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An organotypic 3D assay for primary human mammary epithelial cells that recapitulates branching morphogenesis.
Methods Mol. Biol. 1612, 125-137 (2017)
We have developed a three-dimensional organotypic culture system for primary human mammary epithelial cells (HMECs) in which the cells are cultured in free floating collagen type I gels. In this assay, luminal cells predominantly form multicellular spheres, while basal/myoepithelial cells form complex branched structures resembling terminal ductal lobular units (TDLUs), the functional units of the human mammary gland in situ. The TDLU-like organoids can be cultured for at least 3 weeks and can then be passaged multiple times. Subsequently, collagen gels can be stained with carmine or by immunofluorescence to allow for the analysis of morphology, protein expression and polarization, and to facilitate quantification of structures. In addition, structures can be isolated for gene expression analysis. In summary, this technique is suitable for studying branching morphogenesis, regeneration, and differentiation of HMECs as well as their dependence on the physical environment.
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Publikationstyp
Artikel: Journalartikel
Dokumenttyp
Wissenschaftlicher Artikel
Schlagwörter
3d Assay ; Branching Morphogenesis ; Collagen Gel ; Forskolin ; Mechanotransduction ; Organoid ; Primary Human Mammary Epithelial Cells ; Regenerative Potential ; Rho-associated Protein Kinase
ISSN (print) / ISBN
1064-3745
e-ISSN
1940-6029
Bandtitel
3D Cell Culture
Zeitschrift
Methods in Molecular Biology
Quellenangaben
Band: 1612,
Seiten: 125-137
Verlag
Springer
Verlagsort
Berlin [u.a.]
Nichtpatentliteratur
Publikationen
Begutachtungsstatus
Peer reviewed
Institut(e)
Institute of Stem Cell Research (ISF)