Figure 1.

Effects of brusatol on HIF-1α expression in colorectal cancer cells under hypoxia. (A and B) RKO (A) and HCT116 (B) cells were treated with different concentrations of brusatol (0-100 nM) for 1 h and exposed to 20% or 0.5% O₂. After a 4-h incubation, cells were harvested and whole-cell lysates were analyzed by immunoblotting for the indicated proteins. (C and D) RKO (C) and HCT116 (D) cells were incubated with or without 100 nM brusatol. After an 8 h exposure to 20% or 0.5% O₂, cells were harvested at the indicated times. (E and F) RKO (E) and HCT116 (F) cells were incubated with or without 100 nM brusatol for 1 h, exposed to 20% or 0.5% O₂ for 8 h, and then harvested. qPCR was used to amplify Glut1, PGK1, PDK1, and CA9; 18S rRNA was used as an internal control. Data are presented as means ± SD (*P < 0.05, **P < 0.01, ****P < 0.0001; ANOVA). (G and H) RKO (G) and HCT116 (H) cells were co-transfected with p3×HRE-luc and pCMV-β-galactosidase, cultured for 16 h, and then incubated with or without 100 nM brusatol for 1 h. Cells were then exposed to 20% or 0.5% O₂ for 8 h. Luciferase activity measured in whole-cell lysates was normalized to that of β-galactosidase. Data are presented as means ± SD (****P < 0.0001; ANOVA).