沉默YAP1可抑制鼻咽癌细胞的增殖、迁移及侵袭能力

YAP1 knockdown suppresses the proliferation, migration and invasion of human nasopharyngeal carcinoma cells

目的探究沉默Yes相关蛋白1(YAP1)对鼻咽癌细胞增殖、迁移及侵袭能力的影响。方法采用实时荧光定量PCR和Western blot方法检测YAP1在鼻咽癌细胞系中的表达水平;在鼻咽癌细胞S26和S18中转染3条合成的小干扰RNA:si-NC, siYAP1#1和si-YAP1#2,以转染阴性对照si-NC的细胞作为对照组,转染si-YAP1#1和si-YAP1#2的细胞作为实验组,实时荧光定量PCR和Western blot检测转染后的干扰效率;通过细胞计数、平板克隆形成等方法评估转染si-YAP1#1和si-YAP1#2后的实验组和转染si-NC的对照组对鼻咽癌细胞增殖能力的影响;用划痕和Transwell方法分析各组细胞迁移与侵袭能力的变化;Western blot检测各组细胞中c-myc、上皮性钙黏蛋白(E-cadherin)、神经性钙黏蛋白(N-cadherin)和波形蛋白(Vimentin)等与细胞增殖和上皮间质转化相关蛋白的表达变化。结果 YAP1在鼻咽癌细胞系中高表达;与阴性对照组相比,转染si-YAP1#1和siYAP1#2的实验组细胞中YAP1的mRNA及蛋白水平均显著降低(P<0.05);生长计数、平板克隆、划痕及Transwell等实验结果均显示转染si-YAP1#1和si-YAP1#2的实验组细胞的生长和克隆能力、划痕愈合能力及侵袭能力较阴性对照组细胞明显下降(P<0.01);且相较阴性对照组,实验组细胞中c-myc、N-cadherin和Vimentin等蛋白的表达水平降低,E-cadherin的表达量增加。结论 YAP1在鼻咽癌细胞系中高表达,在鼻咽癌细胞中利用siRNA靶向敲低YAP1的表达后可以抑制鼻咽癌细胞的增殖、迁移及侵袭能力,提示YAP1可以作为一个潜在的鼻咽癌临床治疗干预靶点。

Objective To investigate the effects of Yes-associated protein 1(YAP1) knockdown on the proliferation, migration and invasion in human nasopharyngeal carcinoma(NPC) cells. Methods We detected the expression of YAP1 mRNA and protein in different NPC cell lines and an immortalized nasopharyngeal epithelial cell line using RT-PCR and Western blotting.Two YAP1-targeting small interfering RNAs(siRNA) were transfected into NPC cell lines S26 and S18, and the knockdown efficiency was confirmed by RT-PCR and Western blotting. The effect of YAP1 knockdown on the proliferation of the NPC cells was determined by cell counting and colony formation assay;wound healing assay and Transwell assay were used to analyze the changes in the cell migration and invasion abilities in each group. Western blotting was used to analyze the changes in the expressions of c-myc, E-cadherin, N-cadherin and vimentin in the NPC cells after YAP1 knockdown. Results YAP1 was highly expressed in the NPC cell lines. Compared with the negative control group, the NPC cell lines with YAP1 knockdown showed significantly lowered YAP1 expressions at both the mRNA and protein levels(P<0.05). YAP1 knockdown significantly suppressed the growth, cloning formation, migration and invasion of the NPC cells as compared with control cells(P<0.01).YAP1 knockdown obviously decreased the expression levels of c-myc, N-cadherin and vimentin and increased E-cadherin expression in the NPC cells. Conclusions YAP1 knockdown via siRNA suppresses the proliferation, migration and invasion of NPC cells in vitro, suggesting that YAP1 may serve as a therapeutic target for NPC.