ABSTRACT
Objective:Enriching and isolating breast cancer stem cells from breast cancer transplantation tumors in nude mice.Methods:Human breast cancer MDA-MB-231 cells were injected into the right axilla subcutaneous of 20 nude mice, and the tumor growth was observed .After 30 days, tumors were isolated and stained with hematoxylin and eosin, and then tumor cells from tissues were isolated. DMEM medium containing serum was used to cultivate isolated transplantation tumor cells, cell morphology and growth were also observed. Flow cytometry was used to detect the proportion of stem cells (CD44 +/CD24 -/low cells) in transplantation tumor cells. Serum-free DMEM medium containing epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and B27 cell supplement were used to cultivate transplantation tumor cells and to obtain cell microspheres. The proportion of stem cells on the 10th day in cell microspheres was detected by using flow cell sorter and stem cells were isolated according to the markers of cell surface. Results:After subcutaneously injecting MDA-MB-231 cells into 20 nude mice for 9 days, 17 nude mice had subcutaneous tumors with more parenchymal cells, little interstitial cells, arranged cords tumor cells, large volume of the cell and abundant cytoplasm, the nuclei in different sizes and hyperchromatic state, mitotic more common, the nucleoli clear and obvious pleomorphy. After cultivating transplantation tumor cells with DMEM medium containing serum, the cells began to grow adherent after 24 h, and the adherent proportion rose to 60% after 3 days; after 7 days, the cell proliferation was accelerated; and the cell morphology was more consistent, most of which were spindle shaped and were not significantly different from MDA-MB-231 cells; the proportion of stem cells in transplantation tumor cells was (0.10±0.02)%. After cultivating transplantation tumor cells with serum-free DMEM medium containing cell cultured supplement, the cells grow in spherical patterns, the proportion of stem cells in cell microspheres got up to (70.47±2.03)% on the 10th day.Conclusions:Subcutaneously injecting MDA-MB-231 cells in nude mice can build breast cancer nude mice ectopic transplantation tumor model. Breast cancer stem cells in the transplantation tumors can be enriched from isolated transplantation tumor cells through serum-free medium, and more stem cells can be isolated to provide the research basis for the biological characteristics of breast cancer stem cells.