Objective: To investigate the reactivity of colon cancer cell line SW480 and CD133(+) SW480 subsets to hypoxia in vitro and the changes in the expressions of anti-apoptosis and angiogenesis genes.
Methods: SW480 cells was subjected to CoCl(2) exposure at varying concentrations and for different time lengths to induce hypoxia, and the protein expression of hypoxia induced factor 1α (HIF-1α) was detected by Western blotting. The CD133(+) SW480 cells were sorted by magnetic activated cell sorting (MACS) and their proportion was assayed by flow cytometry (FCM). The CD133(+) SW480 subsets were exposed to CoCl(2) at the optimal concentration with exposure time selected in terms of HIF-1α level, and their tumor stem cell sphere formation ability was evaluated. Real-time PCR was used to compare the mRNA expression levels of the surface markers of colon cancer stem cells (CD133 and PROM1), survivin, and vascular endothelial growth factor (VEGF).
Results: Exposure to 200 µmol/L CoCl(2) for 8 h resulted in the highest HIF-1α expression in SW480 cells, but the same exposure failed to induce HIF-1α expression in CD133(+) SW480 subsets. The CD133(+) SW480 subsets, after CoCl(2)-induced hypoxia, showed significantly enhanced ability of cell sphere formation. Hypoxia of SW480 cells caused significant increases in CD133, survivin and VEGF mRNA levels by 1.607∓0.103, 2.745∓0.370 and 3.798∓0.091 folds, respectively (P<0.05).
Conclusion: CoCl(2) can simulate hypoxia in colon cancer cells in vitro to induce stable HIF-1α expression, which is concentration- and time-dependent. The hypoxia-stimulated tumor stem sells show an enhanced sphere formation and anti-apoptotic and anti-angiogenic abilities.