Biomedical subjects

I Hayata

Publications and source records attributed to I Hayata.

At least 37 records · Page 2Linked to original sources

PubMed1999

Phosphatase inhibitors and premature chromosome condensation in human peripheral lymphocytes at different cell-cycle phases.

The cytogenetical reaction of human peripheral lymphocytes to okadaic acid and calyculin A was examined. Calyculin A could induce PCC about 20 times more effectively than okadaic acid. Their mechanisms of PCC induction were judged similar by their dose-dependent manner and chromosome morphology. Contrary to earlier studies suggesting that chemicals could not induce PCC in G1 cells where little cyclin B is present, the present study showed that calyculin A could induce PCC in lymphocytes not only at S and G2/M but also at the second G1 phase after PHA stimulation in vitro. PCC was induced slightly in lymphocytes both at G0 and the first in vitro G1 phase even when the calyculin A concentration increased one hundred fold. It was found that calcium ionophore A23187 increased frequencies of G0-PCC induced by calyculin A, although a further refinement is necessary to obtain a suitable morphology of G0-PCC for cytogenetic studies.

Calcimycin↗

PubMed1999

Easy biodosimetry for high-dose radiation exposures using drug-induced, prematurely condensed chromosomes.

PURPOSE: To propose a practical method of biodosimetry for high-dose estimation. MATERIALS AND METHODS: Okadaic acid in the range 100 nM to 500 nM was given for 1-6 h to cultured lymphocytes that had been exposed in the range of 0 Gy to 40 Gy. The frequency of the cells with prematurely condensed chromosomes (PCC) was compared with that of metaphases accumulated by 0.05 microg/ml colcemid. The dose-response relationship of the yield of prematurely condensed ring chromosomes (PCC rings) in the irradiated lymphocytes was examined. RESULTS: The 0.05 microg/ml colcemid collected few metaphases when lymphocytes were irradiated to over 10 Gy. However, 500 nM okadaic acid produced a substantial number of analysable PCC in lymphocytes that had been exposed in the range of 0 Gy to 40 Gy. After 1 h of treatment, the 500 nM okadaic acid gave the most suitable chromosome morphology for the analysis. PCC rings were narrow, which facilitated their detection in Giemsa-stained preparations. The yield of PCC rings increased dose-dependently up to 20 Gy. CONCLUSION: PCC rings are a more suitable indicator than dicentrics at high doses over 10 Gy. This technique requires no particular equipment or exceptional skill. After a serious radiation accident, it can be easily performed in a short time in most medical facilities to obtain dose information to assist physicians.

Azure Stains↗

PubMed1999

Induction of apoptosis by beta radiation from tritium compounds in mouse embryonic brain cells.

Induction of apoptosis by tritium exposure was investigated in both cultured embryonic mid brain cells and brain sections of embryos and of newborns in mice. In the cultures of mid brain cells, addition of methyl-3H-thymidine (3H-TdR) (21 kBq mL(-1)) and tritiated water (5.616 MBq mL(-1)) induced late appearances and low percentages of apoptosis when compared to x-irradiation at the ID50 dose, the inhibitory dose that reduced cellular differentiation by 50% of the control. A significant increase in p53 protein was detected about 2 h before the marked appearance of apoptosis. The pregnant mice were given an intraperitoneal injection of tritiated water at the concentration of 481.8 kBq g(-1) of body weight on gestation day 12.5, by which treatment behavioral changes in the offspring occurred. Increased apoptotic cells were observed in the neural tube of embryos from 1 d after the injection to 1 wk postnatal age. Apoptosis induced by x-rays appeared 2 h after irradiation, with a peak at 4 h. Increase of apoptotic cells was also found in the brain cortexes of newborns. The percentage of apoptosis in the brain was higher in the prenatal tritiated water exposed mice than in the prenatal x-irradiated mice. Possible mechanisms on apoptosis and its relation to the higher relative biological effectiveness value of tritium beta-rays are discussed.

Animals↗

PubMed1999

Effect of estradiol on radiation-induced chromosome aberrations in human lymphocytes.

As a part of studies on physiological factors that affect radiosensitivity, we examined the in vitro effect of estradiol (E2) on the yield of radiation-induced chromosome aberrations in human peripheral lymphocytes. Lymphocytes were cultured for 3 days in the medium containing E2 at 0-100,000 ng/ml. On the second day, they were irradiated by X-rays at 3 Gy, and then 2% phytohemagglutinin and 0.05 microgram/ml colcemid were added to the medium. After further 48 h, mitotic indices and the yields of chromosome aberrations were examined at various E2 concentrations. E2 treatment at concentrations above 1000 ng/ml resulted in dose-related inhibition of mitosis. Repeated experiments showed that the yield of dicentrics plus centric rings in the culture containing E2 at 100 ng/ml was significantly higher than the yields at 0 ng/ml. Similarly, the yield of total chromosome breaks in the culture containing E2 at 100 ng/ml was significantly higher than that at 1 ng/ml. This study provides the direct evidence in human that radiosensitivity may vary in relation to hormonal conditions.

