Fig. 3

HIF-1α enhances ANRIL expression in osteosarcoma cells. a Osteosarcoma cells were transfected with HRE-mutant ANRIL promoter constructs. ANRIL promoter activity was detected by luciferase assay (**, P < 0.01, n = 3). b HIF-1α mRNA levels were analyzed by real-time PCR after siRNA knockdown of HIF-1α. β-actin was used as an internal control (**, P < 0.01, n = 3). c HIF-1α protein levels were analyzed by Western blotting after HIF-1α knockdown. β-actin was used as the internal control. (**, P < 0.01, n = 3). d Effect of HIF-1α silencing on ANRIL promoter activity was detected by luciferase assay (**, P < 0.01, n = 3). e Effect of HIF-1α silencing on ANRIL expression was detected by real-time PCR. β-actin was used as an internal control (**, P < 0.01, n = 3). f HIF-1α mRNA levels were analyzed by real-time PCR after treatment with YC-1 HIF-1α-specific inhibitor (40 μM). β-actin was used as an internal control (**, P < 0.01, n = 3). g HIF-1α protein levels were analyzed by Western blotting after treatment with YC-1 (40 μM). β-actin was used as the internal control (**, P < 0.01, n = 3). h Effect of YC- 1 on ANRIL expression was detected by real-time PCR. β-actin was used as an internal control (**, P < 0.01, n = 3)