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Biomedical subjects

E Gotoh

Publications and source records attributed to E Gotoh.

At least 19 recordsLinked to original sources
PubMed2006

Mitotic arrest caused by an X-ray microbeam in a single cell expressing EGFP-aurora kinase B.

Although the highest radiosensitivity of cells in the M phase among the other cell phases, such as the G(1), S and G(2) phases, has been known, the exact mechanism of radiosensitivity in mitotic cells remains unclear. Recently, mitotic arrest caused by DNA-damaging reagents has been shown, and the molecular mechanism in the arrest has been discussed in detail. In this study, abnormal cell-cycle progression in the M phase was investigated when a single mitotic cell in each mitotic stage was irradiated with a 5.35 keV X-ray microbeam focused on the cell nucleus. An X-ray microbeam irradiation system installed at BL-27 in Photon Factory, High Energy Accelerator Research Organization (HEARO, Tsukuba) was used. HeLa cells, genetically modified and expressing enhanced green fluorescent protein-tagged aurora kinase B, were used as irradiated samples in order to recognise the stage of each cell in the M phase. Thus, 10 Gy irradiation concentrated at the nucleus of a single cell elongated the cell-cycle progression in the M phase by delaying the metaphase/anaphase transition. The dose dependence of the elongation of the M phase was also examined. An irregular distribution of DNA in anaphase cells was observed after irradiation.

Aurora Kinase B↗
PubMed2005

Telomere size and telomerase activity in Epstein-Barr virus (EBV)-positive and EBV-negative Burkitt's lymphoma cell lines.

The telomere repeat lengths of BL cell lines were quantified by measuring terminal restriction fragment (TRF). Epstein-Barr virus (EBV)-positive Namalwa, Raji, and EB-3 cell lines have long telomeres, i.e. TRFs 10-19 kbp, whereas the Daudi cell line, producing a transformation-defective EBV mutant, has TRFs approximately 2.2 kbp. EBV-negative BJAB and DG75 cell lines have short TRFs 3.9-5.4 kbp, shorter than the approximately 12 kbp TRFs in PBLs. Telomerase activities of these BL cell lines are similar. TRFs of non-BL lymphoma cell lines are 2.3-5.5 kbp. Fluorescent in situ hybridization (FISH) studies of these cell lines showed remarkable heterogeneity of telomere size in chromosomes in the same BL cell. These results suggest that EBV-positive and EBV-negative BL cell lines have experienced various telomere dynamics.

Base Sequence↗
PubMed2005

Simple biodosimetry method for use in cases of high-dose radiation exposure that scores the chromosome number of Giemsa-stained drug-induced prematurely condensed chromosomes (PCC).

There is a need for quick dose estimation by a simple method in radiation accidents. This study develops a simple and rapid dose estimation protocol for victims of such accidents, in particular those involving high radiation doses. Human peripheral blood lymphocytes (PBL) were gamma-irradiated in vitro at several dose points up to 60 Gy, and were stimulated with phytohaemagglutinin-P (PHA-P) for 2 days to obtain dividing cells. PBL were then forced to condense prematurely, using 50 nM calyculin A, and the obtained chromosome spreads were Giemsa stained. The G2-PCC (prematurely condensed chromosomes) index and chromosome number for each radiation dose point were scored. G2-PCC were stably induced using calyculin A within 24 h delays in stimulation of PBL with PHA-P. The chromosome number of G2-PCC increased steeply with radiation doses up to 30 Gy at a rate of 0.31 Gy(-1) and then decreased at 0.30 Gy(-1) up to 40 Gy. More than 10% of G2-PCC index remained up to a 15 Gy dose. Even after 40 Gy irradiation, about 2% PCC index was obtained, and this value was enough to score a sufficient number of chromosome spreads for analysis. Therefore, the combined use of chromosome number and G2-PCC index allows biodosimetry to be done easily and rapidly. If PCC are not induced using calyculin A, it is strongly suggested that the radiation dose is over 50 Gy. A rapid and easy dose estimation for large dose exposure whole-body was realized by combined analysis of Giemsa-stained chromosome number of G2-PCC and PCC index using calyculin A. This simple method will be of use for rapid decision making of therapy for radiation accident victims. This method also has potential for use as a biodosimeter for partial-body exposure accidents.

