Abstract
IntroductionDuring pregnancy the mammary epithelial compartment undergoes extreme proliferation and differentiation, facilitated by stem/progenitor cells. Mouse mammary epithelium in nonpregnant mice contains long label-retaining epithelial cells (LREC) that divide asymmetrically and retain their template DNA strands. The role of LREC during alveogenesis has not been determined.MethodsWe performed immunohistochemistry and autoradiography on murine mammary glands that had been labeled with 5-bromodeoxyuridine (5BrdU) during allometric ductal growth to investigate the co-expression of DNA label retention and estrogen receptor-α or progesterone receptor during pregnancy. A second DNA label ([3H]-thymidine) was administered during pregnancy to identify label-retaining cells (LRC), which subsequently enter the cell cycle. Use of this methodology allowed us to investigate the co-localization of 5BrdU with smooth muscle actin, CD31, cytokeratin, and desmin in periductal or peri-acinar LRC in mammary tissue from pregnant mice subsequent to a long chase period in order to identify LRC.ResultsEstrogen receptor-α positive and progesterone receptor positive cells represented approximately 30% to 40% of the LREC, which is under 1.0% of the epithelial subpopulation. Pregnancy altered the percentage of LREC expressing estrogen receptor-α. LRC situated in periductal or peri-acinar positions throughout the gland do not express epithelial, endothelial, or myoepithelial markers, and these undefined LRCs persist throughout pregnancy. Additionally, new cycling LREC ([3H]-thymidine retaining) appear during alveologenesis, and LRC found in other tissue types (for example, endothelium and nerve) within the mammary fat pad become double labeled during pregnancy, which indicates that they may also divide asymmetrically.ConclusionsOur findings support the premise that there is a subpopulation of LREC in the mouse mammary gland that persists during alveologenesis. These cells react to hormonal cues during pregnancy and enter the cell cycle while continuing to retain, selectively, their original template DNA. In addition, nonepithelial LRC are found in periductal or peri-acinar positions. These LRC also enter the cell cycle during pregnancy. During alveologenesis, newly created label-retaining ([3H]-thymidine) epithelial cells appear within the expanding alveoli and continue to cycle and retain their original template DNA ([3H]-thymidine) strands, as determined by a second pulse of 5BrdU.
Highlights
During pregnancy the mammary epithelial compartment undergoes extreme proliferation and differentiation, facilitated by stem/progenitor cells
New cycling labelretaining epithelial cells (LREC) ([3H]-thymidine retaining) appear during alveologenesis, and label-retaining cells (LRC) found in other tissue types within the mammary fat pad become double labeled during pregnancy, which indicates that they may divide asymmetrically
Our findings support the premise that there is a subpopulation of LREC in the mouse mammary gland that persists during alveologenesis
Summary
During pregnancy the mammary epithelial compartment undergoes extreme proliferation and differentiation, facilitated by stem/progenitor cells. Mouse mammary epithelium in nonpregnant mice contains long labelretaining epithelial cells (LREC) that divide asymmetrically and retain their template DNA strands. In 1975, Cairns [1] postulated that dividing adult somatic stem cells protected themselves from mutation and cancer risk by segregating their template DNA strands. This property of selective DNA segregation was shown to occur both in vivo and in vitro in a variety of cell and tissue types [2,3,4,5,6,7,8]. In the mouse mammary gland, label-retaining epithelial cells (LREC) in the ducts divide asymmetrically and retain their template.
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