Effect of Small Interfering RNA Targeting COX-2 on the Proliferation of Hepatocellular Carcinoma HepG2 Cell
|Keywords||Cyclooxygenase-2 Extracellular signal regulated protein kinase Small interfering RNA HepG2 cell|
BackgroundPrimary hepatic carcinoma (PHC) is one of the most common malignancies worldwide ,Recent studies have found that Cyclooxygenase-2 (COX-2) protein is highly expressed in hepatocellular carcinoma. It may be involved in tumorigenesis and development of PHC.In this study,we used small interfering RNA (siRNA) to silence COX-2 genetic expression,then observed the effects of the proliferation of hepatocellular carcinoma cell HepG2 and explored the possible mechanism. To provide the experiment evidences for the novel treatment approaches of PHC.ObjectiveTo explore the effects of the proliferation of hepatocellular carcinoma cell line HepG2 and ERK expression by siRNA decreasing COX-2 genetic expression.MethodsExperimental study based on HepG2 cell. HepG2 cell was subject to different treatments with four groups , that was COX-2 siRNA group,control siRNA group,lipo 2000 group and blank group. The Method of MTT (thiazolyl blue) was used to measure the proliferation inhibition rates of four groups cell after incubated 24 hours ,48 hours and 72 hours respectively. The changes of cell cycle were detected 24 hours after intervention by a flow cytometry. RT-PCR analysis was used to detect the RNA expression level of COX-2,ERK1/2 in four groups cell after incubated 24 hours respectively. Western blotting analysis was used to detect the protein expression level of COX-2,ERK1/2 in four groups cell after incubated 24 hours respectively.Results The results of MTT Experiment discovered this: The proliferation inhibition ratio of the COX-2 siRNA group was higher than these of the control siRNA and lipo 2000 group (P <0.01). There was no statistical difference between the control siRNA group and lipo 2000 group (P >0.01).In the COX-2 siRNA group , the inhibition ratio of after incubated 24 hours was higher than 48 hours and 72hours. Flow cytometry analysis showed that cell cycle of HepG2 cell treated with COX-2 siRNA blocked in the spot G1 (DNA presynthetic phase), which was significantly different from the control siRNA group,lipo 2000 group and blank group (P<0.05). The results of RT-PCR analysis discovered this: Compared to the control siRNA group,lipo 2000 group and blank group,the RNA content of COX-2,ERK1/2 in COX-2 siRNA group cell was significantly decreased after incubated 24 hours (P<0.05). The results of Western blotting analysis suggested this: Compared to the control siRNA group,lipo 2000 group and blank group.,the protein content of COX-2,ERK1/2 in COX-2 siRNA group cell was significantly decreased after incubated 24 hours (P<0.01).ConclusionsSiRNA could decreasing COX-2 genetic expression in hepatocellular carcinoma cell line HepG2 and inhibit the proliferation of HepG2. COX-2 promoting the development of PHC was possibly related to the regulation of ERK1/2 pathway.