TRANSFORMATION OF HUMAN BREAST EPITHELIAL CELLS IN VITRO

我们的长期目标是诱导肿瘤前和肿瘤 人乳腺上皮细胞的转化，根据 体外条件，直接和间接作用化学品 致癌物质。 一种持续改变人类乳房的能力 细胞在体外条件下的定义是必不可少的设计 实验来了解细胞分子事件背后的 人乳腺癌的转化和组织发生。 我们 策略是将人类上皮细胞在血清中转化， 胶原凝胶培养系统中的游离条件，以及 使用各种基质的单层培养系统。 具体地说， 我们将：a）分析正常，良性， 胶原蛋白中的（纤维囊性）和癌性人乳腺细胞 在激素存在下，在无血清条件下培养 （雌激素、孕激素、催乳素等），生长因子（EGF， FGF、TGF α、TGF β等）和非激素剂（脂质， TPA，Li+）; B）使用这些终点来设计培养条件 其促进人乳腺上皮细胞转化， 高产量，以类似的方式开发， 转化鼠上皮细胞; c）使用cDNA探针 确定是否扩增、重排或表达 纤溶酶原激活剂（tPA、尿激酶）、TGF α、TGF β、EGF 受体，谷胱甘肽-S-转移酶-P，γ-谷氨酰 转肽酶和0-6烷基-DNA-烷基转移酶可靠， 转化与肿瘤的定量分子标志物 组织发生; d）确定是否在分子水平上发生变化 与表型变化（核型，免疫细胞化学， e）开发合适的体外和体内测定法 系统来识别那些分子细胞的变化， 代表肿瘤前和肿瘤（恶性）转化体。 我们的经验和方法表明， 人乳腺上皮细胞体外转化的研究 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.