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浙江大学学报(医学版)  2012, Vol. 41 Issue (3): 281-288    DOI: 10.3785/j.issn.1008-9292.2012.03.009
论著     
RNA干扰突变型p53基因对胃癌SGC7901细胞生物学行为的影响
李贵玲,张钧,周艳文,王燕忠,何晓红,谢鑫友
浙江大学医学院附属邵逸夫医院检验科,浙江 杭州 310016
Effect of siRNA for mutant p53 gene on biological behavior of gastric cancer cell line SGC7901
LI Gui-ling,ZHANG Jun,ZHOU Yan-wen,WANG Yan-zhong,HE Xiao-hong,XIE Xin-you
Department of Clinical Laboratory,Sir Run Run Shaw Hospital,Zhejiang University School of Medicine,Hangzhou 310016,China
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摘要: 目的:利用RNA干扰(RNAi)技术稳定沉默人胃癌SGC7901细胞中的突变型p53基因,观察其对细胞生物学行为的影响。
方法:p53小干扰RNA(siRNA)重组质粒用脂质体介导法转染SGC7901细胞,另设未转染组和对照组。RT-PCR和Western blot分别检测p53基因mRNA和蛋白的表达,MTT法检测细胞增殖情况及其对奥沙利铂药物敏感性的变化,流式细胞仪检测细胞周期分布和凋亡,平板克隆形成试验、裸鼠移植瘤成瘤实验分析细胞成瘤性变化。
结果:p53 siRNA可显著下调SGC7901细胞中突变型p53基因mRNA和蛋白的表达,实验组较未转染组和对照组细胞生长曲线左移。实验组的G0/G1期细胞数较对照组和未转染组分别减少7.4%和6.7%,进入S期的细胞数增加17.2%,奥沙利铂诱导的细胞凋亡率明显降低,未转染组、对照组和实验组细胞的IC50分别为15.88μmol/L、14.32 μmol/L和20.34 μmol/L。与未转染组和对照组相比,实验组细胞平板克隆形成数量显著增多,裸鼠体内细胞成瘤性增加。
结论:p53 siRNA可有效抑制突变型p53基因在胃癌SGC7901细胞中的表达,但利用RNA干扰技术靶向突变型p53基因在胃癌的治疗上尚存在不确定性。
关键词: 胃肿瘤/病理学 转染 细胞增殖 细胞凋亡 基因p53 RNA干扰 基因疗法 肿瘤细胞培养的 基因表达    
Abstract: Objective: To investigate the effects of silencing the expression of mutant p53 gene with RNA interference technique on biological behavior of gastric cancer SGC7901 cells.
Methods: Recombinant plasmid of mutant p53 gene-targeting siRNA was transfected into gastric cancer SGC7901 cells by LipofectamineTM2000.The expressions of mutant p53 gene mRNA and protein were identified by RT-PCR and Western blot assay.The proliferation of SGC7901 cells and changes of Oxaliplatin (OXA) drug sensitivity were detected by MTT assay.The cell cycle distribution and apoptosis rate were analyzed by flow cytometry.Changes in cell tumorigenicity ability were analyzed by colony formation assay and xenograft tumor formation in nude mice.
Results: The expression of mutant p53 in SGC7901 cells was remarkable suppressed by mutant p53-siRNA.The cell growth curve of the transfected group turned to left compared to untransfected group and control group.The cell number of G0/G1 phase of transfected group was decreased by 7.4% and 6.7% respectively,and that of S phase was increased both by 17.2% compared to control group and untransfected group,and the cell apoptosis rate induced by Oxaliplatin was remarkable decreased.The IC50 of OXA in untransfected group,control group and transfected group were 15.88 μmol/L,14.32 μmol/L and 20.34 μmol/L respectively.The colony formation rate and tumorigenicity ability in nude mice of transfected group increased remarkably.
Conclusion: Mutant p53-siRNA can significantly inhibit the expression of mutant p53 in SGC7901 cells,however,the use of RNA interference targeting mutant p53 gene in the treatment of gastric cancer is still uncertain.
Key words: Stomach neoplasms/pathology; Transfection; Cell proliferation; Apoptosis; Genes    p53; RNA interference; Gene therapy; Tumor cells    cultured; Gene expression
出版日期: 2012-05-25
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李贵玲;张钧;周艳文;王燕忠;何晓红;谢鑫友. RNA干扰突变型p53基因对胃癌SGC7901细胞生物学行为的影响[J]. 浙江大学学报(医学版), 2012, 41(3): 281-288.

链接本文:

https://www.zjujournals.com/med/CN/Y2012/V41/I3/281

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