| RNA干扰靶向沉默HER2/neu基因对卵巢癌细胞株SK-OV-3的影响 |
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| 引用本文： | 衣翠华,侯明,李丽珍,张昕,魏军民,黎莉,郝静. RNA干扰靶向沉默HER2/neu基因对卵巢癌细胞株SK-OV-3的影响[J]. 山东大学学报(医学版), 2006, 44(10): 1048-1052 |
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| 作者姓名： | 衣翠华 侯明 李丽珍 张昕 魏军民 黎莉 郝静 |
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| 作者单位： | 山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012;山东大学齐鲁医院肿瘤防治研究中心,山东,济南,250012 |
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| 摘 要： | 目的：观察siRNA表达质粒稳定转染靶向沉默HER2/neu基因对SK OV 3卵巢癌细胞株的影响。方法：针对HER2/neu基因序列构建短发夹状siRNA真核表达载体(pGenesil 1 HER2/neu)，转染卵巢癌细胞株SK OV 3，G418筛选出稳定转染株后，采用RT PCR和Western Blot法观察HER2/neu基因的沉默效果；CCK 8比色分析检测细胞体外增殖活力；流式细胞仪检测细胞周期。结果：测序证实成功构建HER2/neu的2个短发夹状siRNA真核表达质粒；稳定转染SK OV 3细胞后，转染了特异性HER 2/neu siRNA的细胞HER 2/neu mRNA及p185蛋白水平均明显低于亲本细胞及转染无义对照序列的细胞；CCK 8比色分析显示HER 2/neu基因沉默的细胞存活分数明显低于HER 2/neu基因高表达的细胞（P=0.000）；流式细胞仪细胞周期分析也显示，HER 2/neu基因沉默的细胞处于凋亡状态及非增殖期的比例明显增加（P均<0.01），而处于合成期的比例却显著减少（P≤0.001）。结论：靶向HER2/neu基因的短发夹状siRNA可从转录及翻译水平有效地沉默HER 2/neu基因；HER 2/neu基因沉默的SK OV 3卵巢癌细胞增殖明显受抑制。
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| 关 键 词： | RNA干扰 HER2/neu基因 卵巢癌 基因疗法 |
| 文章编号： | 1671-7554（2006）10-1048-05 |
| 收稿时间： | 2006-07-06 |
| 修稿时间： | 2006-07-06 |
Effects of RNAi on HER2/neu gene and proliferation of SK-OV-3 ovarian cancer cells in vitro |
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| YI Cui-hua,HOU Ming,LI Li-zhen,ZHANG Xin,WEI Jun-min,LI Li,HAO Jing. Effects of RNAi on HER2/neu gene and proliferation of SK-OV-3 ovarian cancer cells in vitro[J]. Journal of Shandong University:Health Sciences, 2006, 44(10): 1048-1052 |
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| Authors: | YI Cui-hua HOU Ming LI Li-zhen ZHANG Xin WEI Jun-min LI Li HAO Jing |
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| Affiliation: | Tumor Research Center, Qilu Hospital of Shandong University, Jinan 250012, Shandong, China |
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| Abstract: | To investigate the effects of HER2/neu gene silencing on SK OV 3 ovarian cancer cells by RNA interference. Methods： Design and synthesize two siRNAs based on the sequence of HER2/neu mRNA. They were separately subcloned into the plasmid of pGenesil 1 containing U6 promoter. The pGenesil 1 vectors of the RNA interference eukaryotic expression specific to HER2/neu gene were constructed. The pGenesil 1 vectors were transfected into SK OV 3 ovarian cancer cells with METAFECTENE in vitro. The positive cell clones were obtained after being screened with 250?μg/ml G418. Then the effects of RNAi on the expression of HER2/neu gene were detected by RT PCR and Western Blot. The cellular growth activities were assayed by CCK 8 colorimetry and flow cytometry in vitro. Results： DNA sequencing of the plasmids verified the successful construction of the HER2/neu siRNA vectors. Compared with the blank and the control group, the expressions of HER2/neu mRNA and protein 185 were remarkably down regulated in the RNAi groups; and the cell proliferations decreased in the RNAi groups. Conclusion： We successfully construct the recombinant plasmids of siRNAs specific to HER2/neu gene and transfect them into the SK OV 3 ovarian cancer cells in vitro. The RNAi inhibits the expression of HER2/neu mRNA and protein 185, and subsequently suppresses the proliferation of SK OV 3 cells. |
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| Keywords: | RNA interference HER2/neu gene Ovarian cancer cell Gene therapy |
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