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Embryonic cells contribute directly to the quiescent stem cell population in the adult mouse mammary gland.

Boras-Granic K, Dann P, Wysolmerski JJ - Breast Cancer Res. (2014)

Bottom Line: Interestingly, long label retaining cells (labeled during puberty) are found just in front of the eLLRCs, near where the ends of the ducts had been at the time of DNA labeling in early puberty.Thus, our studies have identified a putative stem/progenitor cell population of embryonic origin.Further study of these cells will contribute to an understanding of how quiescent stem cells are generated during development and how fetal exposures may alter future breast cancer risk in adults.

View Article: PubMed Central - PubMed

Affiliation: Section of Endocrinology and Metabolism Department of Internal Medicine, Yale University School of Medicine TAC S131, Box 208020, New Haven, CT, 06520-8020, USA. granickata@gmail.com.

ABSTRACT

Introduction: Studies have identified multi-potent stem cells in the adult mammary gland. More recent studies have suggested that the embryonic mammary gland may also contain stem/progenitor cells that contribute to initial ductal development. We were interested in determining whether embryonic cells might also directly contribute to long-lived stem cells that support homeostasis and development in the adult mammary gland.

Methods: We used DNA-label retention to detect long label-retaining cells in the mammary gland. Mouse embryos were labeled with 5-ethynl-2'-deoxyuridine (EdU) between embryonic day 14.5 and embryonic day 18.5 and were subsequently sacrificed and examined for EdU retention at various intervals after birth. EdU retaining cells were co-stained for various lineage markers and identified after fluorescence activated cell sorting analysis of specific epithelial subsets. EdU-labeled mice were subjected to subsequent 5-bromo-2'-deoxyuridine administration to determine whether EdU-labeled cells could re-enter the cell cycle. Finally, EdU-labeled cells were grown under non-adherent conditions to assess their ability to form mammospheres.

Results: We demonstrate embryonically-derived, long label-retaining cells (eLLRCs) in the adult mammary gland. eLLRCs stain for basal markers and are enriched within the mammary stem cell population identified by cell sorting. eLLRCs are restricted to the primary ducts near the nipple region. Interestingly, long label retaining cells (labeled during puberty) are found just in front of the eLLRCs, near where the ends of the ducts had been at the time of DNA labeling in early puberty. A subset of eLLRCs becomes mitotically active during periods of mammary growth and in response to ovarian hormones. Finally, we show that eLLRCs are contained within primary and secondary mammospheres.

Conclusions: Our findings suggest that a subset of proliferating embryonic cells subsequently becomes quiescent and contributes to the pool of long-lived mammary stem cells in the adult. eLLRCs can re-enter the cell cycle, produce both mammary lineages and self-renew. Thus, our studies have identified a putative stem/progenitor cell population of embryonic origin. Further study of these cells will contribute to an understanding of how quiescent stem cells are generated during development and how fetal exposures may alter future breast cancer risk in adults.

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Luminal and myoepithelial lineage marker expression during embryonic mammary gland development. (A) Schematic representation of embryonic mammary rudiment formation in the female from e11.5 to birth. Five pairs of mammary glands form in the female mouse. (B) Immunostaining for K14 (green) and Gata3 (red) (top), K14 (green) and p63 (red) (middle), and K14 (green) and K8 (red) (bottom) in WT MG at e11, e13, e15, and newborn (one day old). Scale bars, 40 μm. e, embryonic day; MG, mammary gland; WT, wild type.
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Fig1: Luminal and myoepithelial lineage marker expression during embryonic mammary gland development. (A) Schematic representation of embryonic mammary rudiment formation in the female from e11.5 to birth. Five pairs of mammary glands form in the female mouse. (B) Immunostaining for K14 (green) and Gata3 (red) (top), K14 (green) and p63 (red) (middle), and K14 (green) and K8 (red) (bottom) in WT MG at e11, e13, e15, and newborn (one day old). Scale bars, 40 μm. e, embryonic day; MG, mammary gland; WT, wild type.

Mentions: In mice, mammary gland development begins around embryonic day 10.5 (e10.5) with the formation of bilateral mammary lines between the fore and hind limb buds along the ventral-lateral borders of the embryo. Cells within the mammary line coalesce into five distinct pairs of placodes (three thoracic and two inguinal). Over the next several days, each mammary placode expands and invaginates into the underlying mesenchyme to form a mammary bud (Figure 1A). Mammary rudiments have very low proliferative activity between e11.25 and e13.5 and the initial phases of mammary development are thought to rely on cell migration from the epidermis rather than proliferation of mammary epithelial cells [1-3]. Active proliferation within the mammary epithelium begins at e14.5 [4]. By e15.5, the distal end of the mammary bud begins to elongate into the underlying dermal mesenchyme to form a sprout. The sprout grows downward into the mammary fat pad, an adipocyte-rich stromal compartment and begins to branch, forming the rudimentary ductal tree by e18.5. By birth, the mammary epithelium consists of a primary duct and about 10 to 15 branches located within the proximal end of the nascent mammary fat pad.Figure 1


Embryonic cells contribute directly to the quiescent stem cell population in the adult mouse mammary gland.

