关键词: 发夹状小分子干扰RNA（shRNA）, VEGF, 人皮肤鳞癌细胞（A431细胞）

Abstract: 【Abstract】Objective The eukaryotic expression plasmids targeting VEGF (psilencer-VEGF1-shRNA，VEGF-s1；psilencer-VEGF2-shRNA，VEGF-s2)were constructed and transfected these plasmids to the human skin squamous cell carcinoma cell line (A431) to observe its affect to a series of biological characteristics of A431 cells for exploring the significance of intervention by VEGF-s1 and VEGF-s2. Method: VEGF-s1,VEGF-s2 and random target sequence of the negative control plasmid (psilencer-Target-off-shRNA，T-off)were constructed, simultaneously. and transfected these plasmids to A431 by Tm2000. The expression of VEGF mRNA and protein were detected by RT-QPCR, western blot and ELISA; the vitality of A431 cells was examined by CCK-8 assay; the percentage of Cell Proliferative Cycle and apoptosis of A431 were tested by flow cytometry; the migration capacity of A431 in two-dimensional and three- dimensional space were inspected by wound healing effect and Transwell assay,respectively; the adhesion potential of A431 was detected by FN adhesion test. Results: After VEGF-s1 or VEGF-s2 treatment, cytoactivity declined, cell cycle arrested, G1 cell proportion increased sharply, S cell proportion decreased obviously, apoptosis increased, the expression of VEGF mRNA and protein decreased significantly, the migration and adhesion of A431 cells was suppressed significantly, as compare with the control group and untreated cells(every ,p<0.05). Conclusion: VEGF is an important growth-related gene of human skin squamous cell carcinoma, silencing VEGF could effectively inhibit the biological behavior of A431 cells.

Key words: short hairpin RNAi, VEGF, human skin squamous cell(A431 cells)