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. Author manuscript; available in PMC: 2019 Feb 13.
Published in final edited form as: Oncogene. 2018 Aug 13;38(3):445–453. doi: 10.1038/s41388-018-0439-1

Figure 4. Overexpression of miR-489 in mammary epithelial cells has reduced Ma-CFC population.

Figure 4

(A) FACS analysis was performed to examine the luminal progenitor cell population (CD49fmedCD24hi) in mammary epithelial cells from 6-week-old WT and MMTV-miR489 transgenic mice (n=3). P=0.0016 (unpaired student t test). (B) Colony formation assay was performed from mammary epithelial cells from 6-week-old WT and MMTV-miR489 transgenic mice (n=3). P=0.0038 (unpaired student t test). (C) Mammary epithelial cells were isolated from 24-week-old Her2 and Her2-miR489 mice and stained with CD49f-FITC and CD24-PE and analyzed for Ma-CFC population (n=3). P=0.0015 (unpaired student t test). (D) FACS analysis of the putative CSC cells (CD49fhiCD61hi population) in the first mammary tumor of each animal (n=8). P=0.0337 (unpaired student t test) Method: Thoracic and inguinal mammary glands were isolated from 6-week-old WT and MMTV-miR-489 mice, single cell mammary epithelial cells suspension were prepared and stained with CD49f and CD24 antibody as described above. Cells were analyzed with BD Accuri C6 flow cytometer for different mammary epithelial cell population. A total of 35,000 events were collected for each sample. Mammary epithelial cells were seeded in plates for Ma-CFC colony formation assay as described in supplementary method. Mammary tumors were harvested, minced and incubated in digestion media to generate single cell suspensions (described above) at 37°C for 2h. Cells were stained with CD49f and CD61 on ice for 30 min. CD61 was obtained from BioLegend (cat#104308).