Adult↗
PubMed2005

Simple biodosimetry method for cases of high-dose radiation exposure using the ratio of the longest/shortest length of Giemsa-stained drug-induced prematurely condensed chromosomes (PCC).

The aim was to develop a simple biodosimetry method for as rapid as possible estimation of absorbed radiation doses in victims of radiation accidents, in particular after high-dose exposure. Human peripheral blood lymphocytes (PBL) were gamma-irradiated in vitro with several doses up to 40 Gy stimulated with phytohaemagglutinin-P (PHA-P) for 2 days and their chromosomes condensed prematurely using 50 nm calyculin A. Chromosome lengths of Giemsa-stained G2 prematurely condensed chromosomes (PCC) were measured using image analysing software and the ratio of the longest/shortest chromosome length was calculated. The length ratio (LR) of the longest/shortest Giemsa-stained chromosome s increased with a good correlation to the square root of the radiation dose (D) up to 40 Gy, i.e. LR = (4.90 x D0.5) + 2.14. The LR of the longest/shortest chromosome might be used as an index for estimating the radiation dose. The blood samples should not be cooled until the start of separation/stimulation of the lymphocytes. A rapid and easy estimation of large doses after whole-body exposure was identified by measuring the ratio of the longest/shortest length of Giemsa-stained G2-PCC induced by calyculin A. This simple protocol will be particularly useful for making therapy decisions for victims of ionizing radiation exposure and has potential for use as a biodosimeter for partial-body exposure accidents.

Azure Stains↗
PubMed2001

Kinetics of chromatid break repair in G2-human fibroblasts exposed to low- and high-LET radiations.

The purpose of this study is to determine the kinetics of chromatid break rejoining following exposure to radiations of different quality. Exponentially growing human fibroblast cells AG1522 were irradiated with gamma-rays, energetic carbon (290 MeV/u), silicon (490 MeV/u) and iron (200 MeV/u, 600 MeV/u). Chromosomes were prematurely condensed using calyculin A. Prematurely condensed chromosomes were collected after several post-irradiation incubation times, ranging from 5 to 600 minutes, and the number of chromatid breaks and exchanges in G2 cells were scored. The relative biological effectiveness (RBE) for initial chromatid breaks per unit dose showed LET dependency having a peak at 55 keV/micrometers silicon (2.4) or 80 keV/micrometers carbon particles (2.4) and then decreased with increasing LET. The kinetics of chromatid break rejoining following low- or high-LET irradiation consisted of two exponential components. Chromatid breaks decreased rapidly after exposure, and then continued to decrease at a slower rate. The rejoining kinetics was similar for exposure to each type of radiation, although the rate of unrejoined breaks was higher for high-LET radiation. Chromatid exchanges were also formed quickly.

Cell Line↗
PubMed2000

High-LET radiation-induced aberrations in prematurely condensed G2 chromosomes of human fibroblasts.

PURPOSE: To determine the number of initial chromatid breaks induced by low- or high-LET irradiations, and to compare the kinetics of chromatid break rejoining for radiations of different quality. MATERIAL AND METHODS: Exponentially growing human fibroblast cells AG1522 were irradiated with gamma-rays, energetic carbon (290MeV/u), silicon (490MeV/u) and iron (200 and 600 MeV/u). Chromosomes were prematurely condensed using calyculin A. Chromatid breaks and exchanges in G2 cells were scored. PCC were collected after several post-irradiation incubation times, ranging from 5 to 600 min. RESULTS: The kinetics of chromatid break rejoining following low- or high-LET irradiation consisted of two exponential components representing a rapid and a slow time constant. Chromatid breaks decreased rapidly during the first 10min after exposure, then continued to decrease at a slower rate. The rejoining kinetics were similar for exposure to each type of radiation. Chromatid exchanges were also formed quickly. Compared to low-LET radiation, isochromatid breaks were produced more frequently and the proportion of unrejoined breaks was higher for high-LET radiation. CONCLUSIONS: Compared with gamma-rays, isochromatid breaks were observed more frequently in high-LET irradiated samples, suggesting that an increase in isochromatid breaks is a signature of high-LET radiation exposure.

Cell Line↗
PubMed2000

Acetabular development after reduction in developmental dislocation of the hip.

