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T Kohno

Publications and source records attributed to T Kohno.

At least 163 records · Page 9Linked to original sources
PubMed1998

Inactivation of the PTEN/MMAC1/TEP1 gene in human lung cancers.

The PTEN/MMAC1/TEP1 gene has been isolated as a tumor suppressor gene that is altered in several types of human tumors including brain, breast, and prostate cancers. In the present study, we report PTEN/MMAC1/TEP1 alterations in human lung cancers. Intragenic homozygous deletions were detected in 6 (40%) of 15 small cell lung carcinoma (SCLC) cell lines and in 2 (8%) of 25 non-small cell lung carcinoma (NSCLC) cell lines. A nonsense mutation and a missense mutation were detected in 2 (8%) NSCLC cell lines. An intragenic homozygous deletion, a 1-bp frameshift mutation, and a nonsense somatic mutation were also detected in three (6%) of 47 surgical specimens. All the lung tumors with PTEN/MMAC1/TEP1 mutations were homozygous for the mutant alleles. These findings suggest that PTEN/MMAC1/TEP1 plays a role as a tumor suppressor gene in the genesis and/or progression of human lung cancer.

Carcinoma, Non-Small-Cell Lung↗
PubMed1998

Association of CDKN2A(p16)/CDKN2B(p15) alterations and homozygous chromosome arm 9p deletions in human lung carcinoma.

To elucidate the possibility of the existence of multiple tumor suppressor genes on chromosome arm 9p, we performed genetic and epigenetic analyses of the CDKN2A/p16/MTS1 and CDKN2B/p15/MTS2 genes as well as homozygous deletion mapping of 9p in human lung carcinoma. To avoid overlooking genetic alterations due to contamination of noncancerous cells, we examined 32 non-small cell lung carcinoma (NSCLC) and 16 cell small cell lung carcinoma (SCLC) cell lines. (CDKN2A was mutated or homozygously deleted in 20 (63%) of 32 NSCLC cell lines, and methylation of the CpG island in the CDKN2A gene was detected in six of the 12 cell lines carrying the wild-type CDKN2A gene. Although homozygous deletions of the CDKN2B gene were also detected in NSCLC cell lines with CDKN2A deletions, mutation and methylation in the CDKN2B gene were infrequent. Thus, it was indicated that the CDKN2A gene rather than the CDKN2B gene plays a critical role as a tumor suppressor gene in NSCLC. Homozygous deletions on 9p were detected in 14 (44%) NSCLC cell lines. It is of note that two common regions of homozygous deletions were mapped proximal to the CDKN2A and CDKN2B loci, suggesting that tumor suppressor genes other than CDKN2A are present on 9p. In contrast to NSCLC, homozygous deletions on 9p as well as CDKN2A and CDKN2B alterations were infrequent in SCLC. Therefore, the pathogenetic significance of 9p alterations is likely to differ between SCLC and NSCLC.

Carcinoma, Small Cell↗
PubMed1998

Development of a highly sensitive and specific two-site enzyme immunoassay for parathyroid hormone (1-34): application to pharmacokinetic study on intranasal parathyroid hormone (1-34) in human.

A highly sensitive and specific two-site enzyme immunoassay for parathyroid hormone (1-34) (PTH(1-34)) and its usability for the pharmacokinetic study are described. Plasma samples were incubated simultaneously with 2,4-dinitrophenylated anti-PTH(1-34) IgG and anti-PTH(1-34) Fab'-beta-D-galactosidase conjugate. The immune complex formed of the three components was trapped onto (anti-2,4-dinitrophenyl group) IgG-coated polystyrene balls. beta-D-Galactosidase activity bound to the polystyrene balls was assayed by fluorometry. The practical detection limit of PTH(1-34) was 50 fg (12 amol)/0.05 ml of sample and 1 pg/ml as the concentration and practically no interference occurred by PTH(1-84) and PTH-related protein (1-34) up to 300 pg/ml and 10 ng/ml, respectively. The application of this method has enabled us to directly estimate the bioavailability of PTH(1-34) dosed intranasally at the prescribed level (0.090 mg). The pharmacokinetic parameters of the intranasal PTH(1-34) (n = 4) thus estimated were as follows: the area under the plasma concentration-time curve (AUC) = 20,500+/-15,900(SD) pg.min/ml; the mean residence time (MRT) = 194+/-16.3(SD) min; and the maximal concentration (Cmax) = 98+/-51 (SD) pg/ml with the maximal time (Tmax) = 35.0+/-12.2(SD) min.

Adult↗
PubMed1998

Determination of the bioavailability of intranasal elcatonin in humans: development of a sandwich transfer enzyme immunoassay for elcatonin.

