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J Miyakoshi

Publications and source records attributed to J Miyakoshi.

At least 19 recordsLinked to original sources
PubMed2001

Transient suppression of X-ray-induced apoptosis by exposure to power frequency magnetic fields in MCF-7 cells.

Epidemiological studies suggest that exposure to power frequency magnetic fields may be a risk factor for breast cancer in humans. To study the relationship between exposure to 60-Hz magnetic fields (MFs) and breast cancer, cell cycle distribution, apoptosis, and the expression of related proteins (p21, Bax, and Bcl-2) were determined in MCF-7 cells following exposure to magnetic fields (60 Hz, 5 mT) alone or in combination with X rays. It was found that exposure of MCF-7 cells to 60-Hz MFs for 4, 8, and 24 h had no effect on cell cycle distribution. Furthermore, 60-Hz MFs failed to affect cell growth arrest and p21 expression induced by X rays (4 Gy). Similarly, 60-Hz MFs did not induce apoptosis or the expression of Bax and Bcl-2, two proteins related to apoptosis. However, exposure of cells to 60-Hz MFs for 24 h after irradiation by X rays (12 Gy) significantly decreased apoptosis and Bax expression but increased Bcl-2 expression. The effects of exposure to 60-Hz MFs on X-ray-induced apoptosis and Bax and Bcl-2 expressions were not observed at 72 h. These data suggest that exposure to 60-Hz MFs has no effects on the growth of MCF-7 cells, but it might transiently suppress X-ray-induced apoptosis through increasing the Bcl-2/Bax ratio.

Apoptosis↗
PubMed2001

Involvement of eddy currents in the mutagenicity of ELF magnetic fields.

Possible carcinogenic and/or mutagenic activity of extremely low frequency magnetic fields was examined using somatic mutation and recombination test system of Drosophila melanogaster. An X-linked semi-dominant DNA repair defective mutation mei-41(D5) was introduced into the conventional mwh/flr test system to enhance mutant spot frequency. Virgin females of w mei-41(D5)/FM6; flr/TM6 were crossed with w mei-41(D5)/Y; mwh jv; spa(pol) males. The F(1) third instar larvae were exposed to a 50Hz, 20mT sinusoidal AC magnetic field for 24h. After moulting from pupal cases, their wings were examined under a bright field microscope to detect hair spots with mwh or flr mutant morphology. The exposure caused a statistically significant enhancement in somatic recombination spot frequency. Mutant spots arising due to chromosomal non-disjunction or terminal deletion also increased but the frequency of spots resulting from point mutation was not altered. The enhancement in the recombination spot frequency was suppressed to the control level when a culture medium without electrolytes was used during exposure. When larvae were exposed to a magnetic field in an annular dish, flies from the outer ring showed more mutant spots compared to those from the inner ring. These results suggest that the detected mutagenic activity was that of the induced eddy current, rather than that of the magnetic field itself.

Animals↗
PubMed2001

Increase in hypoxanthine-guanine phosphoribosyl transferase gene mutations by exposure to electric field.

Previously, we reported that exposure to extremely low frequency magnetic field (400 mT) increased in hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene mutations. However, it is unclear these mutations were induced by magnetic field (MF), electric field (EF), or both. To explore this question, a new exposure apparatus for EF was manufactured. We observed an increase in HPRT gene mutations in Chinese hamster ovary (CHO) cells after exposure to EF (10 V/m, 60 Hz) for 10 h. The mutant frequency by EF-exposure was an approximate 2-fold of that by sham-exposure. Our data suggest that the mutations induced by exposure of cells to the variable magnetic field at 400 mT may be, in part, due to the induced EF.

Animals↗
PubMed2001

Development of a cryopreservation procedure employing a freezer bag for pancreatic islets using a newly developed cryoprotectant.

