CD24 staining of mouse mammary gland cells defines luminal epithelial, myoepithelial/basal and non-epithelial cells

Sleeman, Katherine E; Kendrick, Howard; Ashworth, Alan; Isacke, Clare M.; Smalley, Matthew J.

doi:10.1186/bcr1371

Cited by 284 publications

(313 citation statements)

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“…As previously, CD24 þ /Low basal epithelial cells and CD24 þ /High luminal epithelial cells were identified (Sleeman et al, 2006). Within the basal population, a CD24 þ /Low Sca-1 À CD49f Low population (previously identified as mainly myoepithelial cells) and a CD24 þ /Low Sca-1 À CD49f High population (previously shown to be highly enriched for mammary stem cells) could be recognised Sleeman et al, 2007).…”

Section: Resultssupporting

confidence: 66%

“…Cell suspensions at 10 6 cells/ml were stained with combinations of anti-CD24-FITC (1.0 mg/ml; BD Biosciences, Oxford, UK), anti-Sca-1-APC (1.0 mg/ml; eBioscience, Hatfield, UK), anti-CD45-PE-Cy7 (1.0 mg/ml; BD Biosciences), antiCD49f-PE-Cy5 (5.0 ml/ml; BD Biosciences) and anti-c-Kit-PE (1.0 mg/ml; BD Biosciences). Cells were sorted on a FACSAria (Becton Dickenson, Oxford, UK) and dead cells, CD45 þ leucocytes and non-single cells were excluded as shown in Supplementary Figure 1 (Sleeman et al, 2006). Cells incubated in non-specific IgG were used to set the limits that defined negative and positive staining for each antibody.…”

Section: Preparation Of Mammary Epithelial Cellsmentioning

confidence: 99%

“…Cells incubated in non-specific IgG were used to set the limits that defined negative and positive staining for each antibody. Within the CD24 þ population, two separate cell populations were annotated as CD24 þ /High and CD24 þ /Low as previously described (Sleeman et al, 2006). Within the basal CD49f þ population CD49f High mammary stem cell enriched cells were defined by previous understanding and with reference to the outermost contours of a 5% interval linear density contour plot as described ( Supplementary Figure 1) (Britt et al, 2009).…”

Section: Preparation Of Mammary Epithelial Cellsmentioning

confidence: 99%

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c-Kit is required for growth and survival of the cells of origin of Brca1-mutation-associated breast cancer

et al. 2011

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BRCA1 mutation-associated breast cancer originates in oestrogen receptor-alpha-negative (ER À ) progenitors in the mammary luminal epithelium. These cells also express high levels of the Kit gene and a recent study demonstrated a correlation between Brca1 loss and Kit over-expression in the mammary epithelium. However, the functional significance of c-Kit expression in the mammary gland is unknown. To address this, c-Kit À and c-Kit þ mammary epithelial subsets were isolated by flow cytometry, characterised for expression of lineage-specific cell markers and functionally analysed by in vitro colony forming and in vivo transplantation assays. The results confirm that the majority of luminal ER À progenitors are c-Kit þ , but also that most stem cells and the differentiated cell populations are c-Kit À . A subset of c-Kit þ cells with high proliferative potential was found in the luminal ER þ population, however, suggesting the existence of a distinct luminal ER þ progenitor cell type. Analysis of mouse Brca1 mammary tumours demonstrated that they expressed Kit and its downstream effector Lyn at levels comparable to the most strongly c-Kit þ luminal ER À progenitors. Consistent with c-Kit being a progenitor cell marker, in vitro threedimensional differentiation of c-Kit þ cells resulted in a loss of c-Kit expression, whereas c-Kit over-expression prevented normal differentiation in vivo. Furthermore, c-Kit was a functional marker of proliferative potential, as c-Kit inhibition by short hairpin knockdown prevented normal epithelial growth and caused cells to undergo apoptosis. Therefore, c-Kit defines distinct progenitor populations in the mammary epithelium and is critical for mammary progenitor survival and proliferation. Importantly, c-Kit is only the second mammary epithelial stem/progenitor marker to be shown to have a functional role in the mammary epithelium and the first marker to be shown to be required for progenitor cell function. The c-Kit signalling network has potential as a target for therapy and/or prevention in BRCA1-associated breast cancer.

