沉默NEK2通过调节Wnt信号通路抑制非小细胞肺癌的细胞增殖

Silencing NEK2 Inhibits Cell Proliferation of Non-small Cell Lung Cancer by Regulating Wnt Signaling Pathway

目的探讨沉默有丝分裂相关激酶2(never in mitosis related kinase 2,NEK2)对非小细胞肺癌(non-small cell lung cancer,NSCLC)细胞生物学行为的影响及其可能机制。方法收集27对NSCLC和邻近的正常组织,免疫组化检测组织NEK2表达,收集临床病理数据。生物信息学分析研究NEK2在肺腺癌中的表达和预后价值。通过NCI-H1299和A549细胞体外转染siRNA敲低NEK2,通过免疫印迹及qPCR实验验证敲低效果,CCK-8评估细胞增殖能力,流式细胞术评估细胞凋亡及周期,蛋白印迹测定Wnt信号通路相关蛋白表达。结果在肺腺癌组织中的NEK2表达显著高于癌旁组织,ⅡB-ⅢB级组显著高于ⅠA-ⅡA级组,从TCGA数据库获得的数据也显示肺腺癌患者的NEK2明显高于相应的对照组,并与预后不良显著相关。转染NEK2 siRNA显著降低了NCI-H1299和A549细胞的增殖,细胞阻滞在G0/G1期,细胞凋亡率增加。此外,转染NEK2 siRNA的NCI-H1299和A549细胞中Wnt和β-catenin的表达水平显著下调,而pGSK3β的表达水平上调。结论siRNA干扰NEK2的表达可抑制非小细胞肺癌细胞的增殖,其机制可能是通过抑制Wnt/β-catenin通路的促进细胞凋亡和周期阻滞。

Objective To explore the effect of silencing NEK2 on the biological behavior of non-small cell lung cancer(NSCLC)cells and its possible mechanism.Methods A total of 27 pairs of NSCLC and adjacent normal tissues were collected,the expression level of NEK2 in those tissues were detected immunohistochemically,and the clinicopathological data were collected.Bioinformatics methods were performed to analyze the expression level and the prognostic value of NEK2 in lung adenocarcinoma.NCI-H1299 and A549 cells were transfected with siRNA to knock down the expression of NEK2 in vitro,and the knockdown efficiency was assessed by Western blotting and qPCR experiments.CCK-8 was used to measure the proliferation rate of cells,and flow cytometry was used to detect the cell apoptosis and cell cycle.Western blotting was used to detect the expression levels of Wnt signaling pathway related proteins.Results The expression level of NEK2 in lung adenocarcinoma tissues was significantly higher than that in the adjacent tissues,and its expression level in theⅡB-ⅢB group was significantly higher than that in the IA-ⅡA group.The data obtained from the TCGA database also showed that the expression level of NEK2 in patients with lung adenocarcinoma was significantly higher than that in patients of the corresponding control group,and that its high expression level was significantly associated with a poor prognosis.Transfection of NEK2 siRNA significantly inhibited the proliferation and enhanced the apoptosis of NCI-H1299 and A549 cells,made the cells been arrested in the G0/G1 phase.In addition,the expression levels of Wnt andβ-catenin were significantly lower in the NCI-H1299 and A549 cells transfected with NEK2 siRNA,while the expression level of p-GSK3βwas higher.Conclusion Silencing of NEK2 by siRNA can inhibit the proliferation of non-small cell lung cancer cells,the mechanism may relate to the promotion of cell apoptosis and cell cycle arrest through the inhibition of the Wnt/β-catenin pathway.