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Updated: Oct 29, 2026
Isolation and Functional Assessment of Human Breast Cancer Stem Cells from Cell and Tissue Samples
Published on: October 2, 2020
Breast Cancer Side Population Versus CD44+/CD24- Cells: Functional and Tumorigenic Differences
Yukihiko Osawa1,2,3, Aya Kumazaki4, Konatsu Imanaka5
1Graduate School of Health Sciences, Department of Health Sciences, Kyoto Tachibana University, Kyoto, Japan; osawa-y@tachibana-u.ac.jp.
Background/Aim:
Cancer stem cells (CSCs) are heterogeneous and exhibit distinct phenotypic and functional characteristics. The side population (SP) and CD44+/CD24- phenotypes are widely used to enrich breast cancer cells with CSC-associated properties; however, whether these approaches identify functionally equivalent populations remains unclear. Building on our previous establishment of a DyeCycle Violet (DCV)-based method for isolating SP cells from MCF-7 breast cancer cells, this study compared the biological characteristics of DCV-defined SP cells and a CD44+/CD24--enriched fraction.
Materials And Methods:
SP cells were isolated from MCF-7 cells by fluorescence-activated cell sorting using DCV and fumitremorgin C. A CD44+/CD24--enriched fraction was isolated by magnetic-activated cell sorting. Parental MCF-7 cells, SP cells, and the CD44+/CD24--enriched fraction were compared for proliferation and invasion. Tumor-forming ability was evaluated in nude mice following subcutaneous inoculation of 100 or 500 cells. Xenograft tumors were examined histopathologically and immunohistochemically.
Results:
No significant differences in proliferative activity were observed among parental MCF-7, SP, and the CD44+/CD24--enriched fraction at either 48 or 72 h. Invasion was highest in the CD44+/CD24--enriched fraction (87.63±20.29%), followed by SP cells (51.86±6.83%) and parental MCF-7 cells (32.52±9.95%; p=0.001). Tumors developed in all three mice receiving the CD44+/CD24--enriched fraction at both cell doses, whereas tumor formation was less frequent in the other groups. Xenograft tumors showed morphological and immunohistochemical characteristics consistent with MCF-7 cells.
Conclusion:
DCV-defined SP cells and the CD44+/CD24--enriched fraction exhibited distinct biological characteristics. The CD44+/CD24--enriched fraction showed high invasive activity and frequent tumor formation, whereas SP cells showed relatively high proliferative activity and retained tumor-forming capacity under the experimental conditions examined. These findings suggest that SP and CD44+/CD24- enrichment methods select populations with different functional properties.
