前梯度蛋白2干扰质粒的构建及其对鼻咽癌细胞生物学功能的影响

Construction of shRNA targeting AGR2 gene and effect of AGR2 on biological function of nasopharyngeal carcinoma cells

目的:构建前梯度蛋白2(AGR2)基因干扰重组质粒及建立AGR2基因沉默的鼻咽癌细胞及动物模型,检测AGR2对鼻咽癌细胞的生物学功能的影响。方法:首先以real-time PCR和Western blot验证AGR2在鼻咽癌细胞系5-8F和6-10B中的表达,以高表达AGR2的5-8F细胞作为干扰转染的目标细胞;设计及构建shRNA表达载体pSR-GFP/Neo-AGR2-shRNA,将其转染至5-8F鼻咽癌细胞系中,通过Transwell小室迁移及侵袭实验来检测其迁移和侵袭功能的变化。结果:与6-10B细胞比较,AGR2在5-8F中的表达显著上调(P<0.05);干扰鼻咽癌细胞5-8F中的AGR2可使细胞迁移和侵袭功能明显减弱(P<0.05),使其体外成瘤能力显著降低(P<0.05)。结论:AGR2在鼻咽癌细胞系5-8F中明显高表达;pSR-GFP/Neo-AGR2-shRNA可成功干扰5-8F中AGR2的表达;AGR2可抑制5-8F细胞的迁移、侵袭及体内成瘤能力。

AIM： To construct the shRNA targeting anterior gradient protein 2 （AGR2） gene for exploring the effect of AGR2 on the biological behavior of nasopharyngeal carcinoma （NPC） cells. METHODS： The expression of AGR2 at mRNA and protein levels in NPC cell lines 6-10B and 5-8F was detected by real-time PCR and Western blot. The pSR- GFP/Neo-AGR2-shRNA expression vector targeting AGR2 was constructed. Based on the interference targeting AGR2, the cell migration and motility were determined by Transwell migration and motility assays. RESULTS ： The expression of AGR2 was increased in NPC cell line 5-8F compared with NPC cell line 6-10B （P 〈0. 05） . When the AGR2 expression in 5-8F cells was interfered, the cell migration, invasion and tumorigenicity were weakened. CONCLUSION： The expres-sion of AGR2 is up-regulated in NPC cell line 5-8F. pSR-GFP/Neo-CLU-shRNA successfully inhibits the expression of AGR2 in NPC cell line 5-8F. AGR2 inhibits the migration, invasion and tumorigenicity of 5-8F cells in vivo.