Isolation and purification of mouse mammary epithelial cells for primary culture in 2 and 3 dimensions.
Aurélie Lacouture, Cynthia Jobin, Dominic Bastien, Cindy Weidmann, Isabelle Laverdière, Étienne Audet-Walsh.
Few in vitro models are used to study mammary epithelial cells (MEC) and most of them do not express hormone receptors such as the estrogen receptor α (ERα), thus decreasing their experimental value for studies in molecular endocrinology. The usage of primary MEC is an attractive alternative avenue to overcome such issue. Still, published methods to purify these cells require fluorescence-activated cell sorting (FACS), which are time-consuming, involve access to specialized instruments, and necessitate specific expertise to be developed. In the current study, our objective was to elaborate a FACS-free protocol to purify primary mouse MEC for cell culture in 2 and 3 dimensions. We developed an isolation protocol with sequential digestions and a purification method based on specific antibodies coupled to magnetic beads. These purified cells can be plated for cell culture in 2-dimensions, in which they form colonies of epithelial cells that remain differentiated up to 6 days with an enrichment of luminal epithelial cells of 85%. When cells are seeded in Matrigel discs for 3-dimensions culture, they form visible organoids in 2-3 days which grow up to 14 days and remain viable for at least 1 month in culture. These organoids have lumens, contractile cells as well as lobular structure which recapitulate the morphology of the mammary gland in vivo. Our preliminary results indicate that these organoids express ERα and are responsive to estrogens. Obtaining this new functional study model in both 2 and 3 dimensions will enable us to better understand the functions of the mammary gland, particularly in a lactation context and in breast cancer research.
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(Événement intégré au programme des Midis Parlons Sciences de l’Axe Endo-Néphro !)