ABSTRACT
Contractile myoepithelial cells dominate the basal layer of the mammary epithelium and are considered to be differentiated cells. However, we observe that up to 54% of single basal cells can form colonies when seeded into adherent culture in the presence of agents that disrupt actin-myosin interactions, and on average, 65% of the single-cell-derived basal colonies can repopulate a mammary gland when transplanted in vivo. This indicates that a high proportion of basal myoepithelial cells can give rise to a mammary repopulating unit (MRU). We demonstrate that myoepithelial cells, flow-sorted using two independent myoepithelial-specific reporter strategies, have MRU capacity. Using an inducible lineage-tracing approach we follow the progeny of myoepithelial cells that express α-smooth muscle actin and show that they function as long-lived lineage-restricted stem cells in the virgin state and during pregnancy.
Subject(s)
Epithelial Cells/cytology , Mammary Glands, Animal/cytology , Myocytes, Smooth Muscle/metabolism , Stem Cells/cytology , Actins/metabolism , Animals , Benzamides/pharmacology , Cell Proliferation/drug effects , Cells, Cultured , Dioxoles/pharmacology , Epithelial Cells/metabolism , Female , Flow Cytometry , Gene Expression Profiling , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Interleukin Receptor Common gamma Subunit/deficiency , Interleukin Receptor Common gamma Subunit/genetics , Mammary Glands, Animal/metabolism , Mice, Inbred C57BL , Mice, Inbred CBA , Mice, Inbred NOD , Mice, SCID , Mice, Transgenic , Microscopy, Fluorescence , Myocytes, Smooth Muscle/cytology , Oligonucleotide Array Sequence Analysis , Receptors, Transforming Growth Factor beta/antagonists & inhibitors , Receptors, Transforming Growth Factor beta/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Stem Cells/metabolismABSTRACT
Many normal adult tissues contain rare stem cells with extensive self-maintaining regenerative potential. During development, the stem cells of the hematopoietic and neural systems undergo intrinsically specified changes in their self-renewal potential. In the mouse, mammary stem cells with transplantable regenerative activity are first detectable a few days before birth. They share some phenotypic properties with their adult counterparts but are enriched in a subpopulation that displays a distinct gene expression profile. Here we show that fetal mammary epithelial cells have a greater direct and inducible growth potential than their adult counterparts. The latter feature is revealed in a novel culture system that enables large numbers of in vitro clonogenic progenitors as well as mammary stem cells with serially transplantable activity to be produced within 7 days from single fetal or adult input cells. We further show that these responses are highly dependent on novel factors produced by fibroblasts. These findings provide new avenues for elucidating mechanisms that regulate normal mammary epithelial stem cell properties at the single-cell level, how these change during development, and how their perturbation may contribute to transformation.
Subject(s)
Epithelial Cells/cytology , Mammary Glands, Animal/cytology , 3T3 Cells , Animals , Epithelial Cells/physiology , Female , Immunohistochemistry , Mammary Glands, Animal/physiology , MiceABSTRACT
Isolation of mammary epithelial subpopulations, including stem and progenitor cells, has become a standard technique in recent years. However, a number of methods and approaches for this have developed and the relative benefits of the different approaches, and the reason for their development, have not always been clear. Here, three of the leading laboratories working on the separation of mammary cell subpopulations have summarised their methods, highlighted their differences and similarities and also discussed the reasoning behind the approaches they have taken. This article will assist workers establishing mammary cell separation protocols in their laboratories to make informed choices about the methods they should use.
