Stem cells and the mammary microenvironment - PubMed (original) (raw)

Stem cells and the mammary microenvironment

Brian W Booth et al. Breast Dis. 2008.

Abstract

An entire mammary epithelial outgrowth, capable of full secretory differentiation, may comprise the progeny of a single cellular antecedent. This conclusion is based upon the maintenance of retroviral insertion sites within the somatic DNA of successive transplant generations derived from a single mammary fragment. In addition, dissociation of these clonal dominant glands and implantation of dispersed cells at limiting dilution demonstrated that both duct-limited and lobule-limited outgrowths were developed as well as complete, fully differentiated glands. Thus, transplantation has revealed three distinct mammary epithelial progenitors in the mouse. Recently, using cre-lox conditional activation of reporter genes, the lobule-limited progenitor was lineally marked by lacZ expression. In situ, these cells were shown to regenerate secretory lobules upon successive pregnancies. In transplant studies, they demonstrated the capacity for self- renewal and contributed to the new generation of all of the epithelial cell types among mammary secretory lobules. Using this conditional activation model, cells isolated from other tissues of the WAP-Cre/Rosa26/lacZReporter mice, co-mingled with normal wild type mammary epithelial cells and transplanted into epithelium-divested mammary fat pads, were shown to be amenable to redirection of their cell fate by interaction with the mammary microenvironment in vivo. This suggests the ascendancy of the microenvironment over the intrinsic nature of somatic stem cells.

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Figures

Fig. 1

Fig. 1

Schematic illustration depicting self-renewal and differentiation of stem/progenitor cells in he murine mammary epithelium.

Fig. 2

Fig. 2

Whole mount examples of lobule-limited and duct-only mammary outgrowths with their corresponding cross sections. Note the absence of ductal tree in the lobule only outgrowth and the absence of developing acinar structures in duct only outgrowth. The white arrows in lobule only cross section and black arrows in duct only cross section denote myoepithelial cells. Arrows in duct only whole mount indicate cap cells.

Fig. 3

Fig. 3

Both duct-limited and lobule-limited outgrowths possessed ER_α_ and PR-positive epithelial cells. Arrows indicate cells expressing the nuclear steroid receptors in lobule-limited and ductal-limited mammary outgrowths.

Fig. 4

Fig. 4

Total DNA, from serially transplanted clonal dominant epithelial populations of five successive generations, was subjected to digestion with EcoR1 followed by Southern Blot analysis. The subsequent blot was probed with a P32 – labeled MMTV-LTR-specific probe. EcoR1 cuts within the genome of the MMTV producing 2 host-viral junction fragments of each provirus insertion. In panel A, five specific host-viral restriction fragments (arrows) were found in the original outgrowth, lane 6, and in all of the succeeding generations. Transplant generations 2 through 5 are represented by the middle 4 lanes, # 2 through 5. The DNA in lane 1 is from the mammary tumor that arose in a 4_th_ generation outgrowth. In panel B, DNA from fully developed R12 outgrowth (lane 7) is compared to DNA from a lobule-limited outgrowth in the contra-lateral gland at parturition (lane 8). In both cases, all five MMTV-host restriction fragments are detected.

Fig. 5

Fig. 5

Hyperplastic mammary outgrowths contain both luminal and myoepithelial cells. SEM of HOG demonstrating the presence of MMTV and the development of luminal epithelial cells around the lumen with the underlying myoepithelial cells. A small light cell (SLC) is also present in a suprabasal position.

Fig. 6

Fig. 6

Mouse mammary glands contain nuclear label retaining cells. Nuclei positive for3HtdR alone (A, E) or 5BrdU alone (B–D). Doubly labeled 5BrdU/3HtdR nuclei, singly labeled 3HtdR-positive nuclei and 5BrdU-labeled nuclei were often juxtaposed suggesting that their labeling resulted from a recent mitotic even (double arrows in K–M). Singly labeled3HtdR nuclei in 5BrdU-labeled mammary tissues (E, N). Scale bars =10 _μ_m.

Fig. 7

Fig. 7

The mammary gland microenvironment is dominant over foreign stem cells. Freshly isolated WAP-Cre/Rosa26-fl-stop-fl-LacZ testicular cells were mixed with equal numbers of wild type mammary epithelial cells and transplanted into the “cleared” mammary fat pads of 3-week old Nu/Nu host females. After pregnancy-induced recombination lacZ expressing progeny of the testicular cells are observed throughout the recapitulated mammary gland and these cells remain after involution and into second generation mammary transplants.

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