Objective: To isolate and identify breast cancer initiating cells (CD44(+)/CD24(-/low) cells) in breast cancer cell lines MCF-7 and MDA-MB-231, and to evaluate the activity of the Hedgehog signaling pathway in different subpopulations.
Methods: CD44+/CD24(-/low) subpopulation and non-CD44+/CD24(-/low) subpopulation from breast cancer cell lines of MCF-7 and MDA-MB-231 were separated by fluorescence-activated cell sorting (FACS). Laser Scanning Confocal Microscope was used to identify CD44+/CD24(-/low) cells. Reverse transcription-polymerase chain reaction (RT-PCR) was employed to analyze the expression of Human Patched gene (PTCH), Sonic Hedgehog (SHH), glioma-associated oncogene homoglog-1(GIi-1), smoothened homolog (SMOH) and GAPDH of CD44+/CD24(-/low) subpopulation and non-CD44+/CD24(-/low) subpopulation from MCF-7 and MDA-MB-231.
Results: The CD44+/CD24(-/low) subpopulation in MCF-7 and MDA-MB-231 cell lines were (1.70+/-1.43)% and (94.2+/-1.2)% respectively. The Hedgehog signaling pathways were active in CD44+/CD24(-/low) subpopulation of MCF-7 and MDA-MB-231 cell lines. The expression of transcription factor GIi-1, which was downstream components of the hedgehog pathway, was assayed and the results showed that the level of GIi-1 mRNA of CD44+/CD24(-/low) cells was much higher than that of non-CD44+/CD24(-/low) cells (P = 0.007 in MCF-7, and 0.005 in MDA-MB-231).
Conclusion: Breast cancer initiating cells (CD44+/CD24(-/low) subpopulation) exist in MCF-7 and MDA-MB-231 cell lines. The Hedgehog signaling pathway is active in the subpopulation of MCF-7 and MDA-MB-231 cell lines.