LncRNA HCG18通过靶向miR-140-3p/PD-L1通路对鼻咽癌细胞生物学行为的影响

Effect of lncRNA HCG18 on biological behavior of nasopharyngeal carcinoma cells by targeting miR-140-3p/PD-L1pathway

目的 探讨长非编码(lncRNA)HCG18通过微小RNA-140-3p(miR-140-3p)/程序性死亡受体配体1(PD-L1)对鼻咽癌细胞增殖、迁移与侵袭的影响。方法 选择人鼻咽癌CNE2细胞、人正常鼻黏膜上皮细胞HNEpC细胞,将CNE2细胞分为HCG18组、pcDNA3.1组、sh-HCG18组、sh-NC组、miR-140-3p组、miR-NC组、miR-140-3p inhibitor组、Scramble组、HCG18+miR-NC组、HCG18+miR-140-3p组、miR-140-3p inhibitor+sh-NC组及miR-140-3p inhibitor+sh-PD-L1组。用CCK-8检测细胞增殖能力,Transwell实验检测细胞侵袭及迁移能力,Western blot检测PD-L1蛋白表达,实时荧光定量聚合酶链式反应(RT-qPCR)检测lncRNA HCG18、miR-140-3p及PD-L1 mRNA水平,生物信息学、双荧光素酶报告实验分析lncRNA HCG18、miR-140-3p及PD-L1的靶向关系。结果 CNE2细胞lncRNA HCG18、PD-L1蛋白及其mRNA表达水平高于HNEpC细胞,miR-140-3p表达水平低于HNEpC细胞(P <0.05)。上调lncRNA HCG18或下调miR-140-3p后CNE2细胞增殖、细胞迁移与侵袭能力增强,PD-L1蛋白、PD-L1 mRNA水平升高(P <0.05);下调lncRNA HCG18或上调miR-140-3p后CNE2细胞增殖、细胞迁移与侵袭能力降低,PD-L1蛋白、PD-L1 mRNA水平降低(P <0.05)。miR-140-3p与lncRNA HCG18、PD-L1均有靶向关系。上调miR-140-3p表达可逆转上调lncRNA HCG18对CNE2细胞增殖、迁移与侵袭的促进作用;沉默PD-L1可逆转下调miR-140-3p对CNE2细胞增殖、迁移与侵袭的促进作用。结论 过表达lncRNA HCG18可通过海绵化miR-140-3p,上调PD-L1表达,促进CNE2细胞增殖、迁移与侵袭。

Objective To investigate the effect of lncRNA HCG18 on proliferation,migration,and invasion of nasopharyngeal carcinoma cells via microRNA-140-3p(miR-140-3p)/programmed death receptor ligand 1(PD-L1).Methods Human nasopharyngeal carcinoma CNE2 cells and normal nasal mucosal epithelial cells HNEpC cells were selected.CNE2 cells were divided into HCG18 group,pcDNA3.1 group,sh-HCG18 group,sh-NC group,miR-140-3p group,miR-NC group,miR-140-3p inhibitor group,Scramble group,HCG18+miR-NC group,HCG18+miR-140-3p group,miR-140-3p inhibitor+sh-NC group and miR-140-3p inhibitor+sh-PD-L1 group.The CCK-8 was used to detect cell proliferation.Transwell test was used to detect cell invasion and migration.Western blot was used to detect PD-L1 protein expression,real-time fluorescence quantitative polymerase chain reaction(qRT-PCR)was used to detect levels of lncRNA HCG18,miR-140-3p and PD-L1 mRNA.The bioinformatics and dual luciferase report experiment were used to analyze targeting relationship of lncRNA HCG18,miR-140-3p and PD-L1.Results The levels of lncRNA HCG18,PD-L1 protein and mRNA in CNE2 cells were higher than those in HNEpC cells,while the level of miR-140-3p was lower than those in HNEpC cells(P<0.05).After up-regulation of lncRNA HCG18 or down-regulation of miR-140-3p,the proliferation,migration and invasion of CNE2 cells were enhanced,and levels of PD-L1 protein and PD-L1 mRNA were increased(P<0.05).After down-regulation of lncRNA HCG18 or up-regulation of miR-140-3p,the proliferation,migration and invasion of CNE2 cells were decreased,and levels of PD-L1 protein and PD-L1 mRNA were decreased(P<0.05).MiR-140-3p showed targeted relationship with lncRNA HCG18 and PD-L1.Up-regulation of miR-140-3p expression was reversed the effect of up-regulation of lncRNA HCG18 on proliferation,migration and invasion of CNE2 cells,and knockdown of PD-L1 reversed the effect of down-regulation of miR-140-3p on proliferation,migration and invasion of CNE2 cells.Conclusion It is demonstrated that lncRNA HCG18 overexpression could up-regulate the level of PDL1 via sponge forming miR-140-3p,promote the proliferation,migration,and invasion of CNE2 cells.