TRANSFORMATION OF HUMAN BREAST EPITHELIAL CELLS IN VITRO

我们的长期目标是诱发癌前病变和肿瘤 人乳腺上皮细胞的转化，定义如下 体外条件，含有直接和间接作用的化学物质 致癌物质。始终如一地改变人类乳房的能力 在确定的体外条件下的细胞对于设计是必不可少的 理解潜在细胞-分子事件的实验 人乳腺癌的转化和组织发生(S)。我们的 策略是将在血清中生长的人上皮细胞- 在胶原凝胶培养系统中的游离条件，以及在 使用各种基质的单层培养系统。具体来说， 我们将：a)分析正常、良性的生长规律 (纤维囊性)和癌性人乳腺细胞中的胶原 在激素存在的无血清条件下进行培养 (雌激素、黄体酮、催乳素等)、生长因子(EGF、 成纤维细胞生长因子、转化生长因子α、转化生长因子β等)和非激素性药物(脂类、 TPA，Li+)；b)使用这些端点来设计文化条件 促进人乳腺上皮细胞转化的基因 高产，其方式类似于为 转化小鼠上皮细胞；c)使用cDNA探针 确定是否扩增、重排或表达 纤溶酶原激活物(tPA、尿激活酶)、转化生长因子α、转化生长因子β、表皮生长因子 谷胱甘肽-S-转移酶-P，γ-谷氨酰受体 转肽酶和0-6烷基-DNA-烷基转移酶是可靠的， 转化与肿瘤的定量分子标记物 组织发生；d)确定是否在分子水平上发生变化 与表型变化相关(核型、免疫细胞化学、 超微结构)；e)建立适合体外和体内的检测方法 系统来识别那些分子细胞的变化， 代表癌前病变和肿瘤(恶性)转化子。 我们的经验和方法建议开发一个系统(S) 对人乳腺上皮细胞的体外转化是一种 可实现的目标。

Our long term objective is to induce preneoplastic and neoplastic transformation of human mammary epithelial cells, under defined in vitro conditions, with direct- and indirect-acting chemical carcinogens. An ability to consistently transform human mammary cells under defined in vitro conditions is essential to designing experiments to understand the cellular-molecular events underlying transformation and histogenesis of human breast cancer(s). Our strategy is to transform human epithelial cells grown under serum- free conditions in a collagen gel culture system, as well as in monolayer culture systems using various matrices. Specifically, we will: a) analyze the growth regulation of normal, benign (fibrocystic) and carcinomatous human mammary cells in collagen culture, under serum-free conditions in the presence of hormones (estrogen, progesterone, prolactin, etc.), growth factors (EGF, FGF, TGF alpha, TGF beta, etc.) and nonhormonal agents (lipids, TPA, Li+); b) use these endpoints to design culture conditions which facilitate human mammary epithelial cell transformation at high yield, in a manner similar to those developed for transformation of murine epithelial cells; c) use cDNA probes to determine whether amplification, rearrangement or expression of plasminogen activators (tPA, urokinase), TGF alpha, TGF beta, EGF receptor, glutathione-s-transferase-P, gamma-glutamyl transpeptidase and 0-6 alkyl-DNA-alkyltransferase are reliable, quantitative molecular markers for transformation and tumor histogenesis; d) determine whether changes at the molecular level correlate with phenotype changes (karyotypic, immunocytochemical, ultramicroscopic); e) develop suitable in vitro and in vivo assay systems to identify those molecular-cellular changes which represent preneoplastic and neoplastic (malignant) transformants. Our experience and approach suggest that development of a system(s) for transformation of human mammary epithelial cell in vitro is an achievable goal.