Characterization of side population cells isolated from the gastric cancer cell line SGC-7901

Side population (SP) cells are a subset of stem cells that have been isolated from several different gastrointestinal cancer cell lines. Using flow cytometry and the DNA-binding dye Hoechst 33342, we isolated SP cells from SGC-7901 human gastric tumor cell lines and found that they comprise 2.3±0.78% of the tumor cells. Using the Cell Counting Kit-8 (CCK-8) assay, we demonstrated that SP cells have a stronger proliferative activity than non-SP cells. Additionally, we observed tumor mass formation following the cultivation of SP cells in serum-free medium, indicating the capability of these cells for self-renewal. SP cells were observed to undergo non-symmetrical division, which is characteristic of stem cells. A drug resistance assay revealed that SP cells have a high survival rate when exposed to the chemotherapy drug 5-fluorouracil; the results of western blot analysis suggest that this stems from the abundant expression of the chemoresistance-associated proteins ABCG2 and Bcl-2. We also used fluorescence quantitative PCR to reveal that SP cells have relatively high expression levels of the stem cell-related genes Musashi-1 and CD44. In vivo experiments in mice revealed that the subcutaneous injection of 2×10(3) SP cells resulted in the formation of tumors, while the injection of 2×10(4) non-SP cells did not. Cumulatively, our results suggest that gastric tumorigenesis associated with SGC-7901 may partly be driven by the activity of SP cells, which exhibit certain biological characteristics of stem cells. Our results also show that the SP cell sorting method is an effective means for isolating and identifying gastric cancer stem cells during early screening.