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

Farin Amersi,Bowen Gao,Ying Qu,Armando E Giuliano,Bingchen Han,Liting Jin,Stacey Chung,Catherine Dang,Liliana J Gomez,Xiaojiang Cui,Yiping Gong,Yong Yue,Xuefeng Liu

doi:10.18632/oncotarget.23817

Abstract

PurposeConditional 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.ResultsWe 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 ERα, as well as its ability to respond to estrogen stimulation.Materials and MethodsWe 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.ConclusionsThe 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.

Highlights

The mature human mammary gland is a compound tubuloalveolar structure composed of milk-secreting polarized epithelial cells surrounded by myoepithelial cells, and the two-layered tissue organization is surrounded by a basement membrane [1]
We show that cells derived from fresh normal breast tissues can be propagated and exhibit heterogeneous morphologic features
The cultures are composed of cytokeratin 18 (CK18), desmoglein 3, and cytokeratin 19 (CK19)-positive luminal cells and vimentin, p63, and CK14positive myoepithelial cells, suggesting the maintenance of in vivo heterogeneity

Summary

Introduction

The mature human mammary gland is a compound tubuloalveolar structure composed of milk-secreting polarized epithelial cells surrounded by myoepithelial cells, and the two-layered tissue organization is surrounded by a basement membrane [1]. During the past few decades, studying the physiology of the mammary gland in rodent models has greatly promoted our knowledge about hormone action [2], gene regulation [1, 3], and stem cell biology [4]. Cultured immortalized human mammary cell lines, such as MCF-10A, have been widely used as in vitro models to study the mechanisms that dictate mammary epithelial biology. Normal breast cell lines demonstrate loss of EpCAM+CD49f- and EpCAM+CD24+CD49f+ populations compared to primary breast epithelial cells isolated from reduction mammoplasty. They retain features of bipotent progenitor cells, mammary cell lines such as MCF-10A and HME I/II are unable to differentiate into mature luminal breast epithelial cells [9]. In vitro models that better recapitulate the physiologically relevant heterogeneity of the epithelial cells of human mammary gland tissue are desired

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Conclusion