A sandwich transfer enzyme immunoassay for elcatonin (ECT) and its usability for the pharmacokinetic study are described. The anti-salmon calcitonin (SCT) antibody was used for the present assay. The assay procedure consisted of the reaction of ECT with 2,4-dinitrophenylbiotinyl anti-SCT IgG and anti-SCT Fab'-beta-D-galactosidase conjugate, trapping onto (anti-2,4-dinitrophenyl bovine serum albumin) IgG-coated polystyrene balls, eluting with epsilonN-2,4-dinitrophenyl-L-lysine and transferring to streptavidin-coated polystyrene balls and fluorometric detection of beta-D-galactosidase activity. The practical detection limit of ECT was 0.15 pg (44 amol)/50 microl of sample and 3 pg/ml as the concentration. The application of this method has enabled us to directly estimate the bioavailability of ECT dosed intranasaly at a therapeutic level (100 IU, 17 microg) for its anti-osteoporotic effect as compared to an intramuscular dose (40 IU, 6.7 microg). The pharmacokinetic parameters of the intranasal ECT (n = 6) thus estimated were as follows: the area underthe serum concentration-time curve (AUC) = 2,570 +/- 1,650 (SD) pg x min/ml, and the maximal concentration (Cmax) = 60 +/- 25 (SD) pg/ml with the maximal time (Tmax) = 17.5 +/- 6.9 (SD) min, when the AUC for the intramuscular ECT (n = 9) = 9,460 +/- 5,870 (SD) pg x min/ml and the Cmax = 165 +/- 79 (SD) pg/ml with the Tmax = 16.1 +/- 4.2 (SD) min.

Administration, Intranasal↗
PubMed1998

Suppression of experimental autoimmune encephalomyelitis with a TNF binding protein (TNFbp) correlates with down-regulation of VCAM-1/VLA-4.

The effect of a novel TNF binding protein (TNFbp), a polyethylene glycol-linked form of the type I soluble receptor of TNF, on the expression of adhesion molecules has been investigated with a passive transfer model of experimental autoimmune encephalomyelitis (EAE) in SJL/J mice. The expression of L-selectin, VLA-4 and LFA-1 on spleen cells of EAE animals treated with TNFbp or saline was examined by FACS analysis. The expression of VCAM-1 and ICAM-1 was investigated by immunochemistry in spinal cord tissue of SJL/J mice with EAE. In animals sensitized for EAE and treated with TNFbp, the expression of VCAM-1 in the central nervous system as well as VLA-4 on spleen cells was clearly diminished. Reduction in VCAM-1 staining and VLA-4 expression corresponded to inhibition of inflammation in the spinal cord and to prevention of clinical signs of EAE. The results have also shown that myelin basic protein responses as well as non-antigen-specific responses were not diminished in animals treated with TNFbp. The findings suggest that TNFbp might prevent EAE development by modulating the expression of VCAM-1 and VLA-4.

Animals↗
PubMed1998

Genomic structure and chromosomal localization of the mouse Ogg1 gene that is involved in the repair of 8-hydroxyguanine in DNA damage.

8-Hydroxyguanine (7,8-dihydro-8-oxoguanine: oh8Gua) is a damaged form of guanine induced by oxygen-free radicals and causes GC to TA transversions. Previously we isolated the hOGG1 gene, a human homolog of the yeast OGG1 gene, which encodes a DNA glycosylase and lyase to excise oh8Gua in DNA. In this study, we isolated a mouse homolog (Ogg1) of the OGG1 gene, characterized oh8Gua-specific DNA glycosylase/AP lyase activities of its product, and determined chromosomal localization and exon-intron organization of this gene. A predicted protein possessed five domains homologous to human and yeast OGG1 proteins. Helix-hairpin-helix and C2H2 zinc finger-like DNA-binding motifs found in human and yeast OGG1 proteins were also retained in mouse Ogg1 protein. The properties of a GST fusion protein were identical to human and yeast OGG1 proteins in glycosylase/lyase activities, their substrate specificities, and suppressive activities against the spontaneous mutagenesis of an Escherichia coli mutM mutY double mutant. The mouse Ogg1 gene was mapped to Chromosome (Chr) 6, and consisted of 7 exons approximately 6 kb long. Two DNA-binding motifs were encoded in exons 4 through 5. These data will facilitate the investigation of the OGG1 gene to elucidate the relationship between oxidative DNA damage and carcinogenesis.

Amino Acid Sequence↗
PubMed1998

Choriocapillary changes and hyperfluorescence on indocyanine green angiography.

