COX-2 upregulation in HIPK2 knockdown depends on HIF-1.

<p>(<b>A</b>) Representative RT-PCR analyses of control RKO-siRNA and RKO-siHIPK2 cells transfected with HIF-1α dominant negative (HIF-1αDN) for 36 h. The COX-2 and VEGF mRNA levels are shown. β-actin was used as internal control. (<b>B</b>) RT-PCR analyses of RKO-siHIPK2 cells were transfected with specific siRNA for HIF-1α knockdown or control siRNA. After transfection total RNA was extracted and RT-PCR was performed to assess HIF-1α, COX-2 and VEGF expression levels. β-actin was used as internal control. One representative experiment out of three independent experiments was shown. (<b>C, left panel</b>) RT-PCR analyses of RKO-siHIPK2 cells transfected with VHL expression vector (+) or with empty vector (-). After transfection total RNA was extracted and RT-PCR was performed to assess COX-2 and VEGF expression levels. β-actin was used as internal control. (<b>C, right panel</b>) Densitometric analysis of COX-2 and VEGF levels (as in the left panel) was performed and normalized values to β-actin mRNA levels were plotted. *P = 0.001. (<b>D</b>) 293 cells were transfected with HIF-1α expression vector (+) or with empty vector (−). After transfection total RNA was extracted and RT-PCR was performed to assess HIF-1α, COX-2 and VEGF expression levels. β-actin was used as internal control. One representative experiment out of three independent experiments was shown.