RNA干扰ABCEl基因后可抑制肺癌95-D/NCI-H446细胞的增殖和诱导细胞凋亡
Suppression of ABCE1 Leads to Decreased Cell Proliferation and Increased Apoptosis in 95-D/NCI-H446 Lung Carcinoma Cells
查看参考文献19篇
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文摘
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ABCE1作为RNase L抑制剂首先是在脊椎动物中被发现的.前期研究结果显示ABCE1与肺腺癌的发生率及临床分期显著相关.为了进一步研究ABCE1的新功能,构建了ABCE1基因的siRNA表达质粒(RNAi-ReadypSIREN-DNR-DsRed-Express vector),培养肺癌细胞(95-D和NCI-H446),用FuGENE 6作为转染试剂转染后,使用荧光显微镜观察转染效果,RT-PCR分析ABCE1基因表达,Western blot分析ABCEl蛋白的表达,MTT法检测细胞的活性,流式细胞仪分析细胞周期,ELISA法检测细胞凋亡.结果显示:质粒的转染效果较满意,阳性率约为42.70%;在实验组,细胞活性和生长指数明显受到抑制,细胞凋亡明显增加,与对照组比较差异显著(P<0.05).上述结果显示,RNA干扰ABCE1基因可显著抑制肺癌细胞(95-D/NCI-H446)RNA的转录、蛋白质的表达,并增加细胞凋亡,为进一步研究ABCE1基因提供必要的基础. |
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其他语种文摘
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ATP-binding cassette protein E (ABCE1) has been annotated as an Rnase L inhibitor in eukaryotes. Previous study showed that the overexpression of ABCE1 was related with the occurrence and clinical stage of lung adenocarcinoma. As an initial investigation into the novel functions of ABCE1, siRNA-expressing vectors targeting sites of the ABCE1 gene were constructed from RNAi-Ready pSIREN-DNR-DsRed-Express vector. Cultured 95-D and NCI-H446 lung carcinoma cells were transfected with the siRNA-expressing vectors using FuGENE 6 and transfection efficiency was determined by using fluorescence microscopy. The expression level of ABCE1 protein was determined by Western blot and immunofluorescence staining. Cell viability was determined by MTT, cell cycle was analysed by flow cytometry.The apoptotic rate was observed by ELISA. Fluorescence microscopy showed a satisfactory transfection efficiency which was about 42.70%. Cell viability and the growth fraction were markedly suppressed, whereas the apoptosis was significantly increased in SiRNA-95-D and SiRNA-NCI-H446 cells than controls(P< 0.05). It can be concluded that the siRNA targeting ABCE1 gene shows a dramatic inhibitory effect on RNA transcription and protein expression and a promoting effect on the apoptosis in 95-D/NCI-H446 cells, which offers a reliable base for the further in vivo experiment. |
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来源
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生物化学与生物物理进展
,2009,36(11):1475-1482 【核心库】
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DOI
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10.3724/sp.j.1206.2009.00223
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关键词
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ABCE1
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凋亡
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肺癌细胞
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增殖
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RNA干扰
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地址
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1.
中国医科大学四院胸外科, 辽宁, 沈阳, 110032
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中国医科大学四院病理科, 辽宁, 沈阳, 110032
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中国医科大学一院胸外科, 辽宁, 沈阳, 110032
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语种
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中文 |
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文献类型
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研究性论文 |
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ISSN
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1000-3282 |
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学科
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分子生物学 |
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基金
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国家自然科学基金
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辽宁省教育厅高等学校科研项目专项基金
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文献收藏号
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CSCD:3716647
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