Primary culture of human mammary epithelial cells embedded in collagen gels.
Primary culture of human mammary epithelial cells embedded in collagen gels.
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嵌入胶原凝胶中的人乳腺上皮细胞的原代培养物。
DOI：
10.1093/jnci/65.2.337
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发表时间：
1980
期刊：
影响因子：
--
通讯作者：
S. Nandi
中科院分区：
文献类型：
--
作者：
J. Yang;R. Guzman;J. Richards;V. Jentoft;M. Devault;S. Wellings;S. Nandi
从乳房切除组织中分离人乳腺上皮细胞。使用 Percoll 密度梯度离心去除污染的成纤维细胞，该离心利用上皮细胞和成纤维细胞之间的浮力密度差异。然后将高度富集乳腺上皮细胞的制剂包埋在胶原凝胶中并在含有12.5%马血清、2.5%胎牛血清、0.1微克霍乱毒素/ml、从人男性尿液制备的提取物（L微克蛋白质/ml）以及10微克胰岛素/ml、10微克人胎盘催乳素/ml、1微克的激素组合的Ham's F12培养基中培养。醛固酮/ml，和0.5微克氢化可的松/ml。在原代培养物中实现了持续生长，导致细胞数量比初始值增加十倍至三十倍，并且这种生长甚至在传代至二次培养物后仍得以维持。尿液提取物或激素组合的缺失都会导致生长达不到最佳状态。随后的研究表明，单独使用氢化可的松可以替代激素组合。此外，尿液提取物可以用从人的肾脏或大脑中提取的提取物代替。胶原凝胶系统提供了一种可重复且一致的方法，用于在原代和传代培养物中使人乳腺组织的乳腺上皮细胞持续三维生长。
Human mammary epithelial cells were dissociatd from mastectomy tissues. The contaminating fibroblasts were removed by the use of Percoll density-gradient centrifugation, which utilizes the difference in buoyant densities between epithelial cells and fibroblasts. A preparation highly enriched for mammary epithelial cells ws then embedded in collagen gel and cultured in Ham's F12 medium containing 12.5% horse serum, 2.5% fetal calf serum, 0.1 microgram cholera toxin/ml, an extract prepared from human male urine (L microgram protein/ml), and a hormone combination of 10 microgram insulin/ml, 10 microgram human placental lactogen/ml, 1 microgram aldosterone/ml, and 0.5 microgram hydrocortisone/ml. Sustained growth leading to an increase of tenfold to thirtyfold in cell number over the initial value was accomplished in primary culture, and this growth was maintained even after passage to secondary culture. Deletion of either the urine extract or the hormone combination resulted in less than optimal growth. Subsequent studies showed that hydrocortisone alone could replace the hormone combination. In addition, urine extract could be replaced by extracts prepared from human kidneys or brains. The collagen gel system provies a reproducible and consistent method for sustained three-dimensional growth of mammary epithelial cells from human breast tissue in primary as well as passaged cultures.