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背景与目的:Layilin是一种新发现与肺腺癌侵袭转移有密切联系的特异性透明质酸受体。本研究旨在观察缺氧/放射条件下活化缺氧/放射双敏感性增强子(Epo/e9),调控针对layilin的小干扰RNA(small interfering RNA,siRNA)表达,探讨其对人肺腺癌A549细胞受透明质酸诱导侵袭的影响。方法:构建携带Epo/e9增强子和U6启动子且表达针对layilin的siRNA的RNA干扰载体(Epoo/e9-siLay plasmid)和阴性对照RNA干扰载体(Epo/e9-siCtrl plasmid),分别转染A549细胞,新霉素抗性筛选得到layilin表达受抑制的A549_(Epo/e9-siLay)细胞和layilin表达未受影响的A549_(Epo/e9-siCtrl)细胞。针对非转染A549(A549_(untransfected))、A549_(Epo/e9siLay)、A549_(Epo/e9-siCtrl)三组细胞,在缺氧/放射处理下,分别采用RT-PCR和和Western blot检测layilin mRNA和和蛋白表达,用Transwell模型检测细胞侵袭能力。结果:与A549_(untransfected)组相比,A549_(Epo/e9-siLay)组layilin表达显著减弱(P<0.01),Transwell穿膜细胞数显著减少(P<0.01);A549_(Epo/e9-siCtrl)组layilin表达、Transwell穿膜细胞数皆和A549_(untransfected)组无显著差异(P>0.05)。缺氧/放射处理能进一步增加和RNA干扰的上述效应(P<0.01)。结论:在缺氧/放射条件下,Epo/e9增强子调控layilin siRNA表达能明显抑制A549细胞侵袭行为。
BACKGROUND & PURPOSE: Layilin is a newly discovered specific hyaluronic acid receptor closely related to the invasion and metastasis of lung adenocarcinoma. The purpose of this study was to observe the expression of small interfering RNA (siRNA) targeting layilin in hypoxia / radiation-stimulated hypoxia / radiation double sensitivity enhancer (Epo / e9) Effect of hyaluronic acid on invasion of A549 cells. METHODS: RNA interference vectors (Epoo / e9-siLay plasmid) and Epo / e9-siCtrl plasmid carrying Epo / e9 enhancer and U6 promoter and siRNA against layilin were constructed and transfected into A549 A549_ (Epo / e9-siLay) cells in which layilin expression was inhibited and A549_ (Epo / e9-siCtrl) cells in which layilin expression was not affected were screened by neomycin resistance. The expression of layilin was detected by RT-PCR and Western blot respectively in three groups of cells transfected with A549 (A549 untransfected), A549_ (Epo / e9siLay) and A549_ (Epo / e9-siCtrl) mRNA and protein expression, using the Transwell model of cell invasion. Results: Compared with A549 untransfected group, layilin expression in A549_ (Epo / e9-siLay) group was significantly decreased (P <0.01) and transwell transmembrane cell number was significantly decreased (P <0.01). A549_ (Epo / e9-siCtrl ) Group layilin expression, Transwell number of transmembrane cells and A549 untransfected group was no significant difference (P> 0.05). Hypoxia / radiation treatment further increased the aforementioned effects of RNA interference (P <0.01). Conclusion: Under the condition of hypoxia / radiation, the expression of layilin siRNA by Epo / e9 enhancer can significantly inhibit the invasion of A549 cells.