One of the most important requirements for success in clinical islet transplantation is the use of a large number of viable donor islets. To achieve this, the ability to cryopreserve islets and to establish an islet bank are critical. Previously, we developed a two-step cryopreservation procedure with freezing tubes utilizing low and high concentrations of dimethyl sulfoxide (DMSO) and using a fully automated cryomachine for human pancreatic islets and porcine islet-like cell clusters (ICCs). Based on these experiments, we developed a simple and efficient cryopreservation procedure of a freezer bag for isolated islets using a fully automated computer-controlled cryomachine with a newly developed cryoprotectant consisting of ethylene glycol (EG) instead of DMSO for decreasing injury of the islets by freezing. A 250 ml Cryocyte blood freezer bag and our newly developed cryoprotectant containing ethylene glycol (EG) were used in the freezing procedure. The islets were frozen by a fully automated computer-controlled cryomachine (GE 9,000) with our original program of slow cooling. Nucleation occurred at -8 degrees C, and the frozen islets were stored at -196 degrees C in a liquid nitrogen tank. The frozen-stored islets were subsequently rapidly thawed in a 37 degrees C water bath and cultured before viability testing. In vitro function, the stimulation index of insulin release during the static incubation test for rat islets cryopreserved in a freezer bag vs. nonfrozen islets as control, was 2.13 +/- 0.42 and 2.02 +/- 0.38 (94.8% compared with control), respectively (n = 5, p = NS). The islet recovery compared with the nonfrozen control group was 85% (n = 5) in insulin content. When 1000 rat islets cryopreserved in a freezer bag were transplanted into the renal capsule of diabetic athymic mice, all the mice became normoglycemic within 7 days from transplantation. Before nephrectomy, the intravenous glucose torelance test (IVGTT) was performed. The fractional decay constant of the glucose level (K value) of the frozen-thawed group was 0.42 +/- 0.06%/min. A histological study of renal subcapsular grafts demonstrated the morphological integrity of the islets. These results demonstrate the utility of our cryopreservation procedure of a freezer bag for isolated islets using a fully automated computer-controlled cryomachine with a newly developed cryoprotectant for the maintenance of viability and function of frozen-stored islets both in culture and after transplantation. Cryopreservation using freezer bags with the new cryoprotectant is an effective and simple method for making an islet bank for clinical trials of islet transplantation.

Animals↗
PubMed2000

Increase in X-ray-induced mutations by exposure to magnetic field (60 Hz, 5 mT) in NF-kappaB-inhibited cells.

It is established that extremely low frequency magnetic fields (ELFMF) at the flux densities, i.e., 5 mT and less, are not mutagenic. However, exposure to ELFMF enhances mutations induced by X-rays. In this study, we examined the effects of long-term exposure to 5 mT ELFMF on mutation induction and X-ray-induced mutations in human malignant glioma cells (MO54) with different mutant IkappaB-alpha (a critical inhibitor of NF-kappaB) genes. Cells were exposed or sham-exposed to 5 mT ELFMF for up to 8 days with or without initial X-rays (4 Gy), and the mutant frequency of hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene was analyzed. An obvious increase in X-ray-induced mutations was observed after treatment with ELFMF in combination with X-irradiation in MO54 cells with tyrosine mutant IkappaB-alpha gene other than with serine mutant IkappaB-alpha gene or vector alone. Exposure to ELFMF alone increased mutations significantly in MO54 cells with tyrosine mutant IkappaB-alpha gene. In addition, X-ray-induced apoptoic cells were increased in MO54-V cells after exposure to ELFMF, while an anti-apoptotic effect of magnetic field was found in MO54-SY4 cells. Our data suggest that exposure to 5 mT ELFMF may induce mutations and enhance X-ray-induced mutations, resulting from the inactivation of NF-kappaB through the inhibition of tyrosine phosphorylation.

Electromagnetic Fields↗
PubMed2000

Exposure to extremely low frequency magnetic fields suppresses x-ray-induced transformation in mouse C3H10T1/2 cells.

