Shin Asada

Publications (7)11.71 Total impact

• Article: In vitro clastogenicity and phototoxicity of fullerene (C(60)) nanomaterials in mammalian cells.

  Masamitsu Honma, Toshitaka Takahashi, Shin Asada, Yuzuki Nakagawa, Atsuko Ikeda, Kohji Yamakage
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  ABSTRACT: Carbon nanomaterials such as carbon nanotubes, graphene, and fullerenes (C(60)) are widely used in industry. Because of human health concerns, their toxic potential has been examined in vivo and in vitro. Here we used mammalian cells to examine the in vitro clastogenicity as well as the phototoxicity of C(60). While C(60) induced no structural chromosome aberrations in CHL/IU cells at up to 5mg/ml (the maximum concentration tested), it significantly induced polyploidy at 2.5 and 5mg/ml with and without metabolic activation. In BALB 3T3 cells, C(60) showed no phototoxic potential but the anatase form of titanium oxide did. Since insoluble nanomaterials cause polyploidy by blocking cytokinesis rather than by damaging DNA, we concluded that the polyploidy induced by C(60) in CHL/IU cells was probably due to non-DNA interacting mechanisms.
  Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 08/2012; · 2.85 Impact Factor
• Article: Application of the improved BALB/c 3T3 cell transformation assay to the examination of the initiating and promoting activities of chemicals: the second interlaboratory collaborative study by the non-genotoxic carcinogen study group of Japan.

  Toshiyuki Tsuchiya, Makoto Umeda, Noriho Tanaka, Ayako Sakai, Hiroshi Nishiyama, Isao Yoshimura, Syozo Ajimi, Shin Asada, Masumi Asakura, Hiroshi Baba, [......], Tetsu Nakamura, Hidetoshi Oishi, Toshiaki Sasaki, Sawako Shimada, Chitose Takahashi, Yuko Takeda, Sinobu Wakuri, Nobuhiro Yajima, Satoshi Yajima, Tomoko Yatsushiro
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  ABSTRACT: The Non-genotoxic Carcinogen Study Group in the Environmental Mutagen Society of Japan organised the second step of the inter-laboratory collaborative study on one-stage and two-stage cell transformation assays employing BALB/c 3T3 cells, with the objective of confirming whether the respective laboratories could independently produce results relevant to initiation or promotion. The method was modified to use a medium consisting of DMEM/F12 supplemented with 2% fetal bovine serum and a mixture of insulin, transferrin, ethanolamine and sodium selenite, at the stationary phase of cell growth. Seventeen laboratories collaborated in this study, and each chemical was tested by three to five laboratories. Comparison between the one-stage and two-stage assays revealed that the latter method would be beneficial in the screening of chemicals. In the test for initiating activity with the two-stage assay (post-treated with 0.1microg/ml 12-O-tetradecanoylphorbol-13-acetate), the relevant test laboratories all obtained positive results for benzo[a]pyrene and methylmethane sulphonate, and negative results for phenanthrene. Of those laboratories assigned phenacetin for the initiation phase, two returned positive results and two returned negative results, where the latter laboratories tested up to one dose lower than the maximum dose used by the former laboratories. In the exploration of promoting activity with the twostage assay (pretreated with 0.2microg/ml 3-methylcholanthrene), the relevant test laboratories obtained positive results for mezerein, sodium orthovanadate and TGF-beta1, and negative results for anthralin, phenacetin and phorbol. Two results returned for phorbol 12,13-didecanoate were positive, but one result was negative - again, the maximum dose to achieve the latter result was lower than that which produced the former results. These results suggest that this modified assay method is relevant, reproducible and transferable, provided that dosing issues, such as the determination of the maximum dose, are adequately considered. The application of this two-stage assay for screening the initiating and promoting potential of chemicals is recommended for consideration by other research groups and regulatory authorities.
  Alternatives to laboratory animals: ATLA 03/2010; 38(1):11-27. · 1.58 Impact Factor
• Article: Technical modification of the Balb/c 3T3 cell transformation assay: the use of serum-reduced medium to optimise the practicability of the protocol.

