyl-alpha-D-galactosamine: polypeptide N-acetylgalactosaminyl transferase-3 (Gal NAc-T3) gene, a member of the Gal NAc transferase gene family, is expressed in a tissue-specific manner. To elucidate the function of this gene, we have focused on the molecular mechanism underlying regulation of gene expression. We have cloned Gal NAc-T3 cDNA and used it to show that Gal NAc-T3 mRNA is expressed in tumor cell lines derived from secretory epithelial tissue adenocarcinomas but not in cell lines derived from bladder and epidermoid carcinomas. Using a polyclonal antibody to Gal NAc-T3, we observed protein expression in adenocarcinoma but not non-adenocarcinoma cell lines, and in breast carcinoma cells but not in normal breast tissue. We used Gal NAc-T3 cDNA to isolate three overlapping genomic clones containing the 5'-portion of the human Gal NAc-T3 gene, and we sequenced 1.6 kb around the first exon. A transient expression assay using the luciferase gene showed that promoter activity was much higher in MCF-7 cells than in KB cells. In vivo footprint experiments showed significant protection of a distal GC box, an NRF-1 site, and an AP-2 site in MCF-7 cells. A novel stem and loop structure extending from nucleotide -103 to nucleotide -165 and contiguous to these transcription factor binding sites seemed to be functional in regulating Gal NAc-T3 gene transcription, and a KMnO4 footprint experiment showed that this stem and loop structure could be formed in vivo. We also observed dimethyl sulfate hypersensitive sites in the untranslated region around nucleotide +50 in MCF-7 but not in KB cells. These findings indicate that Gal NAc-T3 gene expression is regulated by multiple systems, including transcription factor binding sites and a stem-and-loop structure, and that this regulation is restricted to cell lines derived from epithelial gland adenocarcinomas but not cells derived from nonsecretory epithelial tissue carcinomas. In addition, our immunohistochemical results suggest that our anti-Gal NAc-T3 antibody may be useful for diagnostic purposes in the early stages of breast cancer.

Adenocarcinoma↗
PubMed1999

Sensitization by prolonged glutathione depletion of kainic acid to potentiate DNA binding of the nuclear transcription factor activator protein-1 in murine hippocampus.

In four of four mice intracerebroventricularly injected with the inhibitor of glutathione synthesis L-buthionine-[S,R]-sulfoximine (BSO) 2 days before, an intraperitoneal injection of kainic acid (KA) invariably led to marked potentiation of DNA binding activity of the nuclear transcription factor activator protein-I (AP1) in the hippocampus at a dose which was ineffective in animals previously injected with vehicle alone. However, KA failed to potentiate binding in animals injected with BSO 1 day before. The intracerebroventricular injection of BSO induced marked and prolonged depletion of a total glutathione content in murine hippocampus for 1-2 days after administration. These results suggest that prolonged depletion of endogenous glutathione for a period longer than 1 day may lead to sensitization of KA signals to potentiate AP1 DNA binding in cell nuclei and thereby modulate de novo synthesis of particular proteins at the level of gene transcription in murine hippocampus.

Animals↗
PubMed1999

Cellular balance of glutathione levels through the expression of gamma-glutamylcysteine synthetase and glutathione thiol transferase genes in human hepatic cells resistant to a glutathione poison.

Buthionine sulfoximine (BSO) is a synthetic amino acid that irreversibly inhibits glutathione biosynthesis and deranges reduced glutathione (GSH) metabolism in liver cells. We isolated two BSO-resistant lines, HLE/BSO2-1 and HLE/BSO2-2, from human hepatic HLE/WT cells. Cellular levels of the Pi class glutathione thiol transferase (GSTP1) were 3-fold lower in BSO-resistant lines than in HLE/WT cells. By contrast, gamma-glutamylcysteine synthetase (GCS) heavy subunit (GCSh) mRNA levels were markedly decreased in HLE/BSO2-1 and HLE/BSO2-2 as compared with HLE/WT. The expression of a dominant-negative mutant of c-Jun inhibited the GCSh promoter activity in HLE/WT, but not in HLE/BSO2-1. Cellular levels of AP-1, however, were not decreased in either BSO-resistant cell line. Transfection of GCSh promoter of various lengths driven reporter constructs showed no sequence-specific increase in the promoter activities in HLE/BSO2-1. However, transfection of GSTP1 cDNA into HLE/BSO2-1 and HLE/BSO2-2 restored the levels of GCSh mRNA and the GCSh promoter activity to those of HLE/WT. Sequences between -315 and -241 bp of the 5' region contained an AP-1 site responsible for the enhanced GCSh promoter activity in GSTP1 transfectants of HLE/BSO2-1. In vivo footprint analysis showed a specific protection of the AP-1 site on GCSh promoter in GSTP1 transfected HLE/BSO2-1. GSH homeostasis thus appears to be maintained by an interaction between GSTP1 and GCS in human hepatic cells resistant to the GSH poison.