We designed and manufactured equipment for exposure of cultured cells to extremely low frequency magnetic fields (ELFMF) at 5, 50, and 400 mT and examined the effect of ELFMF on cellular transformation in mouse C3H10T1/2 cells (clone 8). Transformed foci, Type II and Type III, were independently counted as transformants. The cells were exposed to ELFMF alone at 5, 50, and 400 mT for 24 h or X-irradiated with 3 Gy followed by the ELFMF exposure. No significant difference in the transformation was observed between sham-exposed control and the ELFMF exposure from 5 to 400 mT. The transformation frequency for X-rays plus ELFMF was decreasing compared with X-rays alone. When 12-O-tetra-decanoylphorbol-13-acetate (TPA) was contained in the medium throughout the experiment, the transformation frequency by X-rays alone was elevated more. In the combined treatment with X-rays followed by ELFMF, the transformation frequency was slightly decreased at 50 and 400 mT even in the medium containing TPA. The long-term exposure at 5 mT suppressed both spontaneous and X-ray-induced transformations significantly. It is well known that overexpressing protein kinase C (PKC) failed to yield identifiable transformation of foci induced by ionizing radiation. We demonstrated previously that exposure to high-density ELFMF induced expression of several genes through an increase in PKC activity. From these results, it is suggested that ELFMF might suppress X-ray-induced transformation through activation of PKC by ELFMF.

Animals↗
PubMed2000

Suppression of heat-induced HSP-70 by simultaneous exposure to 50 mT magnetic field.

Effect of extremely low frequency magnetic field (ELFMF) at 50 mT and 60 Hz on heat-induced expression of heat shock protein 70 (hsp-70) was examined in HL60RG cells. No increase in hsp-70 production was observed in the cells after exposure to 50 mT ELFMF alone. Simultaneous exposure to 50 mT ELFMF in combination with mild heat at 42 and 40 degrees C suppressed heat-induced hsp-70 expression. The suppression of hsp-70 occurred when cells were simultaneously exposed to both for longer periods of more than 5 h. However, the suppression of hsp-70 was not observed at a magnetic density of 5 and 0.5 mT. This result suggests that exposure to 50 mT ELFMF may act on a protection against the concomitant mild heat stress in HL60RG cells.

Blotting, Western↗
PubMed2000

Inhibition of I kappaB-alpha phosphorylation at serine and tyrosine acts independently on sensitization to DNA damaging agents in human glioma cells.

Molecular mechanisms and/or intrinsic factors controlling cellular radiosensitivity are not fully understood in mammalian cells. The recent studies have suggested that nuclear factor kappaB (NF-kappaB) is one of such factors. The activation and regulation of NF-kappaB are tightly controlled by IkappaB-alpha, a cellular inhibitory protein of NF-kappaB. Most importantly, phosphorylation regulates activity of the inhibitor IkappaB-alpha, which sequesters NF-kappaB in the cytosol. Two different pathways for the phosphorylation of IkappaB-alpha are demonstrated, such as serine (at residues 32 and 36) and tyrosine (at residue 42) phosphorylations. To assess a role of the transcription factor, NF-kappaB, on cellular sensitivity to DNA damaging agents, we constructed three different types of expression plasmids, i.e. S-IkappaB (mutations at residues 32 and 36), Y-IkappaB (mutation at residue 42) and SY-IkappaB (mutations at residues 32, 36 and 42). The cell clones expressing S-IkappaB and Y-IkappaB proteins became sensitive to X-rays as compared with the parental and vector-transfected cells. The cell clones expressing SY-IkappaB were further radiosensitive. By the treatment with herbimycin A, an inhibitor of phosphorylation, the X-ray sensitivity of cells expressing SY-IkappaB did not change, while that of the cells expressing S-IkappaB and Y-IkappaB and the parental cells was enhanced. Change in the sensitivity to adriamycin and UV in those clones was very similar to that in the X-ray sensitivity. The inhibition of IkappaB-alpha phosphorylation at serine and tyrosine acts independently on the sensitization to X-rays, adriamycin and UV. These findings suggest that the transcriptional activation induced by NF-kappaB may play a role in the DNA damage repair. The present study proposes a possibility that the inactivation of NF-kappaB by inhibition of both serine and tyrosine phosphorylations may be useful for the treatment of cancer in radio- and chemotherapies.