Chromosome Aberrations↗

PubMed1999

Adaptive response in embryogenesis: II. Retardation of postnatal development of prenatally irradiated mice.

We previously reported that a priming dose of 0.3 Gy on gestation day 11 significantly increased the rate of living fetuses and reduced the incidence of congenital malformations caused by exposure to 5 Gy X rays on gestation day 12 in ICR mice. In the present study, postnatal development of the live offspring was investigated using a set of developmental and behavioral parameters. The offspring of the mice irradiated with 0.3 Gy generally showed a delay in the appearance of most of the physiological markers, impaired acquisition of neonatal reflexes, and alteration of adult behavior. However, an increase in body weight in the females was observed 4 weeks postnatally. In the offspring primed with 0.3 Gy followed by a challenging dose of 5 Gy prenatally, a high postnatal mortality was found, and all the survivors had various radiation-induced detrimental effects. The results indicated that the priming dose was advantageous to survival itself, but was disadvantageous to the health of survivor. The results also suggested that studying the whole animal can show the extent of the effects of radiation, i.e. quality of life, in a way that cellular or molecular studies cannot.

Abnormalities, Radiation-Induced↗

PubMed1999

Radiation-induced apoptosis and limb teratogenesis in embryonic mice.

In utero irradiation of the fetus during the period of organogenesis induces a dramatic increase in malformation. However, the mechanisms underlying the teratogenesis remain to be elucidated. In the present study, the correlation between radiation-induced apoptosis and limb malformation was examined in mice. The mice were exposed to X rays in utero on day 11 of gestation during the period of organogenesis of limb buds. A marked increase in the number of apoptotic cells in the predigital regions in the forelimb buds was detected 4 h after irradiation. The preinterdigital regions of the forelimb buds did not show such an increase at the same time. Aphlangy and ectrodactyly were the main types of anomalies observed on day 19 in the limbs of the fetuses irradiated with 5 Gy. The increases in prenatal death and teratogenesis in limb digits in living fetuses were dependent on dose. The possible mechanisms involved are discussed.

Abnormalities, Radiation-Induced↗

PubMed1998

Non-fluorescent chromosome painting using the peroxidase/diaminobenzidine (DAB) reaction.

PURPOSE: To develop and validate non-fluorescent chromosome painting for bright-field microscopy using the peroxidase/diaminobenzidine (DAB) reaction. MATERIALS AND METHODS: Peripheral blood lymphocytes were taken from patients with uterine cancer who had received heavy-ion radiation therapy. Chromosome slides were treated with RNase and pepsin, denatured mildly, hybridized with a biotinylated DNA probe specific for whole-chromosome 4 and stained using the peroxidase/DAB reaction with an avidin-biotin amplification. The slides were analysed under a bright-field microscope and an atomic force microscope. The detection rate of chromosome aberrations by DAB painting was compared with that obtained by dual analysis of Giemsa staining and FISH painting. RESULTS: When chromosomes 4 were painted, 11.5% of unstable aberrations were detected by DAB painting, while 10.8% of them were found by dual analysis of Giemsa staining and FISH painting. CONCLUSION: A DAB painting method that can effectively detect rearranged aberrations was established. It has advantages over FISH painting: the preparations can be analysed by bright-field microscope, can be preserved permanently and are suitable for analysis by an automated system.

3,3'-Diaminobenzidine↗

PubMed1998

Adaptive response in embryogenesis: I. Dose and timing of radiation for reduction of prenatal death and congenital malformation during the late period of organogenesis.

An adaptive response was demonstrated during embryogenesis in mice. Whole-body irradiation at a dose of 0-50 cGy was given to condition pregnant ICR mice on day 9 to day 11 of gestation. Then their whole bodies were exposed to a challenging dose of 5 Gy on the next day. The numbers of living fetuses, prenatal deaths and living fetuses with external gross malformations were determined on day 19. A conditioning dose of 30 cGy on day 11 significantly increased the rate of living fetuses and reduced the incidence of congenital malformations induced by a 5-Gy dose on day 12. This indicates the existence of a critical dose and timing for administering a conditioning dose for radioadaptation during the late period of organogenesis in mice. The possible mechanisms involved are discussed.

Abnormalities, Radiation-Induced↗

PubMed1996

Mapping of metastasis suppressor gene(s) for rat prostate cancer on the short arm of human chromosome 8 by irradiated microcell-mediated chromosome transfer.