BACKGROUND: The purpose of this study is to examine the relationship between pathological changes of the choriocapillaris (CC) and the finding of bright fluorescence on indocyanine green (ICG) angiography. METHODS: An animal model was used, with chorioretinal lesions produced by injecting sodium iodate 3 h, 24 h, 7 days or 14 days previously. The ICG angiographic findings were compared with histology. RESULTS: Three hours after injection, many spots of bright fluorescence were scattered at the posterior pole. Histologically, variable changes in CC endothelial cells were observed. In some regions, the cells remained almost normal morphologically. In other regions, the endothelial cytoplasm was thickened with decreased fenestrations, or precipitation of fibrin in the choroidal interstitium was observed. Twenty-four hours after injection, the area of bright fluorescence had extended beyond that observed after 3 h. Histologically, the cytoplasmic structure of CC endothelial cells was unclear, and the endothelial walls were detached from the basement membrane. When ferritin was administered as a tracer, many ferritin granules were observed in Bruch's membrane. Seven days after injection, the degree of bright fluorescence was reduced as compared with 3 and 24 h after injection. Histologically, the cytoplasm of CC endothelial cells was thinned, but contained few fenestrations. The endothelial walls were detached from the basement membrane. Fourteen days after injection, normal diffuse ICG fluorescence was observed. Histologically, CC endothelial cells exhibited almost normal structure. CONCLUSION: These findings indicate that changes in CC endothelial cells and bright fluorescence on ICG angiography are closely related. ICG angiography should enable clinical evaluation of increased permeability of CC endothelial cells to relatively large molecules.

Animals↗
PubMed1998

Identification of CpG islands hypermethylated in human lung cancer by the arbitrarily primed-PCR method.

DNA hypermethylation is believed to be involved in human carcinogenesis, since it suppresses the transcription of defined genes and is associated with chromosomal instability. In this study, we identified CpG islands that are hypermethylated in human lung cancer by a modified arbitrarily primed-polymerase chain reaction method using genomic DNAs digested with a methylation-sensitive restriction enzyme, HpaII, as templates. When we analyzed genomic DNAs from normal lung tissues and non-small cell lung carcinoma cell lines using three arbitrary primers, three DNA fragments were amplified from lung cancer DNAs but not from normal lung DNAs. Restriction mapping and Southern blot analysis revealed that all of these bands were amplified from CpG islands that were hypermethylated in the lung cancer cell lines. These islands were mapped to chromosomes 4q34, 10q26 and 17p13.1-p13.2, respectively, and these chromosomal regions were also hypermethylated in a subset of primary lung tumors in vivo. Thus, diverse chromosomal regions are hypermethylated in lung cancer cells. The results also indicate that this method is simple and effective for screening of CpG islands that are hypermethylated in cancer cells.

Carcinoma↗
PubMed1998

Choroidopathy after blunt trauma to the eye: a fluorescein and indocyanine green angiographic study.

PURPOSE: To describe the uses of fluorescein angiography and indocyanine green angiography in highlighting traumatic choroidopathy. METHODS: In a prospective study, 21 patients (21 eyes) who presented with traumatic retinal opacity ophthalmoscopically underwent fluorescein angiography and indocyanine green angiography. With indocyanine green angiography, the subtraction method was used for detailed examination. RESULTS: In 11 of 21 eyes, fluorescein angiography showed no abnormalities. On indocyanine green angiography, delayed filling in the choroid was found locally in nine of these 11 eyes. Delayed filling of the choroidal veins was clearly visualized by the subtraction method. In six eyes, intrachoroidal indocyanine green leakage around the choroidal vessels, including vortex veins, was found. In the remaining 10 of the 21 eyes, fluorescein angiography showed fluorescein leakage or a salt-and-pepper appearance in the region of retinal opacity. On indocyanine green angiography, a triangular-shaped area of delayed filling of the choroidal arteries was observed in four eyes. Delayed filling of the choroidal veins was visualized in all 10 eyes, intrachoroidal indocyanine green leakage was found in eight, and dilation and/or narrowing of the choroidal veins and changes in choroidal vasculature were visualized in five eyes. Furthermore, in regions with fluorescein leakage, indocyanine green leakage or hypofluorescence was observed, suggesting damage to the choriocapillaris. CONCLUSION: Indocyanine green angiography allows the analysis of various degrees of choroidal vascular damage. Because the choroidal circulation nourishes the outer retinal layer, traumatic choroidopathy may play a role in the prognosis for visual recovery in eyes affected by contusion retinal opacities.

Adolescent↗
PubMed1998

Repair of ruptured anastomotic aneurysm: elephant trunk technique after endovascular covered stents.