Antineoplastic Agents↗
PubMed2000

Increased chromatid-type chromosomal aberrations in mouse m5S cells exposed to power-line frequency magnetic fields.

PURPOSE: To investigate the induction of chromosomal aberrations in mouse m5S cells after exposure to power-line frequency magnetic fields (extremely low frequency magnetic fields; ELFMF) at high-flux densities. MATERIAL AND METHOD: m5S cells were either untreated or pretreated during the G1 phase with mitomycin C (MMC, 1 microM) for 1 h or 3 Gy X-rays, and then exposed to ELFMF at three different flux densities (5 and 50 mT at 60 Hz, 400 mT at 50 Hz) for 40 h. Unexposed control cells were incubated for the same period in a conventional CO2 incubator. Chromosomal aberrations were analysed in the first post-treatment metaphases. Cell kinetics were assessed by DNA flow cytometry and the mitotic index. RESULTS AND CONCLUSIONS: ELFMF enhanced the formation of spontaneous and MMC- or X-ray-induced chromosomal aberrations, in a flux-density-dependent manner. Statistically significant increases in the frequency of chromosomal aberrations were observed in cells exposed to 400 mT ELFMF with respect to unexposed controls. The aberrations induced by ELFMF were mostly chromatid-type, not chromosome-type. The cells exposed to 400 mT ELFMF exhibited a three-fold higher level of chromatid-type aberrations than did the unexposed cells. Flow cytometric and mitotic index analyses revealed that the S or G2 arrest following MMC or X-irradiation was more profound in ELFMF-exposed cells than in unexposed cells. Our results suggest that ELFMF can interfere with post-replication repair, resulting in increased levels of chromatid-type chromosomal aberrations induced spontaneously and by DNA damaging agents.

Animals↗
PubMed2000

Exposure to strong magnetic fields at power frequency potentiates X-ray-induced DNA strand breaks.

We examined the effect of an extremely low-frequency magnetic field (ELFMF) at 5, 50 and 400 mT on DNA strand breaks in human glioma MO54 cells. A DNA damage analysis was performed using the method of alkaline comet assay. The cells were exposed to X-rays alone (5 Gy), ELFMF alone, or X-rays followed by ELFMF at 4 degrees C or on ice. No significant difference in the tail moment was observed between control and ELFMF exposures up to 400 mT. X-ray irradiation increased DNA strand breaks. When cells were exposed to X-rays followed by ELFMF at 50 and 400 mT, the tail moment increased significantly compared with that for X-rays alone. When the exposure of cells was performed at 37 degrees C, no significant change was observed between X-rays alone and X-rays plus 400 mT. We previously observed that exposure to 400 mT ELFMF for 2 h increased X-ray-induced mutations (Miyakoshi et al, Mutat. Res., 349: 109-114, 1996). Additionally, an increase in the mutation by exposure to the ELFMF was observed in cells during DNA-synthesizing phase (Miyakoshi et al., Int. J. Radiat. Biol., 71: 75-79, 1997). From these results, it appears that exposure to the high density ELFMF at more than 50 mT may potentiate X-ray-induced DNA strand breaks.

Cell Death↗
PubMed1999

Effect of high-density extremely low frequency magnetic field on sister chromatid exchanges in mouse m5S cells.