Our previous studies demonstrated that human chromosome 8 contains metastasis suppressor gene(s) for rat prostate cancer. However, it is still unknown which portion of human chromosome 8 is associated with suppression of metastatic ability, because all of the clones in which metastatic ability is suppressed contain at least one copy of intact human chromosome 8. In the present study, we used the irradiated microcell-mediated chromosome transfer technique to enrich for specific chromosomal arm deletions of selected chromosomes. The resultant series of human chromosomes 8 with a variety of chromosomal deletions was introduced into highly metastatic Dunning rat prostate cancer cells. All of the resultant microcell hybrids showed reduced metastatic ability. To obtain a smaller size of human chromosome 8 and to locate further the region of metastasis suppressor gene(s), the most reduced size of human chromosome 8 that was generated with the initial irradiated chromosome transfer was retransferred into the Dunning cancer cells without irradiation. The resultant microcell hybrids were analyzed to determine which portion of human chromosome 8 suppressed the metastatic ability of the recipient cells. This analysis demonstrates that the portion of human chromosome 8 containing metastasis suppressor gene(s) for rat prostate cancer cells lies on human chromosome segment 8p21-p12, where frequent allelic losses have been detected in allelotype analyses of human prostate cancer. This suggests that one of the metastasis suppressor genes for rat prostate cancer on human chromosome 8 may also play an important role in the progression of human prostate cancer.

Animals↗

PubMed1996

Comparison of the yields of translocations and dicentrics measured using conventional Giemsa staining and chromosome painting.

Inconsistent results have been reported regarding the yields of translocations and dicentrics measured using chromosome painting. This inconsistency may be due to mis-scoring of dicentrics as translocations because painting is unsuited to identify centromere positions. In the present study, chromosome aberrations exclusively in the first mitosis after irradiation were analyzed using human peripheral lymphocytes. When identical metaphases were analyzed by both painting and conventional Giemsa staining, which is the most reliable method to locate centromeres, equal frequencies of translocations and dicentrics were observed.

Centromere↗

PubMed1994

Suppression of metastasis of rat prostatic cancer by introducing human chromosome 8.

In previous allelotype analyses of human prostatic cancer specimens, allelic loss on the short arm of chromosome 8 is frequently observed. However, it is still unclear whether this allelic loss is an initial event or a later one in development of prostatic cancer. Our previous studies demonstrate that introduction of human chromosome 11 into highly metastatic rat prostatic cancer cells results in suppression of metastatic ability without suppression of the in vivo growth rate or tumorigenicity of the hybrid cells (T. Ichikawa et al. Cancer Res., 52: 3486-3490, 1992). To clarify the role of human chromosome 8 in prostatic cancer, this chromosome was introduced into highly metastatic rat prostatic cancer cells using microcell-mediated chromosome transfer. Introduction of human chromosome 8 resulted in suppression of metastatic ability of the microcell hybrids, whereas no suppression of the in vivo growth rate or tumorigenicity was observed. These results demonstrate that human chromosome 8 contains metastasis suppressor gene(s) for prostatic cancer derived from a rat. These also suggest that human chromosome 8 has an important role in development of prostatic cancer.

Animals↗

PubMed1994

A chromosome painting method for human sperm chromosomes using fluorescent in situ hybridization.

A method of chromosome painting on human sperm chromosomes using fluorescent in situ hybridization (FISH) is introduced. Sperm chromosome slides were prepared after in vitro fertilization of hamster eggs with human spermatozoa. The slides were treated by RNase A before FISH. Chromosome 4 was clearly and specifically painted in a majority of sperm-derived metaphase plates after an application of whole chromosome painting DNA probes of this chromosome. This is the first report of successful painting on human sperm chromosomes.

Animals↗

PubMed1994

Effects of colcemid concentration on chromosome aberration analysis in human lymphocytes.

As a part of technical improvements of chromosome aberration analysis on human peripheral lymphocytes for biological radiation dosimetry, we examined the optimal conditions for the use of colcemid in chromosome preparation in order to obtain enough number of cells at metaphase in the first cell division. When treated with colcemid at concentrations below 0.01 microgram/ml from the beginning of culture, cultures harvested at 48 hours had low mitotic indices. Colcemid treatment at 0.025 to 0.05 microgram/ml during 48 hours resulted in high mitotic indices (8 to 15%) and almost of the mitotic cells remaining in the 1st cell division, suggesting that this range of colcemid concentration was appropriate for continuous treatment with colcemid. We further examined the effect of colcemid concentration on the quantitative consistency of the yields of radiation-induced chromosome aberration. Repeated experiments showed that the yield of dicentrics and centric rings in the culture having colcemid at 0.025 microgram/ml concentration were larger than that at 0.05 microgram/ml. These data indicate the importance of assuring the accuracy of colcemid concentration in the lymphocyte culture for cytogenetic radiation dosimetry.

