背景与目的:已证实非小细胞肺癌中表皮生长因子受体(epidermal growth factor receptor,EGFR)的过度表达和活化与肿瘤分期、放化疗敏感度下降密切相关。许多阻滞和下调EGFR的方法被用来抑制肿瘤增殖以改善预后,EGFR目前成为公认的肿瘤基因治疗的重要靶点。本研究采用靶向EGFR的短发夹RNA(short hairpinRNA,shRNA)表达重组质粒,观察能否在人肺腺癌细胞株A549细胞中引发RNA干扰(RNAi)效应及其对A549细胞生长及药物敏感性的影响。方法:构建靶向EGFR的shRNA重组质粒(pShEGFR)和非特异性的shRNA重组质粒(pShNEG),阳离子脂质体将其转染至肺腺癌A549细胞分别命名为A549/pShEGFR、A549/pShNEG和对照组A549,免疫荧光和Western blot法检测转染后细胞中EGFR表达变化;克隆形成实验观察细胞生长变化;流式细胞仪检测细胞周期和凋亡;MTT法检测细胞对顺铂、多柔比星、紫杉醇的敏感性。结果:与对照组A549细胞和A549/pShNEG相比,A549/pShEGFR可显著抑制A549细胞中EGFR表达,转染后第6天抑制率达74.1%(P<0.01);A549/pShEGFR组细胞克隆形成抑制率为62.8%。与对照组A549细胞和A549/pShNEG相比,A549/pShEGFR组细胞阻滞在G0/G1期(63.2±1.1,64.5±1.4vs.74.2±0.8;P<0.01),凋亡细胞比例显著增加(5.8±1.4,7.7±1.2vs.25.6±1.2;P<0.01);与A549组相比,A549/pShEGFR可将A549细胞对顺铂、多柔比星、紫杉醇的敏感性分别提高6.7、5.5、6.6倍。结论:瞬时转染靶向EGFR基因的shRNA表达重组质粒能够通过下调EGFR蛋白水平抑制肺腺癌A549细胞生长,并提高细胞对不同化疗药物的敏感性。 BACKGROUND & OBJECTIVE: It has been demonstrated that the overexpression and activation of epidermal growth factor receptor (EGFR) in non-small cell lung cancer are closely related to tumor stage and decreased sensitivity to radiochemotherapy. Many methods that block and downregulate EGFR are used to inhibit tumor proliferation and improve prognosis. EGFR is now recognized as an important target for tumor gene therapy. In this study, short hairpin RNA (shRNA) targeting EGFR was used to express recombinant plasmids to observe whether RNA interference (RNAi) effect could be induced in human lung adenocarcinoma cell line A549 and its sensitivity to A549 cell growth and drug sensitivity. The impact of sex. METHODS: The shRNA recombinant plasmid (pShEGFR) targeting EGFR and the non-specific shRNA recombinant plasmid (pShNEG) were constructed and transfected into lung adenocarcinoma A549 cells by cationic liposome. They were named A549/pShEGFR, A549/pShNEG and control respectively. In group A549, the expression of EGFR in the transfected cells was detected by immunofluorescence and Western blot; cell growth was observed by cloning formation assay; cell cycle and apoptosis were detected by flow cytometry; cells were assayed by cisplatin and doxorubicin by MTT assay. Paclitaxel sensitivity. RESULTS: Compared with A549 cells and A549/pShNEG in control group, A549/pShEGFR significantly inhibited the expression of EGFR in A549 cells, and the inhibition rate reached 74.1% on the 6th day after transfection (P<0.01); the cell clones were formed in A549/pShEGFR group. The inhibition rate was 62.8%. Compared with control A549 cells and A549/pShNEG cells, A549/pShEGFR group cells arrested at G0/G1 phase (63.2±1.1, 64.5±1.4 vs. 74.2±0.8; P<0.01), and the percentage of apoptotic cells increased significantly ( 5.8±1.4,7.7±1.2vs.25.6±1.2; P<0.01); Compared with A549 group, A549/pShEGFR can increase the sensitivity of A549 cells to cisplatin, doxorubicin and paclitaxel by 6.7, 5.5, respectively. 6.6 times. CONCLUSION: Transfection of shRNA expression recombinant plasmid targeting EGFR gene can inhibit the growth of lung adenocarcinoma A549 cells by down-regulating the EGFR protein level, and increase the sensitivity of cells to different chemotherapeutic drugs.