The induction of sister chromatid exchanges (SCEs) was evaluated in the cultured mouse m5S cells after exposure to extremely low frequency magnetic field (ELFMF; 5, 50 and 400 mT). Exposure to 5 mT and 50 mT ELFMF led to a very small increase in the frequency of SCEs, but no significant difference was observed between exposed and unexposed control cells. The cells exposed to 400 mT ELFMF exhibited a significant elevation of the SCE frequencies. There was no significant difference between data from treatments with mitomycin-C (MMC) alone and from combined treatments of MMC plus ELFMF (400 mT) at any MMC concentrations from 4 to 40 nM. These results suggest that exposure to highest-density ELFMF of 400 mT may induce DNA damage, resulting in an elevation of the SCE frequencies. We suppose that there may be a threshold for the elevation of the SCE frequencies, that is at least over the magnetic density of 50 mT.

Animals↗
PubMed1999

Long-term exposure to a magnetic field (5 mT at 60 Hz) increases X-ray-induced mutations.

Exposure to extremely low frequency magnetic field (ELFMF) at 400 mT has been shown to induce mutations (Mutat. Res., 349: 109-114, 1996; Int. J. Radiat. Biol., 71: 75-79, 1997; and Biochem. Biophys. Res. Commun., 243: 579-584, 1998). However, whether ELFMF at low flux densities (under 1 mT) induces mutations is debatable. We investigated the effect of long-term exposure to 5 mT ELFMF at 60 Hz on mutant frequency. Chinese hamster ovary K1 (CHO-K1) cells were exposed or sham-exposed to 5 mT ELFMF for up to 6 weeks with or without X-irradiation (3 Gy), and the mutant frequency of the hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene was analyzed. Long-term exposure to 5 mT ELFMF did not increase mutations, suggesting a threshold for mutation induction greater than 115 mA/m2 or a magnetic density of 5 mT. However, enhancement of the X-ray-induced mutation rate was observed after treatment with X-irradiation followed by long-term exposure to 5 mT ELFMF. At little as a 1-week exposure to ELFMF after X-irradiation enhanced the mutation rate. We also found that 400 mT exposure enhanced the mutation rate induced by X-irradiation (Mutat. Res., 349: 109-114, 1996). These results suggest that exposure to more than 5 mT ELFMF may promote X-ray-induced mutations.

Animals↗
PubMed1999

Decreased host-cell reactivation of UV-irradiated adenovirus in human colon tumor cell lines that have normal post-UV survival.

An ongoing study in our laboratories is to examine the relationship of DNA repair defects to human cancer. Our underlying hypothesis has been that human tumors may arise that lack interesting DNA repair pathways if these pathways are important in preventing cancer. In this study, we found that the UV-irradiated adenoviruses showed hypersensitivity when assayed on monolayers of certain human colon tumor cell lines, including three that are reported to have defects in long patch DNA mismatch repair genes and one with no reported defect in mismatch repair. The survival curves showed two components. The first sensitive component was characteristic of 77-95% of the infections depending upon the cell line and the experiment and had an average slope indicating 4.8-fold hypersensitivity to UV. The average of the second-component slopes indicated that the remainder of the infections was accompanied by near-normal repair. Although the value of the first component indicated that the colon tumor lines supported the growth of UV-damaged adenoviruses poorly, the cell lines themselves showed the same post-UV colony-forming ability as did normal human fibroblasts, and their ability to support the growth of N-methyl-N'-nitro-N-nitrosoguanidine-damaged adenoviruses was normal, i.e. it parallelled their ability to repair O6-methylguanine in vitro. We previously observed two-component survival curves when assaying UV-irradiated adenovirus on monolayers of all of seven strains of fibroblasts from Cockayne's syndrome patients. By contrast, single-component curves have been obtained using 21 strains of normal human fibroblasts and seven other tumor lines. We interpret the two-component survival curves in terms of the defective transcription-coupled repair of UV-induced DNA damage that is characteristic both of Cockayne's and certain colon tumor cell lines. In addition, four mismatch repair-deficient colon tumor lines were resistant to killing by elevated levels of dG.

Adenoviruses, Human↗
PubMed1998

Suppression of high-density magnetic field (400 mT at 50 Hz)-induced mutations by wild-type p53 expression in human osteosarcoma cells.