Cells, Cultured↗

PubMed1993

Removal of stainable cytoplasmic substances from cytogenetic slide preparations.

A method to remove stainable cytoplasmic substances from cytogenetic preparation using RNase A treatment is reported. The preparations processed with this method are especially useful for the automated analysis of micronuclei of cultured cells with cytochalasin B and of chromosome aberrations induced by radiation.

Chromosome Aberrations↗

PubMed1992

A high resolution chromosome image processor for study purposes, NIRS-1000:CHROMO STUDY, and algorithm developing to classify radiation induced aberrations.

Since 1989 we have promoted a project to develop an automated scoring system of radiation induced chromosome aberrations. As a first step, a high resolution image processing system for study purposes, NIRS-1000:CHROMO STUDY, has been developed. It is composed of: (1) CHROMO MARKER whose main purpose is to mark on images to make image data base, (2) CHROMO ALGO whose purpose is algorithm development, and (3) METAPHASE RANKER whose purposes are metaphase finding and ranking with a high power objective lens. However, METAPHASE RANKER is presently under development. The system utilizes a high definition video system so as to realize the best spatial resolution that is achievable with an optical microscope using an objective lens (x 100, numerical aperture 1.4). The video camera has 1024 effective scan lines to realize 0.1 microns sampling on a specimen. The system resolution achieved on the hard copy is less than 0.3 microns on a specimen. A preliminary algorithm has been developed to classify the aberrations on the system using projection information of gray level. The preliminary test results on excellent 10 metaphases show that the correct classification ratio is 92.7%, that the detection rate of the aberrations is 83.3% and that the false positive rate is 6.1%.

Algorithms↗

PubMed1992

Robot system for preparing lymphocyte chromosome.

Towards the automatization of the scoring of chromosome aberrations in radiation dosimetry with the emphasis on the improvement of biological preparations, the conventional culture and harvesting method was modified. Based on this modified method, a culture and harvest robotic system (CHROSY) for system (CHROSY) for preparing lymphocyte chromosome was developed. The targeted points of the modification are as in the following. 1. Starting culture with purified lymphocytes in a fixed cell number. 2. Avoiding the loss of cells in changing the liquids following centrifugalization. 3. Keeping the quantity of the liquids to be applied to the treatments of cells fixed. 4. Building a system even a beginner can handle. System features are as follows. 1. Operation system: Handling robot having 5 degrees of freedom; a rotator incubator with an automatic sliding door; units for setting and removing pipette tips; a centrifuge equipped with a position adjuster and an automatic sliding door; two aluminum block baths; two nozzles as pipettes and aspirators connected to air pumps; a capping unit with a nozzle for CO2 gas; a compressor; and an air manipulated syringe. 2. Control system: NEC PC-9801RX21 with CRT; and program written in Basic and Assembly languages on MS-DOS. It took this system 2 hours and 25 minutes to harvest 2 cultures. A fairly good chromosome slide was made from the sample harvested by CHROSY automatically.

Chromosomes↗

PubMed1992

[Progress on chromosome image analysis systems].

This is a review of the development of automated systems of chromosome analysis. The history of its development reflects the progress of image analysis systems and that of method of chromosome analysis in man. It can be chronologically summarized as follows. Period I: 1960s-early 1970s when chromosome banding methods were discovered. Period II: early 1970s-early 1980s when the first image analysis system of chromosome was commercialized. Period III: early 1980s-present. Research and development of the automated analysis system of radiation induced chromosome aberrations, conducted under the national project of cross-over research on underlying technology of nuclear energy, Science and Technology Agency, Japan, is briefly reported.

Chromosomes↗

PubMed1990

Enhancement of an anti-tumor effect of interferon by dipyridamole in established human malignant melanoma cell lines.

Enhancement of the anti-proliferative effect of human interferon (HuIFN) preparations (alpha, beta and gamma) by dipyridamole was detected in a human malignant melanoma cell line, MM-ICB, which we originally established. Cell growth was inhibited by HuIFN alone, but a marked increase in inhibition was noted in vitro and in vivo when dipyrydamole was added. Cellular DNA synthesis, as determined by 3H-deoxythymidine incorporation into the acid-insoluble cellular fraction, was more inhibited by combined treatment than by any of the agents used alone. Two other melanoma cell lines that we established, MM-2CB and MM-3CB, also exhibited sensitivity to combined treatment both in vitro and in vivo. Furthermore, the HMV-I and SEKI melanoma cell lines were susceptible to the combination. Even non-cytotoxic concentrations of dipyridamole could enhance the effect of HuIFN on MM-ICB, MM-2CB, and SEKI cells.

Aged↗