Boras-Granic K, Dann P, Wysolmerski JJ - Breast Cancer Res. (2014)

Luminal and myoepithelial lineage marker expression during embryonic mammary gland development. (A) Schematic representation of embryonic mammary rudiment formation in the female from e11.5 to birth. Five pairs of mammary glands form in the female mouse. (B) Immunostaining for K14 (green) and Gata3 (red) (top), K14 (green) and p63 (red) (middle), and K14 (green) and K8 (red) (bottom) in WT MG at e11, e13, e15, and newborn (one day old). Scale bars, 40 μm. e, embryonic day; MG, mammary gland; WT, wild type.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4308878&req=5

Fig1: Luminal and myoepithelial lineage marker expression during embryonic mammary gland development. (A) Schematic representation of embryonic mammary rudiment formation in the female from e11.5 to birth. Five pairs of mammary glands form in the female mouse. (B) Immunostaining for K14 (green) and Gata3 (red) (top), K14 (green) and p63 (red) (middle), and K14 (green) and K8 (red) (bottom) in WT MG at e11, e13, e15, and newborn (one day old). Scale bars, 40 μm. e, embryonic day; MG, mammary gland; WT, wild type.
Mentions: In mice, mammary gland development begins around embryonic day 10.5 (e10.5) with the formation of bilateral mammary lines between the fore and hind limb buds along the ventral-lateral borders of the embryo. Cells within the mammary line coalesce into five distinct pairs of placodes (three thoracic and two inguinal). Over the next several days, each mammary placode expands and invaginates into the underlying mesenchyme to form a mammary bud (Figure 1A). Mammary rudiments have very low proliferative activity between e11.25 and e13.5 and the initial phases of mammary development are thought to rely on cell migration from the epidermis rather than proliferation of mammary epithelial cells [1-3]. Active proliferation within the mammary epithelium begins at e14.5 [4]. By e15.5, the distal end of the mammary bud begins to elongate into the underlying dermal mesenchyme to form a sprout. The sprout grows downward into the mammary fat pad, an adipocyte-rich stromal compartment and begins to branch, forming the rudimentary ductal tree by e18.5. By birth, the mammary epithelium consists of a primary duct and about 10 to 15 branches located within the proximal end of the nascent mammary fat pad.Figure 1

Bottom Line: Interestingly, long label retaining cells (labeled during puberty) are found just in front of the eLLRCs, near where the ends of the ducts had been at the time of DNA labeling in early puberty.Thus, our studies have identified a putative stem/progenitor cell population of embryonic origin.Further study of these cells will contribute to an understanding of how quiescent stem cells are generated during development and how fetal exposures may alter future breast cancer risk in adults.

View Article: PubMed Central - PubMed

Affiliation: Section of Endocrinology and Metabolism Department of Internal Medicine, Yale University School of Medicine TAC S131, Box 208020, New Haven, CT, 06520-8020, USA. granickata@gmail.com.

ABSTRACT

Introduction: Studies have identified multi-potent stem cells in the adult mammary gland. More recent studies have suggested that the embryonic mammary gland may also contain stem/progenitor cells that contribute to initial ductal development. We were interested in determining whether embryonic cells might also directly contribute to long-lived stem cells that support homeostasis and development in the adult mammary gland.

Methods: We used DNA-label retention to detect long label-retaining cells in the mammary gland. Mouse embryos were labeled with 5-ethynl-2'-deoxyuridine (EdU) between embryonic day 14.5 and embryonic day 18.5 and were subsequently sacrificed and examined for EdU retention at various intervals after birth. EdU retaining cells were co-stained for various lineage markers and identified after fluorescence activated cell sorting analysis of specific epithelial subsets. EdU-labeled mice were subjected to subsequent 5-bromo-2'-deoxyuridine administration to determine whether EdU-labeled cells could re-enter the cell cycle. Finally, EdU-labeled cells were grown under non-adherent conditions to assess their ability to form mammospheres.

Results: We demonstrate embryonically-derived, long label-retaining cells (eLLRCs) in the adult mammary gland. eLLRCs stain for basal markers and are enriched within the mammary stem cell population identified by cell sorting. eLLRCs are restricted to the primary ducts near the nipple region. Interestingly, long label retaining cells (labeled during puberty) are found just in front of the eLLRCs, near where the ends of the ducts had been at the time of DNA labeling in early puberty. A subset of eLLRCs becomes mitotically active during periods of mammary growth and in response to ovarian hormones. Finally, we show that eLLRCs are contained within primary and secondary mammospheres.

Conclusions: Our findings suggest that a subset of proliferating embryonic cells subsequently becomes quiescent and contributes to the pool of long-lived mammary stem cells in the adult. eLLRCs can re-enter the cell cycle, produce both mammary lineages and self-renew. Thus, our studies have identified a putative stem/progenitor cell population of embryonic origin. Further study of these cells will contribute to an understanding of how quiescent stem cells are generated during development and how fetal exposures may alter future breast cancer risk in adults.

Show MeSH
Related in: MedlinePlus