Buthionine Sulfoximine↗
PubMed1999

Control of O-glycan branch formation. Molecular cloning of human cDNA encoding a novel beta1,6-N-acetylglucosaminyltransferase forming core 2 and core 4.

A novel human UDP-GlcNAc:Gal/GlcNAcbeta1-3GalNAcalpha beta1, 6GlcNAc-transferase, designated C2/4GnT, was identified by BLAST analysis of expressed sequence tags. The sequence of C2/4GnT encoded a putative type II transmembrane protein with significant sequence similarity to human C2GnT and IGnT. Expression of the secreted form of C2/4GnT in insect cells showed that the gene product had UDP-N-acetyl-alpha-D-glucosamine:acceptor beta1, 6-N-acetylglucosaminyltransferase (beta1,6GlcNAc-transferase) activity. Analysis of substrate specificity revealed that the enzyme catalyzed O-glycan branch formation of the core 2 and core 4 type. NMR analyses of the product formed with core 3-para-nitrophenyl confirmed the product core 4-para-nitrophenyl. The coding region of C2/4GnT was contained in a single exon and located to chromosome 15q21.3. Northern analysis revealed a restricted expression pattern of C2/4GnT mainly in colon, kidney, pancreas, and small intestine. No expression of C2/4GnT was detected in brain, heart, liver, ovary, placenta, spleen, thymus, and peripheral blood leukocytes. The expression of core 2 O-glycans has been correlated with cell differentiation processes and cancer. The results confirm the predicted existence of a beta1,6GlcNAc-transferase that functions in both core 2 and core 4 O-glycan branch formation. The redundancy in beta1,6GlcNAc-transferases capable of forming core 2 O-glycans is important for understanding the mechanisms leading to specific changes in core 2 branching during cell development and malignant transformation.

Amino Acid Sequence↗
PubMed1999

Transcription factor Y-box binding protein 1 binds preferentially to cisplatin-modified DNA and interacts with proliferating cell nuclear antigen.

The Y-box binding protein (YB-1) binds to inverted CCAAT box sequences that are present in the promoter region of many genes. We previously showed that YB-1 is overexpressed in human cancer cell lines that are resistant to cisplatin and that the depletion of YB-1 by transfection of a vector expressing YB-1 antisense RNA increases the sensitivity of human cancer cells to cisplatin. To determine whether YB-1 can bind to cisplatin-modified DNA, we fused YB-1 cDNA to glutathione S-transferase (GST) cDNA and purified the resulting GST fusion protein. When we tested the fusion protein with unmodified or cisplatin-modified oligonucleotides, we found that GST-YB-1 bound more strongly to cisplatin-modified oligonucleotides, as did GST fusion proteins of high mobility group 1 (HMG1), HMG2, and xeroderma pigmentosum group A protein. When we assayed the ability of proliferating cell nuclear antigen (PCNA) to interact with the GST fusion proteins, we observed binding to YB-1 but not to HMG1, HMG2, or xeroderma pigmentosum group A. Subsequent experiments demonstrated that YB-1 and PCNA interact directly via the COOH-terminal region of YB-1. Using immunochemical coprecipitation methods, we observed binding of YB-1 and PCNA in vivo. These results suggest that YB-1 can function as a recognition protein for cisplatin-damaged DNA and that it may be important in DNA repair or in directing the cellular response to DNA damage.

CCAAT-Enhancer-Binding Proteins↗
PubMed1999

Effect of selegiline on dopamine concentration in the striatum of a primate.

