Objective: To construct an expression plasmid of siRNA against activator protein 4(AP-4) gene and to investigate its biological behavioral effect on human colon carcinoma cell line SW480.
Methods: The specific siRNA target exon 7 of AP-4 gene was constructed and transfected into SW480 cells by liposome. Expression levels of AP-4 mRNA and protein from SW480 after transfection were examined by RT-PCR and Western blot respectively. Cell proliferation was detected by cell counting and MTT assay. Flow cytometry(FCM) and Western blot were used to detect cell apoptosis and cell cycle. The invasiveness of SW480 cells in vitro was measured quantitatively by the matrigel invasion assay.
Results: After AP-4 siRNA transfection into SW480 cells for 96 hours, the cell AP-4 mRNA reduced by 57.8% and the AP-4 protein concentration in culture supernatant decreased by 75.2%. The cell growth was significantly inhibited by 61%-78%. The apoptosis rate of SW480 cells was significantly higher in AP-4 siRNA group than that in negative control group[(21.7 +/- 2.51)% vs. (2.31 +/- 0.14)%, P<0.01]. Cells in G0-G1 phase increased by 22.43% and in G2-M phase decreased by 14.52% (P<0.05). AP-4 siRNA significantly inhibited AP-4-induced invasion of SW480 cells to matrigel (P<0.05).
Conclusions: Silencing AP-4 gene by the siRNA technology can actively suppress the expression of AP-4 gene, and then inhibit the growth and proliferation of SW480 cell, and induce apoptosis of SW480 cell. The successful application of AP-4 siRNA extends the list of available therapeutic modalities in the treatment of human colon cancer.