  Kumiko Hayashi, Kiyoshi Sasaki, Shin Asada, Toshiyuki Tsuchiya, Makoto Hayashi, Isao Yoshimura, Noriho Tanaka, Makoto Umeda
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  ABSTRACT: The two-stage Balb/c 3T3 model of cell transformation can mimic the two-stage carcinogenicity bioassay, and has been recognised as a screening method for detecting potential tumour initiators and promoters. A technical modification to the original protocol (which involved the use of M10F medium, consisting of MEM plus 10% fetal bovine serum [FBS]) has been previously proposed, in order to increase its efficacy, namely: the introduction of enriched, serum-reduced medium (DF2F medium, comprising DMEM/F12 plus 2% FBS and other supplements). The aim of this study was to further modify the protocol, so as to attain higher practicability for the assay. The protocol was further optimised by: a) reducing the number of plates required, through the use of larger plates; b) reducing the cost of the assay by retaining the reduced serum concentration and by using 2microg/ml insulin, rather than the more-complex insulin-transferrin-ethanolamine-sodium selenite (ITES) supplement (i.e. DF2F2I medium); and c) extending the culture period from 24-25 days to 31-32 days, resulting in clearer foci (the number of medium changes did not increase, as less-frequent medium changes were performed during the extended culture period). Growth curve construction revealed that variations in the saturation densities of the parental Balb/c 3T3 cell line and its three transformed clones were highest when M10F medium was replaced with DF2F2I medium just before cells reached confluence. We applied this newly-optimised protocol to the assessment of: a) the tumour initiating activity of 3-methylcholanthrene (MCA), N-methyl-N'-nitro-N-nitrosoguanidine, mitomycin C, methylmethane sulphonate, CdCl(2) and phenacetin, combining a post-treatment of 100ng/ml 12-O-tetradecanoylphorbol-13-acetate at the promotion stage; and b) the tumour promoting activity of insulin, lithocholic acid, CdCl(2) and phenobarbital, with pre-treatment of 0.2microg/ml MCA at the initiation stage. In the present study, only phenobarbital was negative when tested by using the modified protocol.
  Alternatives to laboratory animals: ATLA 01/2009; 36(6):653-65. · 1.58 Impact Factor
• Article: Activity related to the carcinogenicity of plastic additives in the benzophenone group.

  Daisuke Nakajima, Shin Asada, Shiho Kageyama, Takashi Yamamoto, Hidetoshi Kuramochi, Noriho Tanaka, Ken Takeda, Sumio Goto
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  ABSTRACT: This study examines the activities relating to the carcinogenicity of six types of benzophenone derivatives (benzophenone, 2-hydroxy-4-octyloxybenzophenone, 2-hydroxy-4-methoxybenzophenone, 2,4-dihydroxybenzophenone, 2,2'-dihydroxy-4-methoxybenzophenone and 2,2'-dihydroxy-4,4'-dimethoxybenzophenone) currently used in plastic products as additives to serve as ultraviolet absorbing agents. The evaluation of the initiation activity used a light absorption umu-test, a luminescent umu-test and the Ames test. The promotion activity was examined by a Bhas assay, a method that uses Bhas 42 cells for the formation of transformation foci. The luminescent umu-test indicated positive initiation activity of 2-hydroxy-4-methoxybenzophenone, and pseudo-positive activity of 2,4-dihydroxybenzophenone and 2,2'-dihydroxy-4-methoxybenzophenone. In the Ames test, 2-hydroxy-4-octyloxybenzophenone showed pseudo-positive initiation activity. Conversely, 2,4-dihydroxybenzophenone indicated weak promotion activity at 10 microg/ml concentration.
  Journal of UOEH 07/2006; 28(2):143-56.
• Article: Detection of initiating as well as promoting activity of chemicals by a novel cell transformation assay using v-Ha-ras-transfected BALB/c 3T3 cells (Bhas 42 cells).

