Development of important genes during breast carcinogenesis
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1 Nagoya Med. J., 107 Development of important genes during breast carcinogenesis AYA NAIKI-ITO Department of experimental pathology and tumor biology Nagoya City University Graduate School of Medical Sciences Carcinogen exposure induces many gene expression changes in either target or non-target organ, some of them may be important for carcinogenic process but others not. In this study, we attempt to find crucial genes which may be important for mammary carcinogenesis. For this purpose, we used human c-ha-ras proto-oncogene transgenic rats Hraswhich are highly sensitive for mammary carcinogens, N-methyl-N-nitrosourea MNU,, -dimethyl benz a anthracene DMBA and -amino- methyl phenylimidazo-b pyridine PhIP. We found Gpx as an up-regulated gene commonly in all mammary carcinomas induced the different three carcinogens by DNA microarray analysis, and its up-regulation was confirmed by quantitative RT-PCR. In addition, immunohistochemically highly expression of GPX was detected in not only rat mammary carcinomas but also human breast cancers. Knockdown of GPX expression by sirna resulted in significant growth inhibition in both rat and human mammary carcinoma cell lines by p dependent manner. Thus, those data clearly demonstrated that GPX is involved in mammary carcinogenesis and cell proliferation in both rat and human, indicating that GPX may be a novel target of prevention and therapy for breast cancers. HER c-ha-ras 107
![108 Hras Hras N-methyl-N-nitrosourea MNU dimethylbenz a - anthracene DMBA amino- -methyl- -phenylimidazo-b]pyridine PhIP Hras RNA DNA CodeLink rat whole genome Hras Glutathione peroxidase Gpx Hras](/docs-images/94/118324784/images/2-0.jpg "MNU DMBA PhIP RNA Gpx mrna RT- PCR Figure Hras - DMBA RMC- Gpx mrna Table Western blotting Gpx Gpx Figure MCF- T D MDA-MB- GPX mrna Table Figure.")
2 108 Hras Hras N-methyl-N-nitrosourea MNU dimethylbenz a - anthracene DMBA amino- -methyl- -phenylimidazo-b]pyridine PhIP Hras RNA DNA CodeLink rat whole genome Hras Glutathione peroxidase Gpx Hras MNU DMBA PhIP RNA Gpx mrna RT- PCR Figure Hras - DMBA RMC- Gpx mrna Table Western blotting Gpx Gpx Figure MCF- T D MDA-MB- GPX mrna Table Figure. Relative Gpx expression levels in carcinogeninduced mammary cancers. The data were obtained by quantitative RT-PCR as described in the materials and methods. Gpx mrna expression level was adjusted by cytokeratin.*, p,**, p compared with control value Student s t test. Table. GPX mrna level of rat and human mammary carcinoma cell lines.
3 109 Figure. A. Western blot of normal mammary tissue and carcinoma in Hras rats. Lane to, normal mammary tissue. Lane to, carcinogen-induced mammary cancer; to, MNU, to, DMBA,and to, PhIP-induced. C. Immunohistochemical findings of Gpx in rat mammary carcinomas induced by MNU, DMBA and PhIP. Gpx was strongly positive in the cytoplasm of tumor cells. Non-tumorous mammary glands of Hras rats were completely negative. GPX GPX GPX Figure GPX GPX sirna Figure. Histological appearance of human breast cancers and GPX immunohistochemical staining. A, C, E : H&E,and B, D, F : immunohistochemistry. GPX staining was positive in the cytoplasm of human breast cancer cells regardless of tumor type. A to F were invasive ductal carcinoma; A and B were papillotubular carcinoma, C and D were solid-tubular carcinoma and E and F were scirrhous carcinoma. GPX GPX p p p p GPX p GPX Figure DNA
4 110 GPX GPX GPX p in vitro GPX GPX p Figure. GPX affects p -dependent cell proliferation in both rat and human mammary cancer cells. A. Rat mammary cancer cell lines, C with wild-type p showed clear inhibition of cell proliferation by Gpx silencing but not in the C with mutant p. *: Student s t test, p, **: p. Lower graphs show knockdown efficiency by quantitative RT-PCR. B. Human mammary carcinoma cell line, MCF- with wild-type p, showed significant inhibition of cell proliferation by GPX suppression, while another human breast cancer cell line, T D with mutant p showed no apparent change **: Student s t test, p. Lower graphs show knockdown efficiency by quantitative RT-PCR. Gpx GPX mrna GPX MCF- T D MDA-MB- GPX mrna Dumitrescu RG, Cotarla I: Understanding breast cancer risk wheredowestandin? JCell Mol Med : -, Asamoto M, Ochiya T, Toriyama-Baba H, et al.: Transgenic rats carrying human c-ha-ras protooncogenes are highly susceptible to N-methyl-Nnitrosourea mammary carcinogenesis Carcinogenesis : -, Naito A, Suzuki A, Ueda S, et al.: Preferential mammary carcinogenic effects of -amino- methyl- -phenylimidazo, -b pyridine PhIP in human c-ha-ras proto-oncogene transgenic rats Cancer Sci : -, Tsuda H, Asamoto M, Ochiya T, et al.: Highsusceptibility of transgenic rats carrying the human c- Ha-ras proto-oncogene to chemically-induced mammary carcinogenesis Mutat Res : -, YanW,ChenX:GPX, a direct target of p, inhibits oxidative stress-induced apoptosis in a p dependent manner J Biol Chem : -,
5 111 Chu FF, Esworthy RS, Doroshow JH: Role of Sedependent glutathione peroxidases in gastrointestinal inflammation and cancer Free Radic Biol Med :-, Chu FF, Doroshow JH, Esworthy RS: Expression, characterization, and tissue distribution of a new cellular selenium-dependent glutathione peroxidase, GSHPx-GI J Biol Chem : -, Banning A, Kipp A, Schmitmeier S, et al.: Glutathione Peroxidase Inhibits Cyclooxygenase- - Mediated Migration and Invasion of HT- Adenocarcinoma Cells but Supports Their Growth as Tumors in Nude Mice Cancer Res : -, Dewa Y, Nishimura J, Muguruma M, et al.: Gene expression analyses of the liver in rats treated with oxfendazole Arch Toxicol : -, Lacroix M, Toillon RA, Leclercq G: p and breast cancer, an update Endocr Relat Cancer : -,
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