Mammary Stem Cells
(Scientific Summary)

Breast cancer is the most common form of cancer in women, and every year hundreds of thousands of women are newly diagnosed (American Cancer Society 2006).  Most breast cancers originate in the mammary ducts, while a smaller subset originates in the lobules (Colditz 1993).  Since cancer is a stem cell disorder, understanding the basic stem cell biology of mammary cells will help researchers find treatments and maybe even a cure for breast cancer.

During development, mammary glands are derived from ectoderm.  Prior to sexual maturity, the mammary gland consists of a rudimentary ductal system.  Enhanced secretions of estrogen at puberty help develop the mammary ductal system.  The basic functional units of the branching ducts of the mammary gland are terminal ductal lobulo-alveolar units (TDLUs).  Luminal or ductal epithelial cells form the inner layer of the ducts, while myoepithelial cells compose the outer basal layer of the ducts.  During pregnancy, estrogen and progesterone cause further branching of the ductal system and the development of milk-secreting structures known as lobules.

Figure 1.  Basic breast anatomy.  Breast profile: A) Ducts, B) Lobules, C) Dilated section of lobules, D) Nipple, E) Fat, F) Pectoralis Major Muscle, and G) Chest wall/rib cage.

Mammary stem cells (MaSCs) have a fundamental role in the regeneration of mammary tissues during cycles of pregnancy, lactation and involution.  In the mammary epithelium of mice, the number of potential MaSCs is approximately 1 in 2,500 cells (Smith 1996, Kordon 1998).  The existence of MaSCs in mice has been confirmed by limiting-dilution transplantation studies (Smith 1996, Shackleton 2006), clonal analysis (Kordon 1998) and X-chromosome inactivation pattern analysis (Tsai 1996).  Retroviral tagging has shown that clonally derived MaSCs are found throughout the fully developed mammary gland (Kordon 1998).

The existence of a pluripotent progenitor of three distinct self-renewing, multipotent epithelial cells (lobule-limited, ductal-limited, fully competent) has been suggested (Smith 2002). These three distinct self-renewing, multipotent epithelial cell types have been isolated in the murine model (Smith 1996).  Lobule-limited stem cells are able to produce secretory lobules, while ductal-limited stem cells are able to produce mammary ducts.  Fully competent mammary stem cells have the ability to generate the entire mammary epithelium.  The exact lineage of mammary stem cells has not been entirely determined, but a couple of models have been proposed (Figure 2). 

Figure 2. Possible lineages of mammary stem cells.

In order to distinguish mammary stem cells from other cells, distinctive characteristics that MaSCs possess have to be revealed.  Identifying these unique markers will aid in future research and possibly play an important role in breast cancer treatment.  MaSCs are pale or light-staining cells (Smith 1988).  They can either be undifferentiated large light cells (ULLC) or small light cells (SLC) (Chepko 1997).  These stem cells are CD29Hi, CD24+, CD49fHi and Sca-1+ (Shackleton 2006, Stingl 2006, Welm 2002).  Also, MaSCs like other stem cells lack lineage markers.  Since high levels of CD29 and CD49f indicate the excessive presence of cell adhesion molecules (β1 and α6-integrins, respectively), these molecules might play an important role in MaSC homing and maintenance.  MaSCs also express epithelial-specific antigen (ESA) and keratin K19, but are negative for sialomucin (MUC-1) (Gudjonsson 2002).  In addition, MaSCs can either be estrogen receptor positive (ER+) or estrogen receptor negative (ER-).  Gudjonsson et al has found long-term repopulating stem cells to be ER-, while short-term stem cells are ER+.

MaSCs are located throughout the mature mammary gland (Kordon, 1998).  ULLC are located between the basal and luminal layer of the mammary gland, and they possess large nuclei.  SLC, on the other hand, have been located in the basal layer (Chepko 1997). 

There is strong support for the presence of multipotent tumorigenic stem cells within leukemia (Bonnet 1997) and solid tissue tumors including breast (Singh 2004, Galli 2004, Ponti 2005).  Upon transplantation, these self-renewing, tumor initiating cells give rise to additional tumorigenic stem cells as well as heterogeneous population of differentiated, non-malignant cells indicative of the developmental lineage of the tissue (Reya 2001).  Breast cancer cells of CD44+/CD24-/low/ESA+/Lin- phenotype were found to generate proliferative tumors in NOD/SCID mice (Al-Hajj 2003), and have since been estimated to represent 10-20% of total cells in a breast tumor population (Abraham 2005). CD44 and CD 24 cell surface markers are both adhesion molecules, and ESA is the epithelial specific antigen.  Such breast cancer stem cells have also recently been identified in distributed tumor cells present in the bone marrow of early breast cancer patients (Balic 2006), further suggesting the role these cells play in not only the origin but also the progression of breast cancer.

The origin of cancer stem cells is unclear, and tumorigenic stem cells may arise from normal stem cells or progenitor cells that acquire the capacity for self-renewal (Kristov 2006).  Other models surmise that tumor initiating cells are the result of cell-cell fusions or phagocytic gene-transfer from tumor cells (Bjerkvig 2005, Bergsmedh 2001).  Data supports that tumor initiating cells in the breast are phenotypically similar to early multipotent epithelial cells (Gudjonsson 2002, Stingl 1998, Aigner 1997) and that ER+ breast cancers likely originate in ER+ ULLC mammary stem cell populations (Clarke 2005, Gudjonsson 2002).  Gene pathways implicated in the regulation of stem cell self-renewal such as Bmi-1, Wnt/ß-catenin, Notch, PTEN and Hedgehog are tightly regulated in normal stem cells, and disruption of these pathways appears essential for development of tumor initiating cells (Bjerkvig 2005, Al-Hajj 2004).

The presence of rare cells in breast cancers responsible for tumorigenesis indicates the need of targeted therapeutics capable of selectively addressing the malignant cell population.  Specific cell surface markers have been identified for mammary stem cells and breast cancer stem cells, opening the way for vastly improved, targeted cytotoxic therapeutics and accurate breast cancer diagnostics.  However current tools are not readily available to exploit the full potential of these findings.  Also, findings reported in this summary employed murine models rather than human models.  Hopefully, these shortcomings will be ameliorated through continued efforts in the promising field of stem cells.

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Summarized by:  Joyce Kim, Jason Moore and Ajitha Patlolla, Graduate Course in Stem Cell Biology, Fall 2006

Teaching Assistant: Steve Greco