Adenovirus vector-mediated RNA interference inhibits expression of vascular endothelial growth factor receptor in colon carcinoma cells and tumor growth / 第二军医大学学报

ABSTRACT

Objective: To study the inhibitory effects of an adenovirus (Ad)-based short hairpin RNA (shRNA) expression system on expression of vascular endothelial growth factor receptor (VEGFR) and on growth of colon carcinoma cells in vitro and in vivo. Methods: RNA interference pAd-Easy/VEGFR vector was used to transfect 293 cells via Lipofectamine 2000. The adenoviral vector carrying VEGFR was used to transfect CW2 cells and the transfection efficiency was determined by fluorescent microscope and flow cytometry. The expression of VEGFR was examined by RT-PCR and Western blotting. The cell growth was observed with MTT method and the growth curve was plotted. Nude mouse was transplanted with CW2 cells to establish colon cancer tumor model and the growth of tumor was observed daily. Micro-vessel density (MVD) was detected by immunohistochemistry. Results: The recombinant pAd-Easy carrying shRNA against VEGFR was verified by sequencing. Flow cytometry showed that the transfection efficiency of CW2 cells was 99.7%. RT-PCR and Western blotting showed that the expression of VEGFR in pAd-Easy/VEGFR group was obviously decreased. The growth curve showed that the cell growth in the pAd-Easy/ VEGFR group was obviously slowed down. We also found that the tumor growth in the nude mouse model was obviously inhibited and the MVD was also decreased. Conclusion: pAd-Easy/VEGFR-mediated VEGFR shRNA can effectively inhibit the expression of VEGFR in CW2 cells and suppress tumor growth in vivo.