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Section: Resultssupporting

confidence: 66%

Section: Preparation Of Mammary Epithelial Cellsmentioning

confidence: 99%

Section: Preparation Of Mammary Epithelial Cellsmentioning

confidence: 99%

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c-Kit is required for growth and survival of the cells of origin of Brca1-mutation-associated breast cancer

et al. 2011

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“…Experimental animal models have been invaluable in mammary gland research (reviewed in Hennighausen and Robinson, 2001), enabling the identification of mammary stem (Smalley et al, 2005;Shackleton et al, 2006;Stingl et al, 2006); and luminal progenitor cells (Sleeman et al, 2006(Sleeman et al, , 2007Asselin-Labat et al, 2007) and the molecular changes that are associated with the different phases of post-pubertal mammary development (Master et al, 2002;Clarkson et al, 2004;Stein et al, 2004). Mouse models displaying defects in these processes have also been very useful, for example, Gata-3, Elf-5 and Stat5a knockout mice have helped define the cellular events involved in the development of luminal and secretory cells, respectively (Asselin-Labat et al, 2007;Oakes et al, 2008;Yamaji et al, 2009), whereas PrlR and Gal knockout mice have shed light on the molecular events associated with the secretory activation stage of lactation (Naylor et al, 2005).…”

Section: Introductionmentioning

confidence: 99%

Analysis of Brca1-deficient mouse mammary glands reveals reciprocal regulation of Brca1 and c-kit

et al. 2010

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Disruption of the breast cancer susceptibility gene Brca1 results in defective lobular-alveolar development in the mammary gland and a predisposition to breast tumourigenesis in humans and in mice. Recent evidence suggests that BRCA1 loss in humans is associated with an expansion of the luminal progenitor cell compartment in the normal breast and tumours with a luminal progenitorlike expression profile. To further investigate the role of BRCA1 in the mammary gland, we examined the consequences of Brca1 loss in mouse mammary epithelial cells in vitro and in vivo. Here, we show that Brca1 loss is associated with defective morphogenesis of SCp2 and HC11 mouse mammary epithelial cell lines and that in the MMTV-Cre Brca1 Co/Co mouse model of Brca1 loss, there is an accumulation of luminal progenitor (CD61 þ CD29 lo CD24 þ ) cells during pregnancy. By day 1 of lactation, there are marked differences in the expression of 1379 genes, with most significantly altered pathways and networks, including lactation, the immune response and cancer. One of the most differentially expressed genes was the luminal progenitor marker, c-kit. Immunohistochemical analysis revealed that the increase in c-kit levels is associated with an increase in c-kit positivity. Interestingly, an inverse association between Brca1 and c-kit expression was also observed during mammary epithelial differentiation, and small interfering RNA-mediated knockdown of Brca1 resulted in a significant increase in c-kit mRNA levels. We found no evidence that c-kit plays a direct role in regulating differentiation of HC11 cells, suggesting that Brca1-mediated induction of c-kit probably contributes to Brca1-associated tumourigenesis via another cellular process, and that c-kit is likely to be a marker rather than a mediator of defective lobular-alveolar development resulting from Brca1 loss.

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“…Strong support for this model comes from fluorescence-activated cell sorting (FACS) experiments, in which dissociated mammary cells were separated on the basis of differences in cell surface antigen expression. High CD24 glycoprotein expression marks luminal cells, whereas low CD24 expression marks myoepithelial cells 58 . The luminal CD24 hi cells can be further separated on the basis of the expression of the cell surface marker stem cell antigen 1 (SCA1; also known as LY6A) 59 .…”

Section: Box 2 | Progesterone Receptor Isoformsmentioning

confidence: 99%

Progesterone signalling in breast cancer: a neglected hormone coming into the limelight

2013

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CD24 staining of mouse mammary gland cells defines luminal epithelial, myoepithelial/basal and non-epithelial cells

Cited by 284 publications

References 14 publications

c-Kit is required for growth and survival of the cells of origin of Brca1-mutation-associated breast cancer

c-Kit is required for growth and survival of the cells of origin of Brca1-mutation-associated breast cancer

Analysis of Brca1-deficient mouse mammary glands reveals reciprocal regulation of Brca1 and c-kit

Progesterone signalling in breast cancer: a neglected hormone coming into the limelight

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