A radiologic study was conducted on 56 patients with developmental dislocation of the hip (63 hips). Fifty hips in which neither acetabular nor femoral osteotomy was performed were classified as satisfactory (Severin Groups I and II) or unsatisfactory (Severin Groups III and IV) based on radiographs when growth was completed. The sequential changes in the center edge angle and the acetabular index were compared when the patients were ages of 5, 10, and 15 years. There was a significant relationship between the center edge angle and the acetabular index when the patients were 5 years of age and at final outcome. Most (85.7%) patients with a center edge angle less than 8 degrees and an acetabular index greater than 26 degrees at 5 years of age eventually were classified as Severin Groups III and IV at skeletal maturity. These findings suggest that radiologic results at the time when growth is completed can be predicted based on the center edge angle and the acetabular index in radiologic measurements at 5 years of age. The authors recommend that if at 5 years of age the center edge angle is less than 8 degrees and the acetabular index is greater than 26 degrees, consideration be given to an osteotomy to bring these values to a more normal range to improve final outcome.

Acetabulum↗
PubMed2000

Angiotensin-(1-7) attenuates vasoconstriction evoked by angiotensin II but not by noradrenaline in man.

Angiotensin-(1-7) has been suggested to be a novel vasodilating peptide. We investigated the direct vascular effect of angiotensin-(1-7) in human forearm resistant vessels, particularly with regard to the interaction with angiotensin II, in healthy normotensive men by strain-gauge venous occlusion plethysmography with intra-arterial infusions of peptides. Intra-arterial infusion of angiotensin-(1-7) at 0.1 to 2000 pmol/min did not cause vasodilatation but rather reduced forearm blood flow by approximately 10% at the highest dose. A placebo-controlled study showed that angiotensin-(1-7) at 0.5 to 40 nmol/min caused weak but significant vasoconstriction (P=0.0016 by ANOVA). Angiotensin-(1-7) at 100 pmol/min, but not at 10 pmol/min, significantly shifted the angiotensin II dose-response curve toward the right (mean+/-SD of percent changes in forearm blood flow: -19+/-17%, -33+/-22%, -55+/-12%, -63+/-10%, and -68+/-5% at 5, 10, 25, 50, and 100 pmol/min of angiotensin II, respectively, with saline; 5+/-13%, 0. 9+/-18%, -40+/-16%, -54+/-9%, and -61+/-6% with angiotensin-(1-7), P=0.0021 by ANOVA). Angiotensin-(1-7) did not affect the dose-response curve of noradrenaline [3+/-12%, 5+/-16%, -20+/-22%, -31+/-18%, and -40+/-12% at 25, 50, 100, 300, and 600 pmol/min of noradrenaline, respectively, with saline; -4+/-15%, -2+/-23%, -29+/-22%, -34+/-16%, and -42+/-9% with angiotensin-(1-7)]. Our results suggest that angiotensin-(1-7) antagonizes vasoconstriction by angiotensin II in human resistant vessels and might act as an endogenous angiotensin II antagonist.

Angiotensin I↗
PubMed1999

Targeting double-strand breaks to replicating DNA identifies a subpathway of DSB repair that is defective in ataxia-telangiectasia cells.

The critical cellular defect(s) and basis for cell killing by ionizing radiation in ataxia-telangiectasia (A-T) are unknown. We use the topoisomerase I inhibitor camptothecin (CPT), which kills mainly S-phase cells and induces DSBs predominantly in replication forks, to show that A-T cells are defective in the repair of this particular subclass of DSBs. CPT-treated A-T cells reaching G2 have abnormally high levels of chromatid exchanges (viewed as prematurely condensed G2 chromosomes); aberrations in normal cells are mostly chromatid breaks. Transfectants of A-T cells with the wild-type ATM cDNA are corrected for CPT sensitivity, chromatid aberrations, and the DSB repair defect. These data suggest that in normal cells ATM, the A-T protein, probably recognizes DSBs in active replicons and targets the repair machinery to the breaks; in addition, the ATM protein is involved in the suppression of low-fidelity, adventitious rejoining between replication-associated DSBs. The loss of ATM functions therefore leads to genome destabilization, sensitivity to DSB-inducing agents and to the cancer-promoting illegitimate exchange events that follow.

Ataxia Telangiectasia↗
PubMed1999

Chromatid break rejoining and exchange aberration formation following gamma-ray exposure: analysis in G2 human fibroblasts by chemically induced premature chromosome condensation.