Exposure of cultured human osteosarcoma cells (Saos-LP-12) to high-density (400 mT at 50 Hz) extremely low frequency magnetic fields (ELFMF) induced mutations in the hypoxanthine-guanine phosphoribosyl transferase gene. Saos-LP-12 cells, which are isolated from parental Saos-2 cells and have a deletion in the coding region of the p53 gene, are introduced to the wild-type (wt) p53 expression plasmid (pOPRSVp53). The mutation in Saos-LP-12 cells was suppressed by expression of the introduced wt p53 gene during 400 mT ELFMF exposure. No marked difference in the mutation spectrum was observed among the treatments of ELFMF [p53 (-)], ELFMF [p53 (+)], and sham exposures. Our findings suggest that wt p53 has a function in suppression of DNA replication errors and/or in maintenance of genomic stability after high-density ELFMF exposure.

DNA Mutational Analysis↗
PubMed1998

Reduced UV-induced mutations in human osteosarcoma cells stably expressing transfected wild-type p53 cDNA.

We constructed the plasmid which can express human wild-type p53 cDNA and introduced it into the human osteosarcoma cell line SAOS-2 that lacks the chromosomal p53 gene. A cell clone stably expressing p53 protein was isolated and UV sensitivity and UV-induced mutation frequencies of the clone were examined. The UV sensitivity of the clone was slightly higher and UV-induced hprt mutation frequencies of the clone were markedly lower than those of parental SAOS-2 cells. The capability to repair UV-induced DNA damage assessed by the amount of unscheduled DNA synthesis or DNA single strand breaks as well as cell cycle progression after UV irradiation were not different between the clone and SAOS-2 cells. These results indicate that wild-type p53 protein would be involved in the human DNA damage-processing pathway other than the genome-overall excision repair.

Cell Survival↗
PubMed1998

Enhanced NOR-1 gene expression by exposure of Chinese hamster cells to high-density 50 Hz magnetic fields.

Enhanced expression of neuron derived orphan receptor (NOR-1) gene was observed by exposure of Chinese hamster ovary K1 (CHO-K1) cells to an extremely low frequency magnetic field (ELFMF) of 50 Hz at 400 mT, but not at 5 mT. The enhanced expression, reaching the maximum at 6 h, was transient and reduced to the control level after exposure to 400 mT ELFMF for 24 h. The NOR-1 expression induced by treatment with forskolin and TPA was further enhanced by the simultaneous treatment with 400 mT ELFMF, in which the maximum response was at 3 h. The NOR-1 expression by these treatments was induced more earlier than that by 400 mT ELFMF alone. When cells were treated with an inhibitor of the protein kinase C (calphostin C or crocetin) and Ca2+ entry blockers (nifedipin and dantrolen) during the 400 mT ELFMF exposure, the enhanced NOR-1 expression was not observed. Exposure of CHO-K1 cells to the high-density 400 mT ELFMF may affect the signal transduction in the cells, resulting in the enhanced NOR-1 gene expression.

Animals↗
PubMed1997

Increase in radiation sensitivity of human malignant melanoma cells by expression of wild-type p16 gene.

The influence of wild-type p16 expression on the radiation sensitivity of human melanoma cell lines was investigated. MeWo cells, which alone expressed intrinsic wild-type p16 among six melanoma cell lines examined, showed higher radiosensitivity in comparison with the other five melanoma cells. The introduction of human wild-type p16 cDNA into A875 cells, which homozygously lost p16 genes, and AKI cells, which retained p16 gene but did not express p16 mRNA, led to increased sensitivity of those cells to X-ray irradiation. The radiosensitizing effect by the p16 introduction to those cells was prominent after rather higher doses of X-rays (8 and 10 Gy). In both A875 and AKI, no significant difference in sensitivities to UVC and cisplatin was observed between the parental and p16-transfectant cells. These results suggest that the loss or dysfunction of p16 gives melanoma cells the radioresistant characteristics.

Chromosomes, Human, Pair 9↗