中华皮肤科杂志 ›› 2011, Vol. 44 ›› Issue (4): 263-266.

• 论著 • 上一篇    下一篇

RNA干扰抑制表皮生长因子受体表达对Colo-16细胞凋亡及化疗敏感性的影响

王慧1,魏志平2,刘彦群2   

  1. 1. 青岛市城阳区人民医院皮肤科
    2. 徐州医学院附属医院皮肤科
  • 收稿日期:2010-05-25 修回日期:2010-11-06 出版日期:2011-04-15 发布日期:2011-04-01
  • 通讯作者: 魏志平 E-mail:xzwzp1961@yahoo.com.cn

Effects of RNA interference targeting epidermal growth factor receptor on the apoptosis in and chemosensitivity of a human cutaneous carcinoma cell line Colo-16

  • Received:2010-05-25 Revised:2010-11-06 Online:2011-04-15 Published:2011-04-01

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摘要：

目的 探讨靶向表皮生长因子受体（EGFR）基因的短发夹RNA（EGFR-shRNA）对皮肤鳞状细胞癌Colo-16细胞生长和雷帕霉素敏感性的影响。方法 构建针对EGFR序列特异的shRNA表达载体，用脂质体转染Colo-16细胞。实验分正常细胞组、脂质体组、阴性对照组、阳性干扰组。分别采用免疫细胞化学和Western印迹测定EGFR蛋白的表达；噻唑蓝（MTT）法检测Colo-16细胞对雷帕霉素的敏感性；流式细胞仪检测细胞凋亡率。结果 EGFR-shRNA可明显抑制Colo-16细胞EGFR的表达，抑制率达43.3%（F = 44.6，P < 0.05），并将Colo-16细胞对雷帕霉素的敏感性提高2.44倍。阳性干扰组Colo-16细胞的凋亡率为（12.65 ± 0.091）%，与正常细胞组差异有统计学意义（F = 2042.9，P < 0.05）。结论 针对EGFR的质粒表达载体可以有效抑制Colo-16细胞EGFR的表达，增强Colo-16细胞对雷帕霉素的敏感性，并诱导其凋亡。

关键词: 雷帕霉素

Abstract:

Objective To investigate the effects of short hair RNA targeting epidermal growth factor receptor （EGFR-shRNA） on Colo-16 cell apoptosis and sensitivity to rapamycin. Methods The expression vector of EGFR-specific shRNA was constructed. Colo-16 cells were classified into 4 groups, normal control group remaining untreated, liposome group transfected with lipofectamine 2000, negative control group transfected with shRNA-NC/Lipofectamine 2000 and positive interference group transfected with the expression vector of shRNA-EGFR/Lipofectamine 2000. After additional culture, immunocytochemistry and Western blot were conducted to detect the protein expression of EGFR, and flow cytometry to measure the apoptosis in Colo-16 cells. MTT assay was performed to measure the sensitivity of Colo-16 cells to rapamycin. Results Compared with the normal control group, the expression of EGFR was down-regulated by 43.3% in positive interference group （F = 44.6, P < 0.05）, and the sensitivity to rapamycin was increased by 2.44 folds. The apoptosis rate in positive interference group was （12.65 ± 0.091）%, significantly different from that in the normal control group （F = 2042.9, P < 0.05）. Conclusion The plasmid expression vector containing shRNA targeting EGFR can effectively suppress the expression of EGFR by Colo-16 cells, enhance the sensitivity of Colo-16 cells to rapamycin and induce the apoptosis in Colo-16 cells.

Key words: rapamycin