【摘要】 为了研究膜联蛋白A2(ANXA2)基因表达水平与人肝癌细胞生物学行为之间的关系,采用已优化转染条件的磷酸钙法将针对ANXA2的siRNA重组质粒导入人肝癌细胞SMMC-7721并观察对靶基因表达及细胞生物学行为的影响。以半定量RT-PCR和Western blotting检测转染后不同时间(24h、48h、72h和96h)ANXA2mRNA及蛋白表达变化;以MTT法、Hoechast33258染色及体外损伤修复实验等分析抑制ANXA2表达后对SMMC-7721细胞生物学行为的影响。结果显示:所设计的四条siRNA序列均不同程度抑制了ANXA2的表达(p<0.05),且呈时间依赖性;细胞生长能力及运动能力被显著抑制(p<0.05),其抑制效应与ANXA2表达敲低程度呈正相关。结论:ANXA2的高表达水平与肝癌细胞的凋亡、生长及运动能力密切相关,以RNAi方法抑制该基因的表达可以有效地抑制肝癌细胞的恶性生物学行为。上述结果为肝癌的实验性基因治疗研究提供了新的思路和对策。更多还原

【Abstract】 Objective:Investigate the correlation between ANXA2 and Hepatocellular Carcinoma(HCC)proliferation,apoptosis and motility.Method:Four siRNAs recombinants targeting to ANXA2 were introduced into SMMC-7721 cells using an optimized calcium phosphate transfection method.ANXA2 mRNA and protein levels were evaluated by using semi-quantitative RT-PCR and Western blotting.The cell proliferation,motility,and apoptosis were determined with MTT,wound healing assay,and Hoechast33258 staining methods.Results:Four siRNAs suppressed ANXA2 expression effectively with time dependence(p<0.05).Cell proliferation and motility were inhibited after ANXA2 knockdown(p<0.05),and the inhibitory efficiency was positively correlated with ANXA2 knockdown level.Conclusion:It is concluded that the over-expression of ANXA2 in HCC is closely associated with the cell malignant behaviors,and siRNA can inhibit the ANXA2 expression efficiently in SMMC-7721 cells.更多还原