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目的:探讨VEGFR-3对人结肠癌细胞黏附力和侵袭性的影响。方法:构建携靶向VEGFR-3基因siRNA(small interfering RNA)表达载体,转染人结肠癌LoVo细胞,半定量RT-PCR和Western blotting检测转染前后LoVo细胞VEGFR-3 mRNA和蛋白表达的变化,基质-黏附实验检测细胞转染后的黏附能力,细胞侵袭实验检测转染后肿瘤细胞侵袭性的改变。结果:携靶向VEGFR-3基因siRNA的表达载体成功构建,RT-PCR检测转染siRNA后LoVo细胞VEGFR-3 mRNA表达水平降低;Western blotting检测转染siRNA后72 h LoVo细胞VEGFR-3蛋白表达下降,其表达相对值由(1.26±0.19)降至(0.39±0.12)(P<0.05)。转染siRNA 72 h后LoVo细胞的黏附能力显著下降[(0.626±0.047)vs(0.407±0.029),P<0.05];LoVo细胞穿膜细胞数(6.38±3.25)明显低于空白对照组(24.82±3.44)、非特异性对照组(23.58±3.73)(P<0.05)。结论:siRNA能够在LoVo细胞中引发RNA干扰效应,下调VEGFR-3基因的表达,进而抑制LoVo细胞的黏附能力和侵袭性。
Objective: To investigate the effect of VEGFR-3 on the adhesion and invasiveness of human colon carcinoma cells. Methods: The siRNA targeting small interfering RNA of VEGFR-3 gene was constructed and transfected into LoVo human colon cancer cells. The expression of VEGFR-3 mRNA and protein in LoVo cells was detected by semi-quantitative RT-PCR and Western blotting The adhesion ability of the cells after transfection was detected by matrix-adhesion assay. The cell invasion assay was used to detect the invasiveness of tumor cells after transfection. Results: The siRNA targeting VEGFR-3 gene was successfully constructed and the expression of VEGFR-3 mRNA was down-regulated in LoVo cells transfected with siRNA by RT-PCR. The expression of VEGFR-3 protein in LoVo cells was detected by Western blotting at 72 h Decreased from (1.26 ± 0.19) to (0.39 ± 0.12) (P <0.05). The adhesion ability of LoVo cells was significantly decreased after 72 h of transfection ([(0.626 ± 0.047) vs (0.407 ± 0.029), P <0.05]. The number of transmembrane cells in LoVo cells (6.38 ± 3.25) was significantly lower than that in the blank control group ± 3.44), nonspecific control group (23.58 ± 3.73) (P <0.05). Conclusion: siRNA can induce RNA interference in LoVo cells and down-regulate the expression of VEGFR-3 gene, thereby inhibiting the adhesion and invasiveness of LoVo cells.