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目的:研究CSK结合蛋白(CSK-binding protein,CBP)基因转染对人肺鳞癌NCI-H520细胞体外生长侵袭的影响。方法:构建CBP真核表达质粒pcDNA3.0-CBP,以脂质体转染法转染体外培养的人肺鳞癌细胞株NCI-H520,G418筛选出抗性克隆。Western-blotting和RT-PCR分别检测转染前后CBP蛋白和mRNA水平的变化,MTT法分析细胞生长抑制作用,Transwell体外侵袭实验和Wound-healing实验观察细胞的侵袭和迁移能力。结果:稳定转染CBP基因的细胞株有外源目的基因的整合和相应蛋白的高表达。MTT检测表明,pcDNA3.0-CBP转染组活细胞数低于未转染组和pcDNA3.0(-)空载体质粒细胞转染组(P<0.01)。细胞侵袭、迁移实验表明转染pcDNA3.0-CBP的瘤细胞侵袭与迁移能力均明显下降(P<0.01)。结论:外源性CBP基因稳定转染可抑制人肺鳞癌NCI-H520细胞增殖、侵袭的恶性表型。
Objective: To investigate the effect of CSK-binding protein (CBP) gene transfection on the growth and invasion of human lung squamous cell carcinoma cell line NCI-H520 in vitro. Methods: The CBP eukaryotic expression plasmid pcDNA3.0-CBP was constructed and transfected into human lung squamous carcinoma cell line NCI-H520 by lipofection. The resistant clones were screened by G418. The levels of CBP protein and mRNA were detected by Western-blotting and RT-PCR before and after transfection. The cell growth inhibition, MTT assay, Transwell invasion assay and Wound-healing assay were used to detect the invasion and migration of CBP. Results: The cell lines stably transfected with CBP gene had the integration of exogenous gene and high expression of corresponding protein. MTT assay showed that the number of viable cells in pcDNA3.0-CBP transfection group was lower than that in non-transfection group and pcDNA3.0 (-) empty vector plasmid transfection group (P <0.01). Cell invasion and migration experiments showed that the invasion and migration ability of pcDNA3.0-CBP transfected cells were significantly decreased (P <0.01). CONCLUSION: Stable transfection of exogenous CBP gene can inhibit the malignant phenotype of human lung squamous cell carcinoma cell line NCI-H520 in proliferation and invasion.