Alternative to breast carcinogenicity test using a tissue culture method

我们开发了一种新的体外微核试验，使用培养的小鼠乳腺上皮细胞作为组织特异性筛查可疑的乳腺癌致癌物。将妊娠小鼠乳腺上皮细胞培养于无血清的Ham’s F-12/Dulbecco’s改良Eagle培养基中，培养基中添加胰岛素、表皮生长因子、转铁蛋白、霍乱毒素、牛血清白蛋白和抗生素。在培养的第6天，将几种乳腺癌致癌物和其他化学物质添加到培养物中，并在处理后24小时对每1000个细胞的微核数量进行评分。在本研究中，五种乳腺癌致癌物n -甲基-n -亚硝基脲、n -丁基-n -亚硝基脲、7,12 -中二甲基苯(a)蒽、20-甲基胆蒽和2-乙酰氨基芴，以及一种直接致癌物n -甲基-n '-硝基-硝基胍增加了培养物中微核的发生率。由于所有的乳腺癌致癌物和一种直接致癌物在培养的小鼠乳腺上皮细胞中都以剂量相关的方式诱导微核，但其他致癌物通常不会，因此这些结果证明了使用微核作为短期、体外筛查可疑乳腺癌致癌物是合理的。

We developed a new in vitro micronucleus test using cultured mouse mammary epithelial cells as a tissue-specific screen for suspected breast carcinogens.Mammary epithelial cells from pregnant mice were cultured in serumfree Ham's F-12/Dulbecco's modified Eagle's medium supplemented with insulin, epidermal growth factor, transferin, cholera toxin, bovine serum albumin and antibiotics. Several breast carcinogens and other chemicals were added to the culturds at day 6 of culture, and the number of micronuclei per 1, 000 cells was scored at 24hr after treatment.In the present study, five breast carcinogens, N-methyl-N-nitrosourea, N-butyl-N-nitrosourea, 7, 12-mdimethylbenz (a) anthracene, 20-methylcholanthrene and 2-acetamidofluorene, and one direct carcinogen, N-methyl-N'-nitro-Nnitrosoguanidine increased the micronuclei incidence in the cultures.Since all breast carcinogens and a direct carcinogen induced micronuclei in the culture of mouse mammary epitherial cells in a dose related manner, but other agents generally do not, these results justify the use of micronuclei as a short term, in vitro screen of suspected breast carcinogens.