NRP-1在调控人舌鳞状细胞癌细胞分化中的作用  

Effects of NRP-1 on Mediating the Differentiation of Human Tongue Squamous Cell Carcinoma

作　　者：唐子春[1] 王子露[1] 刘来奎[1] 宋晓萌[1] 朱丽芳[1] 叶金海[1] 吴煜农[1] 

机构地区：[1]南京医科大学口腔研究所,江苏南京210018

出　　处：《口腔医学研究》2012年第6期601-604,共4页Journal of Oral Science Research

基　　金：江苏省自然科学基金(编号:BK2010531)

摘　　要：目的:构建神经纤毛素1(Neuropilin-1,NRP-1)短发夹RNA(short hairpin RNA,shRNA)慢病毒表达载体,体外评价其对人舌鳞状细胞癌细胞株SCC25(squamous cell carcinoma 25)分化的影响。方法:根据shRNA设计原则,设计合成用于体内干扰NRP-1编码序列的shRNA,重组入慢病毒载体PLKO.1。将构建正确的PL-KO.1/NRP-1-shRNA感染SCC25,western blot和实时荧光定量PCR检测NRP-1的表达,划痕实验和实时荧光定量PCR检测其对SCC25分化的影响。结果:测序证实重组质粒构建成功;体外试验表明,SCC25转染了NRP-1-shRNA后NRP-1表达水平降低,干扰效率达65%。划痕实验结果表明NRP-1下调明显降低了SCC25的迁移能力。实时荧光定量PCR结果表明NRP-1下调可以导致SCC25上皮标记蛋白钙粘附蛋白(epi-thelial cadherin,E-cadherin)表达上调,而间充质标记蛋白纤粘蛋白(fibronectin,FN)和波形蛋白(Vimentin,VIM)表达下调,与对照组相比具有显著性差异(P<0.05)。结论:慢病毒介导的NRP-1-shRNA成功在体外抑制了目的基因的表达,并影响SCC25的分化。Objective：To construct a lentiviral plasmid of RNA interference of neuropilin-1（NRP-1）gene,and evaluate the interference effect of NRP-1-shRNA on the differentiation of squamous cell carcinoma 25（SCC25）in vitro.Methods：According to the target sequence and design principle of shRNA,the short hairpin RNA was designed and synthesized to interfere the NRP-1expression,and the shRNA duplex was ligated into the PLKO.1 vector.SCC25 was transfected with PLKO.1/NRP-1-shRNA,and the expression level of NRP-1was detected by real time RT-PCR and western blot;scratch test and real time RT-PCR were used to investigate the differentiation potential of SCC25.Results：The recombinant vector was successfully constructed and confirmed by sequencing.In vitro experiment demonstrated that the expression level of NRP-1was significantly downregulated,and the interfering efficiency was 65%;the down-regulation of NRP-1not only significantly decreased the migration of SCC25,but also increased the expression level of E-cadherin（epithelial cadherin）while decreased the expression level of FN（fibronectin）and VIM（Vimentin）（P〈0.05）.Conclusion：RNA interference can successfully interfere with the expression of NRP-1and affect the differentiation of SCC25.

关 键 词：短发夹RNA 慢病毒载体 神经纤毛素1 SCC25 

分 类 号：R739.86[医药卫生—肿瘤]