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Characterization of primary human mammary epithelial cells isolated and propagated by conditional reprogrammed cell culture.

ABSTRACT: Conditional reprogramming methods allow for the inexhaustible in vitro proliferation of primary epithelial cells from human tissue specimens. This methodology has the potential to enhance the utility of primary cell culture as a model for mammary gland research. However, few studies have systematically characterized this method in generating in vitro normal human mammary epithelial cell models.We show that cells derived from fresh normal breast tissues can be propagated and exhibit heterogeneous morphologic features. The cultures are composed of CK18, desmoglein 3, and CK19-positive luminal cells and vimentin, p63, and CK14-positive myoepithelial cells, suggesting the maintenance of in vivo heterogeneity. In addition, the cultures contain subpopulations with different CD49f and EpCAM expression profiles. When grown in 3D conditions, cells self-organize into distinct structures that express either luminal or basal cell markers. Among these structures, CK8-positive cells enclosing a lumen are capable of differentiation into milk-producing cells in the presence of lactogenic stimulus. Furthermore, our short-term cultures retain the expression of ER?, as well as its ability to respond to estrogen stimulation.We have investigated conditionally reprogrammed normal epithelial cells in terms of cell type heterogeneity, cellular marker expression, and structural arrangement in two-dimensional (2D) and three-dimensional (3D) systems.The conditional reprogramming methodology allows generation of a heterogeneous culture from normal human mammary tissue in vitro. We believe that this cell culture model will provide a valuable tool to study mammary cell function and malignant transformation.

SUBMITTER: Jin L

PROVIDER: S-EPMC5837767 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Characterization of primary human mammary epithelial cells isolated and propagated by conditional reprogrammed cell culture.

Jin Liting L Qu Ying Y Gomez Liliana J LJ Chung Stacey S Han Bingchen B Gao Bowen B Yue Yong Y Gong Yiping Y Liu Xuefeng X Amersi Farin F Dang Catherine C Giuliano Armando E AE Cui Xiaojiang X

Oncotarget 20171222 14

<h4>Purpose</h4>Conditional reprogramming methods allow for the inexhaustible <i>in vitro</i> proliferation of primary epithelial cells from human tissue specimens. This methodology has the potential to enhance the utility of primary cell culture as a model for mammary gland research. However, few studies have systematically characterized this method in generating <i>in vitro</i> normal human mammary epithelial cell models.<h4>Results</h4>We show that cells derived from fresh normal breast tissu ...[more]

PMID: 29545915

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Project description:The ability to culture normal human mammary epithelial cells (HMEC) greatly facilitates experiments that seek to understand both normal mammary cell biology and the many differences between normal and abnormal human mammary epithelia. To maximize in vivo relevance, the primary cell culture conditions should maintain cells in states that resemble in vivo as much as possible. Towards this goal, we compared the properties of HMEC strains from two different reduction mammoplasty tissues that were grown in parallel using different media and culture conditions. Epithelial organoids were initiated into three different media: two commonly used serum-free-media, MCDB 170-type (e.g. MEGM) and WIT-P, and a low stress media, M87A. Growth, lineage heterogeneity, p16 protein expression, and population doublings to senescence were measured for each culture condition. MCDB 170 caused rapid senescence and loss of heterogeneity within 2 to 3 passages, but some cultures went through the 1 to 2 month process of selection to generate clonal finite post-selection post-stasis cells. WIT-P caused impressive expansion of luminal cells in 2nd passage followed by their near complete disappearance by passage 4 and senescence shortly thereafter. M87A supported as much as twice the number of population doublings compared to either serum-free medium, and luminal and myoepithelial cells were present for as many as 8 passages. Thus, of the three media compared, WIT-P and MCDB 170 imposed rapid senescence and loss of lineage heterogeneity, phenotypes consistent with cells maintained in high-stress conditions, while M87A supported cultures that maintained multiple lineages and robust growth for up to 60 population doublings. In conjunction with previous studies examining the molecular properties of cultures grown in these media, we conclude that M87A medium is most able to support long-term culture of multiple lineages similar to in vivo conditions, thereby facilitating investigations of normal HMEC biology relevant to the mammary gland in situ.

Dataset Information

Characterization of primary human mammary epithelial cells isolated and propagated by conditional reprogrammed cell culture.

Publications

Characterization of primary human mammary epithelial cells isolated and propagated by conditional reprogrammed cell culture.

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