Monoamine oxidase (MAO) has two subtypes, A and B, that have different distributions between the rodent and the human. In the striatum, dopamine (DA) of the rat seems to be metabolized by MAO A, and DA of the human is largely deaminated by MAO B. MAO in the striatum of common marmosets is also type B. Using in vivo microdialysis, we investigated the pharmacological activity of selegiline, a selective irreversible inhibitor of MAO B, in the striatum of marmosets. Intraperitoneal co-administration of selegiline (1 mg kg-1, i.p.) with levodopa/carbidopa (10/2.5 mg kg-1, i.p.) did not significantly increase extracellular concentration of DA in the striatum of common marmosets compared with control animals receiving levodopa/carbidopa alone. Daily pretreatment with 0.1 mg kg-1 (i.p.) selegiline for two weeks, however, dramatically increased extracellular concentration of DA to about seven times that of control animals treated with levodopa/carbidopa alone in marmosets. Such an increase in extracellular concentrations of DA could not be observed in a similar study with Wistar rats. This study showed that chronic administration of a small dose of selegiline caused a marked increase in extracellular DA concentration in the striatum of primates, but not in the rodents.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1999

Reduction of telomeric repeats as a possible predictor for development of hepatocellular carcinoma: convenient evaluation by slot-blot analysis.

Hepatocellular carcinoma (HCC) mainly arises from the liver with chronic inflammation. Because telomere reduction reflects replicative history in somatic cells, we analyzed the possibility that liver tissues surrounding HCC consist of the cells carrying substantial reduction of telomere. We studied 20 HCC and surrounding noncancerous liver tissues (SL) obtained by surgical resection, and 10 laparoscopically obtained needle biopsy specimens of the liver with chronic inflammation including no overt HCC (CI). Five liver tissues without chronic liver diseases (ND) were also examined. Extracted genomic DNAs were blotted on a nylon membrane, and probed at first with radio-labeled d(TTAGGG)(3) and reprobed with radio-labeled d(CCT)(7). The intensity caused by d(TTAGGG)(3) was divided by that of d(CCT)(7). The ratio was defined as telomeric repeats content (TC). Dilution experiments reproducibly revealed almost the same TC. The reduction rate of telomere length through aging estimated by regression analysis of TC was 0.62% per year. Concomitant analyses of TC and average telomere length revealed that both values were significantly correlated (r =.45; P =.009). To compare TC in the liver with respect to chronic inflammation, the value was divided by TC in peripheral blood leukocytes (PBL) from the same donor. The ratio was defined as relative TC (RTC). There was a statistically significant decrease of RTC in CI compared with that in ND (P =.03). Furthermore, RTC in SL was significantly lower than that in CI (P =.0001). These observations suggest that RTC value in liver tissues may digitally indicate a replicative history of hepatocytes under chronic inflammation, and a risk of HCC development.

Adult↗
PubMed1999

Organ-specific pancreatic tumor growth properties and tumor immunity.

We established a model of orthotopic injection of a syngeneic pancreatic tumor cell line in C57BL/6 mice and evaluated the effects of organ site on induction of immunity to a tumor-specific antigen, MUC1. Mice were challenged with a syngeneic pancreatic adenocarcinoma cell line that expressed MUC1 (Panc02-MUC1) by orthotopic injection into the pancreas, or by subcutaneous injection. Tumor cells injected into the pancreas grew much faster than those injected subcutaneously. Mice challenged subcutaneously with Panc02-MUC1 rejected tumors or developed slowly growing tumors that were negative for MUC1 expression. In contrast, mice challenged orthotopically into the pancreas developed progressive tumors that were positive for MUC1 expression. Sera from mice that rejected PancO2-MUC1 (tumor-immune mice) showed no detectable IgG1 and IgM titers against the MUC1 tandem-repeat peptide, whereas mice with progressive tumor growth had significant titers of IgG1 and IgM specific for MUC1. This suggests that the humoral immune response was ineffective in mediating tumor rejection. The results show that the growth properties and immunological rejection of pancreatic tumors is affected by the organ site at which the tumor grows.

Amino Acid Sequence↗
PubMed1999

Isolation and characterization of a human hepatic epithelial-like cell line (AKN-1) from a normal liver.

The isolation and characterization of human liver cell lines are rather difficult due to limited material and poor growth in cell culture. In this report, we present the isolation, culture and characterization of a new epithelial-like liver cell line (AKN-1) with a heterogeneous cell population and many characteristics of the biliary epithelium. The AKN-1 cell line stained positively with antibodies to epithelial cytokeratin polypetides CK 8, 18, and 19. In addition, the cell line expressed the anti-human epithelial-related antigen (MOC-31), the human epithelial antigen (HEA), and the gamma-glutamyl transpeptidase, the hematopoietic growth factor, stem cell factor, and also its receptor, c-kit. The cell line failed to express albumin and factor 8 by immunohistochemistry. It did show, however, a twofold increase in 7-ethoxyresorufin-O-deethylase activity. Cytogenetic characterization revealed rare breakpoints in chromosome 2, which to our knowledge, have not yet been reported in liver cells.

Biomarkers↗