Isolating Mammary Gland Stem Cells

Female mammary glands are home to a remarkable population of stem cells that grow in culture as small balls of cells called “mammospheres.” Clayton and others were able to identify these stem cells in 2004 (Clayton, Titley, and Vivanco, Exp Cell Res 297 (2004): 444-60), and Max Wicha’s laboratory at the University of Michigan showed that a signaling molecule called Sonic Hedgehog and a Polycomb nuclear factor called Bmi-1 are necessary for the self-renewal of normal and cancerous mammary gland stem cells (Lui, et al., Cancer Res June 15, 2006 66; 606). The biggest problem with mammary gland stem cells is isolating them from the rest of the mammary tissue.

Mammary gland stem cells or MaSCs are very important for mammary gland development and during the induction of breast cancer. Getting cultures of MsSCs is really tough because the MaSCs share cell surface markers with normal cells and they are also quite few in number.

Gregory Hannon and his co-workers at Cold Spring Harbor Laboratory used a mouse model to identify a novel cell surface protein specific to MaSCs. By exploiting this unusual marker, Hannon and his team were able to isolate MaSCs from mouse mammary glands of rather high purity.

Camila Do Santos, the paper’s first author, said that “We are describing a marker called Cd1d.” Cd1d is also found on the surfaces of cells of the immune system, but is specific to MaSCs in mammary tissue. Additionally, MaSCs divide slower than the surrounding cells. Do Santos and her colleagues used this feature to visually isolate MaSCs from cultured mammary cells.

They used a mouse strain that expresses a green glowing protein in its cells and then made primary mammary cultures from these green glowing mice. After shutting of the expression of the green glowing protein with doxycycline, the cultured cells divided, and diluted the quantity of green glow protein in the cells. This caused them to glow less intensely. However, the slow-growing MaSCs divided much more slowly and glowed much more intensely. Selecting out the most intensely glowing cells allowed Dos Santos and her colleagues to enrich the culture for MaSCs.

“The beauty of this is that by stopping GFP expression, you can directly measure the number of cell divisions that have happened since the GFP was turned off,” said Dos Santos. She continued: “The cells that divide the least will carry GFP the longest and are the ones we characterized.”

Using this strategy, Dos Santos and others selected stem cells from the mammary glands in order to examine their gene expression signature. They also confirmed that by exploiting Cd1d expression in the MaSCS, in combination with other techniques, they could enhance the purity of the cultures several fold.

Hannon added, “With this advancement, we are now able to profile normal and cancer stem cells at a very high degree of purity, and perhaps point out which genes should be investigated as the next breast cancer drug targets.”

Will we be able to use these cell for therapeutic purposes some day?  Possibly, but at this time, more must be known about them and MaSCs must be better characterized.

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Professor of Biochemistry at Spring Arbor University (SAU) in Spring Arbor, MI. Have been at SAU since 1999. Author of The Stem Cell Epistles. Before that I was a postdoctoral research fellow at the University of Pennsylvania in Philadelphia, PA (1997-1999), and Sussex University, Falmer, UK (1994-1997). I studied Cell and Developmental Biology at UC Irvine (PhD 1994), and Microbiology at UC Davis (MA 1986, BS 1984).