Figure 6
The effects of physapubescin on reducing cell viabilities, inducing apoptosis and modulating expression of related biomarkers are enhanced under hypoxia vs. normoxia conditions.
786-O cells were seeded at a density of 5 × 104 cells/well in six well plates under normoxic (21% O2), hypoxic (1% O2) or hypoxia mimic (250 μM CoCl2) conditions. (A) After 24 hours of seeding, cells were treated with 0.05% DMSO or physapubescin at the indicated concentrations for 72 hours. Cell densities were measured by MTT assay. Each value represents mean ± SEM of three samples for each treatment. (B) After 24 hours of seeding, the protein expression of HIF-1α, HIF-2α, DR5, and cleaved PARP at indicated treatments for 24 hours was analyzed by Western blotting. β-Tubulin was detected as a loading control. A representative blot was shown from three independent experiments. (C) After 24 hours of seeding, the secretion of VEGF in conditioned medium at indicated treatments for 24 hours was analyzed by ELISA. Each value represents mean ± SEM of three samples for each treatment. (D) RCC4/pcDNA3 and RCC4/VHL cells were seeded at a density of 5 × 104 cells/well in six well plates under normoxic (21% O2) or hypoxic (1% O2) conditions. After 24 hours of seeding, the secretion of VEGF in conditioned medium at indicated treatments for 24 hours was analyzed by ELISA. Each value represents mean ± SEM of three samples for each treatment. “*” and “**” denote “P < 0.05” and “P < 0.01” respectively.