PURPOSE: To analyse the kinetics of chromatid break induction, rejoining, and misrejoining after y-irradiation in G2 phase human cells using premature chromosome condensation induced by calyculin A. MATERIALS AND METHODS: Human fibroblast AG1522 cells were irradiated with gamma-rays and chromosomes were then prematurely condensed by calyculin A. The number of chromatid breaks and chromatid exchanges in G2 chromosomes were scored, and fitted curves were calculated. RESULTS: Calyculin A induced premature chromosome condensation in cells immediately after irradiation. Kinetics of rejoining of chromatid breaks demonstrated two exponential components with rapid and slow time constants. Within 5 min after irradiation, the number of chromatid breaks fell rapidly to about one-half, then gradually decreased. Chromatid exchanges were formed very quickly, reaching a plateau within 20 min from exposure. CONCLUSIONS: Chemically induced premature chromosome condensation technique allows a simple, rapid and precise analysis of chromatid breakage and rejoining. The rapid kinetic component was particularly well characterized.

Cells, Cultured↗
PubMed1999

Association between G2-phase block and repair of radiation-induced chromosome fragments in human lymphocytes.

We have studied the induction of chromosomal aberrations in human lymphocytes exposed in G0 to X rays or carbon ions. Aberrations were analyzed in G0, G1, G2 or M phase. Analysis during the interphase was performed by chemically induced premature chromosome condensation, which allows scoring of aberrations in G1, G2 and M phase; fusion-induced premature chromosome condensation was used to analyze the damage in G0 cells after incubation for repair; M-phase cells were obtained by conventional Colcemid block. Aberrations were scored by Giemsa staining or fluorescence in situ hybridization (chromosomes 2 and 4). Similar yields of fragments were observed in G1 and G2 phase, but lower yields were scored in metaphase. The frequency of chromosomal exchanges was similar in G0 (after repair), G2 and M phase for cells exposed to X rays, while a lower frequency of exchanges was observed in M phase when lymphocytes were irradiated with high-LET carbon ions. The results suggest that radiation-induced G2-phase block is associated with unrejoined chromosome fragments induced by radiation exposure during G0.

Adult↗
PubMed1999

Minimum 10-year results of Sugioka's osteotomy for femoral head osteonecrosis.

Sugioka's transtrochanteric rotational osteotomy was performed on 14 hips (12 patients) before 1987 for avascular necrosis of the femoral head involving a large part of the weight-bearing area. Three hips required a secondary total hip arthroplasty within 5 years after the osteotomy. The remaining 11 hips were examined at a mean of 13.2 years after surgery (range, 10-17.7 years). The clinical and radiologic results were related to the preoperative radiographic stage of the disease. In the hips with less than 2 mm of collapse, highly satisfactory results were maintained more than 15 years after surgery, with minimal development of degenerative changes. In hips with 2 mm or more of collapse or with acetabular changes, the results tended to deteriorate gradually during the long course of observation but were fairly acceptable. This study shows the osteotomy can enable hip joints to survive and function well for more than 10 years with proper patient selection and operative procedure.

Adult↗
PubMed1998

A simple method for simultaneous interphase-metaphase chromosome analysis in biodosimetry.

PURPOSE: To find a simple protocol for measuring chromosome damage both in G1 and in G2/M chromosomes, to overcome problems related to low mitotic index and cell-cycle alterations in biodosimetric tests. MATERIALS AND METHODS: The protocol is based on the use of calyculin A to induce premature chromosome condensation in human peripheral blood lymphocytes in different phases of the cell cycle. Chromosome exchanges were measured by fluorescence in situ hybridization (chromosomes 2 and 4) in lymphocytes from four different donors. Cells were exposed to 4Gy X-rays and the results were compared to aberrations in M phase (colcemid block) and G0 (premature chromosome condensation induced by fusion to mitotic hamster cells). RESULTS: Treatment with calyculin A produced a high fraction of chromosome condensation in different phases of the cell cycle. Cells in G1 and G2/M could be scored simultaneously for biodosimetry by chromosome painting. The condensation index was 5-20 times higher than the mitotic index (colcemid alone). The calyculin A treatment did not produce a significant increase in the background of chromosomal aberrations or modify the yield of chromosomal aberrations scored after exposure to X-rays. CONCLUSIONS: Induction of chromosome condensation by calyculin A is a powerful biodosimetric tool, which provides a high number of spreads for analysis and overcomes problems related to poor in vitro growth or cell-cycle alterations.

Cell Cycle↗