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目的研究同源结构域相互作用蛋白激酶2(HIPK2)对非小细胞肺癌(NSCLC)细胞上皮间质转化(EMT)以及迁移侵袭的影响及机制。方法应用Lipofectamine 2000将HIPK2高表达质粒pc DNA3.1-HIPK2和对照质粒pc DNA3.1-Vector转染至A549、H520细胞,分别为高表达组和对照组;应用慢病毒转染HIPK2干扰质粒GV248 sh HIPK2和对照质粒GV248 sh Vector,在A549细胞中分别构建沉默组和对照组;在A549细胞沉默组中应用Lipofectamine 2000转染GV248 sh ZEB1质粒和GV248 sh Vector质粒,分别为干扰组和对照组。Western blotting检测HIPK2表达水平及其对EMT标记蛋白的影响;应用细胞划痕实验、Transwell实验、Matrigel实验检测细胞迁移、侵袭能力。结果与对照组相比,高表达组上皮细胞标记蛋白表达增强(P<0.01)、间质细胞标记蛋白表达下降(P<0.01),EMT受到抑制。高表达组迁移距离小于对照组(P<0.05),高表达组细胞迁移和侵袭至下室的细胞数也明显减少(P<0.01)。沉默HIPK2则促进EMT发生、增强细胞的迁移和侵袭。干扰锌指E盒结合蛋白(ZEB1)的表达可以缓解HIPK2沉默引起的EMT和迁移侵袭现象。结论 HIPK2具有抑制NSCLC细胞EMT和迁移侵袭的作用,其发生机制可能与转录因子ZEB1有关。
Objective To investigate the effect and mechanism of HIPK2 on epithelial-mesenchymal transition (EMT) and migration and invasion in non-small cell lung cancer (NSCLC). Methods The HIPK2 high expression plasmid pcDNA3.1-HIPK2 and the control plasmid pcDNA3.1-Vector were transfected into A549 and H520 cells by Lipofectamine 2000 respectively, and were transfected with HIPK2 plasmid GV248 sh HIPK2 and control plasmid GV248 sh Vector respectively, and silencing and control groups were constructed respectively in A549 cells. In A549 cell silencing group, GV248 sh ZEB1 plasmid and GV248 sh Vector plasmid were transfected by Lipofectamine 2000 respectively, which were interference group and control group respectively. Western blotting was used to detect the expression of HIPK2 and its effect on EMT marker protein. Cell scratch assay, Transwell assay and Matrigel assay were used to detect cell migration and invasion. Results Compared with the control group, the expression of epithelial marker protein increased (P <0.01) and the expression of interstitial marker protein decreased (P <0.01). EMT was inhibited in the high expression group. The migration distance of high expression group was less than that of control group (P <0.05), and the number of cells migrating and infiltrating into lower compartment of high expression group was significantly decreased (P <0.01). Silencing HIPK2 promotes EMT and enhances cell migration and invasion. Interfere with the expression of Zinc finger-box binding protein (ZEB1) can alleviate the EMT and migration-induced phenotypes induced by HIPK2 silencing. Conclusion HIPK2 can inhibit the EMT and migration of NSCLC cells, and its mechanism may be related to the transcription factor ZEB1.