  Shin Asada, Kiyoshi Sasaki, Noriho Tanaka, Ken Takeda, Makoto Hayashi, Makoto Umeda
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  ABSTRACT: Cell transformation assay using BALB/c 3T3 cells, C3H10T1/2 cells and others, can simulate the two-stage carcinogenesis utilized for formation of transformed foci. A sensitive cell transformation assay for tumor initiators as well as promoters has been developed using a v-Ha-ras-transfected BALB/c 3T3 cell line, Bhas 42; these cells are regarded as initiated in the two-stage paradigm of carcinogenesis. To distinguish between initiation and promotion, the initiation assay involves a 2-day treatment of low-density cells, obtained one day after plating, with a test chemical, and the promotion assay involves treatment of near-confluent cells with a test chemical for a period of 12 days (Day 3-14). When Bhas 42 cells were treated with tumor initiators, N-methyl-N'-nitro-N-nitrosoguanidine and 3-methylcholanthrene, transformed foci were induced in the initiation assay but not in the promotion assay. In contrast, tumor promoters, 12-O-tetradecanoylphorbol-13-acetate, lithocholic acid and okadaic acid, gave negative responses in the initiation assay but positive responses in the promotion assay. The results were reproducible with various treatment protocols. Sixteen polycyclic aromatic hydrocarbons were examined using both assays. Benzo[a]pyrene and 7,12-dimethylbenz[a]anthracene induced focus formation only in the initiation assay. Increase of focus formation was observed in the promotion assay with benzo[e]pyrene, benzo[ghi]perylene, 1-nitropyrene and pyrene. Benz[a]anthracene, benz[b]anthracene, chrysene and perylene showed positive responses in both initiation and promotion assays. Results of initiation and promotion assays of acenaphthylene, anthracene, coronene, 9,10-diphenylanthracene, naphthalene and phenanthrene were negative or equivocal. The present Bhas assays for the detection of either/both initiating and promoting activities of chemicals are sensitive and of high performance compared with other cell transformation assays.
  Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 01/2006; 588(1):7-21. · 2.85 Impact Factor
• Article: Tumor-promoting activity and mutagenicity of 5 termiticide compounds.

  Sumio Goto, Shin Asada, Yuichi Fushiwaki, Yasuaki Mori, Noriho Tanaka, Makoto Umeda, Daisuke Nakajima, Ken Takeda
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  ABSTRACT: The tumor-promoting activities of 5 commercial compounds used in termiticides were measured by a cell-transformation assay employing Bhas 42 cells. Their initiating activities were also measured by the microsuspension assay employing S. typhimurium TA98 and TA100 strains. The results of the transformation assay confirmed the tumor-promoting activities of fenitrothion, silafluofen and bifenthrin. Furthermore, the mutagenicity of S-421 and fenitrothion were also confirmed. Consideration of 2-stage carcinogenesis suggests that concurrent use of and long-term exposure to these compounds that have tumor-promoting and initiator activity, and compounds exhibiting either type of activity individually should be avoided as much as possible.
  Journal of UOEH 01/2005; 26(4):423-30.
• Article: An assay method for the prediction of tumor promoting potential of chemicals by the use of Bhas 42 cells.

  Kiyomi Ohmori, Kiyoshi Sasaki, Shin Asada, Noriho Tanaka, Makoto Umeda
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  ABSTRACT: It has become an important task to develop a simple in vitro method for the detection of non-genotoxic carcinogens, among which tumor promoters are included. Bhas 42 cells are v-Ha-ras-transfected BALB/c 3T3 cells and are regarded as initiated cells in the 2-stage transformation paradigm. We designed a method for detecting tumor promoters by the use of Bhas 42 cells at advanced passage generation. In this method, the cells are cultured in six-well plates for 17 days during which test chemicals are added in the medium for 11 days from days 3 to 14. The end-point of the assay is the induction of transformed foci. When the tumor promoter TPA was used, a significant number of transformed foci were induced concentration-dependently, whereas only a few foci were observed in control cultures. When various chemicals were examined by the method, a reasonable correlation was observed with the reported tumor-promoting ability in animal experiments. We propose that the Bhas 42 cell transformation method is practical and useful for the detection of tumor promoters.
  Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 03/2004; 557(2):191-202. · 2.85 Impact Factor