microRNA-214在晚期下咽鳞状细胞癌中的表达及作用机制

Expression and effect of microRNA-214 in advanced hypopharyngeal carcinoma

目的 研究microRNA-214（miR-214）在晚期下咽鳞状细胞癌（鳞癌）组织中的表达情况,及其对下咽癌FaDu细胞系侵袭、迁移、克隆形成能力的影响,并分析其可能机制.方法 利用qRT-PCR技术检测30例晚期下咽鳞癌患者肿瘤组织及正常黏膜组织中miR-214的表达;通过转染过表达载体hsa-mir-214上调FaDu细胞中miR-214的表达;采用Transwell侵袭、迁移实验和平板克隆实验,分别检测miR-214表达上调后对FaDu细胞侵袭、迁移及克隆形成能力的影响;利用Western blot检测上调miR-214表达对Twist蛋白表达的影响.结果 miR-214在晚期下咽鳞癌组织中相对表达（（x）±s）为（0.311±0.206）明显低于下咽正常黏膜组织中的相对表达为（1.620±1.394）,差异有统计学意义（=5.09,P＜0.05）.实验组FaDu细胞转染hsa-mir-214后,miR-214的表达较阴性对照组明显上调（t=6.347,P＜0.05）.Transwell迁移和侵袭实验显示,实验组FaDu细胞迁移能力及侵袭能力均明显低于对照组（t=11.6,P＜0.01;t=6.499,P＜0.05）.平板克隆实验显示,实验组与对照组相比,平均克隆形成数及克隆形成率差异均无统计学意义（t=0.592,P＞0.05）,miR-214对FaDu细胞增殖能力无明显影响.Western blot结果显示,miR-214上调后的实验组细胞Twist蛋白的表达明显低于对照组细胞中的表达（t=6.545,P＜0.05）.结论 miR-214在晚期下咽鳞癌组织中低表达,且可明显抑制FaDu细胞的侵袭、迁移能力,其作用机制可能与抑制Twist蛋白表达有关.miR-214对FaDu细胞增殖能力无明显影响.

Objective To investigate the expression of microRNA-214 （miR-214） in advanced hypopharyngeal carcinoma tissues and its effects on the invasion,migration and colone formation of FaDu cells.Methods miR-214 expression in 30 cases of advanced hypopharyngeal carcinoma tissues and normal hypopharyngeal mucosa tissues was detected by real-time quantitative polymerase chain reaction （RT-qPCR）.miR-214 was upregulated through transfecting the overexpression vector hsa-mir-214 into FaDu cells.The influences of miR-214 upregulation on the invasion,migration,clone formation and Twist expression were measured by Transwell invasion,Transwell migration,plate clone formation and Western blot assays,respectively.Results The expression of miR-214 in advanced hypopharyngeal carcinoma tissues （0.31 1 ±0.206） was significantly less than normal hypopharyngeal mucosa tissues（1.620 ± 1.394;t =5.09,P ＜ 0.05）.The expression of miR-214 was notably upregulated after tranfected with hsa-mir-214 compared with the negative control group （t =6.347,P ＜ 0.05）.The migration and invasion ability of FaDu cells transfeced with hsa-mir-214 was decreased by comparison with negative control cells （t =1 1.6,P ＜0.01; t =6.499,P ＜ 0.05）.There was no significant difference of the average clony number and the cloning efficiency between the experimental and negative control groups （t =0.592,P ＞ 0.05）.Results of Western blot assay showed that,Twist expression in the miR-214-overexpressed group was apparently decreased compared with that in the control group （t =6.545,P ＜ 0.05）.Conclusions miR-214 is expressed at a low level in advanced hypopharyngeal carcinoma tissues,and can obviously inhibit the invasion and migration abilities of FaDu cells,possibly because of its inhibiting effect on Twist expression.Additionally,miR-214 plays no significant role in the proliferation of FaDu cells.