Endovascular covered stents were successfully applied to temporarily halt hemoptysis and postpone surgical intervention in a 69-year-old man with a ruptured anastomotic false aneurysm of the distal aortic arch. Surgical graft implantation was performed successfully by the elephant trunk technique 14 days after the endovascular stent-grafting, at which time aspiration pneumonia had subsided.

Aged↗
PubMed1998

Accelerated entry into S phase associated with up-regulation of cyclin D1 as a mechanism for granulocyte colony-stimulating factor (G-CSF)-induced apoptosis of murine myeloid leukemia cells.

We previously reported that injection of recombinant granulocyte colony-stimulating factor (G-CSF) suppressed the development of leukemia in mice transplanted with C2M-A5 (C2M) myeloid leukemia cells and that the anti-leukemic effect of G-CSF was ascribed to the induction of apoptosis of C2M cells. These observations make a striking contrast with other previous reports on the biological activities of G-CSF. In the present study, in order to further clarify the G-CSF-induced apoptosis of C2M cells, we studied the effects of G-CSF on the cell cycle as well as the molecular events involving D-type cyclines and their cyclin-dependent kinases (cdk) in G-CSF-treated C2M cells. Cell cycle analysis revealed that G-CSF treatment of C2M cells resulted in accelerated entry from the first gap (G1) phase into the DNA synthesis (S) phase. Western blotting disclosed that G-CSF treatment resulted in down-regulation of cyclin D2 and cdk2 and up-regulation of cyclin D1 and cdk4. The reciprocal relationship between the up-regulation of cyclin D1 and down-regulation of cyclin D2 was closely associated with accelerated entry into S phase and subsequent apoptosis of C2M cells. These results suggest that G-CSF-induced apoptosis of C2M cells might be ascribed to imbalanced cell cycle progression due to deregulated expression of D-type cyclins and their cdks.

Animals↗
PubMed1998

Contrast-enhancement for the image of human immunodeficiency virus from ultrathin section by immuno electron microscopy.

A simple contrast-enhancement method is described for electron microscopic imaging of the human immunodeficiency virus (HIV) from a sample embedded in Lowicryl K4M resin, by immuneelectron microscopy. Ultrathin sections were treated with a mixture of ruthenium red dye (RR) and osmium tetroxide (OSO4). This treatment provided good contrast enhancement of the entire ultrastructural image of virus particles without the loss of immunolabelling. RR/OsO4 solution is simple to prepare and provides a better contrast than that which is achieved during conventional post-embedding immunoelectron microscopy. Treatment of ultrathin sections from low temperature-embedded samples with RR/OsO4 solution is recommended.

Cell Line↗
PubMed1998

Dynamics of the low altitude secondary proton radiation belt.

At the interface between the upper atmosphere and the radiation belt region, there exists a secondary radiation belt consisting mainly of energetic ions that have become neutralized in the ring current and the main radiation belt and then re-ionized by collisions in the inner exosphere. The time history of the proton fluxes in the 0.64-35 MeV energy range was traced in the equatorial region beneath the main radiation belts during the three year period from 21 February 1984 to 26 March 1987 using data obtained with the HEP experiment on board the Japanese OHZORA satellite. During most of this period a fairly small proton flux of -1.2 cm-2 S-1 sr-1 was detected on geomagnetic field lines in the range 1.05 < L < 1.15. We report a few surprisingly deep and rapid flux decreases (flux reduction by typically two orders of magnitude). These flux decreases were also long in duration (lasting up to three months). We also registered abrupt flux increases where the magnitude of the proton flux enhancements could reach three orders of magnitude with an enhancement duration of 1-3 days. Possible reasons for these unexpected phenomena are discussed.

Atmosphere↗
PubMed1998

A new general method for the biosynthesis of stable isotope-enriched peptides using a decahistidine-tagged ubiquitin fusion system: an application to the production of mastoparan-X uniformly enriched with 15N and 15N/13C.

A new strategy is described for the production of peptides enriched with stable isotopes. Peptides of interest are expressed in Escherichia coli (E. coli) cells as recombinant fusion proteins with Saccharomyces cerevisiae ubiquitin. This method yields as much as 30-100 mg/l of isotope-enriched fusion proteins in minimal media. A decahistidine tag attached to the N-terminus of ubiquitin enables a one-step purification of the fusion protein via Ni(2+)-chelating affinity chromatography. The ubiquitin moiety is then easily and specifically cleaved off by a protease, yeast ubiquitin hydrolase. Since this enzyme is also expressed at a high level in E. coli cells and can be purified in one step, the presented strategy has an advantage in view of costs over others that use commercially available proteases. In addition, since ubiquitin fusion proteins easily refold, the fusion protein can be expressed either in a soluble form or as inclusion bodies. This flexibility enables us to prepare peptides that are unstable in a soluble state in E. coli cells. As an example, the expression and the uniform stable isotope enrichment with 15N and/or 13C are described for mastoparan-X, a tetradecapeptide known to activate GTP-binding regulatory proteins. An amide group at the C-terminus of this peptide can also be formed by our method. The presented system is considered powerful for the stable isotope enrichment of short peptides with proton resonances that are too severely overlapped to be analyzed solely by proton NMR.

