The Mammary Gland Contains Distinct Ductal and Alveolar Progenitor Cells

An in vivo transplantation system was used to evaluate the developmental capacities of specific mouse mammary epithelial cell populations. Specifically, mouse mammary epithelial cells with distinctly limited developmental potentials were identified using this procedure. Two distinct epithelial cell progenitors have been identified by experiments designed to determine whether basal lobular and ductal phenotypes could develop independently under conditions imposed by a limiting dilution. The prediction that these separate epithelial progenitors must exist was based upon the results from transplantation experiments carried out in epithelium-divested mammary fat pads of syngeneic mice with mammary epithelium from two different transgenic mouse models; WAP-TGF-b1 and WAP-Int3. In both of these transgenic models, lobular development is curtailed while ductal growth and extension is relatively unaffected. Transplantation demonstrated that this phenotype is a property of the transgenic epithelium and is not due to extracellular effects mediated within the transgenic environment.

The repopulating capacity of primary mammary epithelial cell cultures derived from immature female virgin mice was tested by limiting dilution transplantation into the cleared mammary fat pads of virgin hosts who were subsequently impregnated. Primary cell cultures were developed from WAP-LacZ transgenic virgin female mice. The secretory epithelium in WAP-LacZ mouse mammary glands uniformly express LacZ at parturition. The endpoint dilution for obtaining epithelial growth in a cleared fat pad was 2000 epithelial cells from a primary culture. Non-transgenic mammary fibroblasts were used as non-epithelial diluting cells so that approximately 1 Million cells were contained in each inoculum placed in a given mammary fat pad. Twenty-two of 34 (64.7%) mammary fat pads showed epithelial growth. The extent of epithelial growth varied greatly and in some fat pads, more than one site of epithelial growth was observed.

Transplantation into the cleared mammary fat pads of virgin hosts

Several types of outgrowths occurred (see Figure to the left); secretory lobule proliferation only (10 0f 22) (Figure 1B) (enlarged image); ductal phenotype only (1 of 22) (Figure 1C) (enlarged image), and those comprised of a normal mixture of ductal and lobular growth and development (11 of 22) (Figure 1A) (enlarged image). Figure 1D (enlarged image)) shows a chimera comprised of lacZ positive and lacZ negative cells which arose from a mixture of transgenic and non-transgenic epithelial cells that occasionally contaminated the dilutary fibroblasts.

These results demonstrate the following points:

• lobular i.e. secretory progenitor cells are present as distinct entities among the mammary epithelial cells found in immature virgin female mice
• similarly, ductal epithelial progenitors are present within the same population
• lobular progenitors are present in greater numbers (at least in primary cultures), although both cell populations are extremely small,
• as expected, some inocula produced outgrowths with simultaneous development of both lobular and ductal phenotypes. The latter result may indicate cooperative interaction between the two epithelial progenitors or signal the presence of a third progenitor type, i.e. a mammary epithelial stem cell capable of producing both ductal and lobular- committed daughters (see model below).
• these findings have important consequences in the design of experiments aimed at testing the effects of known and putative mammary oncogenes and tumor suppressor genes, using techniques which include cellular transformation in vitro followed by in vivo cultivation and evaluation.

Conclusions

Understanding the determined epithelial stem cells of the mammary gland is an important link in our ability to perceive the cellular pathways leading to malignant transformation in mammary epithelium. In the present work and in a growing number of transgenic and gene-knockout mouse models the multifaceted nature of epithelial growth and differentiation in mammary tissue is being gradually revealed. The application of the mammary fat pad transplantation techniques and genetic manipulation of the mouse genome present a powerful methodology for furthering our understanding of the regulatory and signaling strategies inherent in normal mammary growth, development and differentiation.

Stem Cell Model

Citations:

Smith, G.H. (1996) Experimental mammary epithelial morphogenesis in an in vivo model: Evidence for distinct cellular progenitors of the ductal and lobular phenotype. Breast Cancer Research and Treatment 39: 21-31.
Chepko, G and Smith, G.H. (997) Three divivsion-competent, structurally distinct cell populations contribute to the murine mammary epithelial renewal. Tissue and Cell 29: 239-253.

Keywords:
stem cells, progenitors, lobulogenesis, branching morphogenesis

Contributed by:

Gilbert H. Smith
in April 1996
Gilbert H. Smith
tel. (301) 496-2385
FAX (301) 402-0711
(e-mail smithg@ltiblp.nci.nih.gov)