Amino Acid Sequence↗
PubMed1998

Phenotypic alterations of small cell lung carcinoma induced by different levels of wild-type p53 expression.

p53 induces both growth arrest and apoptosis in cancer cells. To clarify whether the level of p53 expression determines the response of small cell lung carcinoma (SCLC) cells, we assessed the effect of various p53 levels on a p53-null SCLC cell line, N417, using a tetracycline (Tc)-regulated inducible p53 expression system. Apoptosis was induced in SCLC cells with high p53 expression. Although low levels of p53 induced G1 arrest accompanied by p21 expression, cells with G1 arrest seemed to undergo apoptosis after further cultivation. Expression of exogenous p21 induced G1 arrest but not apoptosis in SCLC cells, suggesting that p53-mediated G1 arrest was induced through p21 expression. Moreover, high level of p53 expression down-regulated Bcl-2 expression in SCLC cells, while Bax was consistently expressed irrespective to the level of p53 expression. These results suggest that p53-mediated apoptosis and G1 arrest depend on level of p53 expression in SCLC cells and that the relative dominancy of Bax to Bcl-2 is involved in the induction of apoptosis by high level of p53 expression.

Apoptosis↗
PubMed1998

Identification and characterization of families with aggregation of lung cancer.

BACKGROUND: To clarify genetic factors involved in the susceptibility to lung cancer, it is essential to identify families with lung cancer clustering and to characterize the mode of clustering. Since somatic mutations of the p53, RB and p16 genes occur frequently in lung cancer and the replication error phenotype is seen in a subset of lung cancer, it is possible that germ-line mutations of the p53, RB, p16 and mismatch repair genes influence the susceptibility to lung cancer. METHODS: In this work, cases with familial clustering of lung cancer were selected from 1068 families with primary lung cancer cases in analogy with the criteria for hereditary non-polyposis colorectal cancer (HNPCC). Cases with Li-Fraumeni syndrome, familial retinoblastoma, familial melanoma and HNPCC were also searched among these 1068 families. RESULTS: There were only four families (0.4%) in which more than three relatives were affected by lung cancer. Two successive generations were affected in 36 families (3.4%). Patients with lung cancer before the age of 50 were present in 165 families (15.5%). However, no family conformed to all three criteria. There was only one family with Li-Fraumeni syndrome and no family with familial retinoblastoma, familial melanoma and HNPCC. CONCLUSION: Familial aggregation of lung cancer is rare and germ-line mutations of the p53, RB, p16 and mismatch repair genes may not contribute greatly to susceptibility to lung cancer.

Colorectal Neoplasms, Hereditary Nonpolyposis↗
PubMed1998

Infrequent mutations of the hOGG1 gene, that is involved in the excision of 8-hydroxyguanine in damaged DNA, in human gastric cancer.

DNA glycosylase, encoded by the hOGG1 gene, repairs 8-hydroxyguanine (oh8Gua), which is an oxidatively damaged mutagenic base. To clarify whether the DNA repair activity of hOGG1 protein is involved in gastric carcinogenesis, we examined 9 gastric cancer cell lines and 35 primary gastric cancers for mutations and genetic polymorphisms of the hOGG1 gene by polymerase chain reaction-single strand conformation polymorphism analysis. A G-to-A transition was detected in a gastric cancer cell line, MKN1. This nucleotide change caused the conversion of the amino acid from Arg to His at codon 154, which is located in a domain highly conserved among human, mouse, and yeast OGG1 proteins. No mutation was detected in primary gastric cancers. We compared the distribution of the polymorphic alleles associated with enzymatic activity (hOGG1-Ser326 vs. hOGG1-Cys326) between 35 gastric cancer patients and 42 healthy individuals. Although the frequency of the Cys326 allele, associated with low enzymatic activity, in gastric cancer patients was a little higher than that in healthy individuals, the difference did not reach statistical significance. These results suggest that low hOGG1 activity due to mutations and genetic polymorphisms is involved in the development of only a small subset of gastric cancers.

DNA Damage↗