摘要
[目的]探讨PD-L1过表达对HeLa细胞迁移的影响。[方法]通过分子克隆构建PD-L1质粒载体(p EGFPPD-L1);通过PEI法转染质粒,调节质粒转染量(1μg、3μg、5μg)和转染时间(24 h、36 h、48 h、72 h),优化转染条件;通过细胞划痕实验检测细胞融合率,Western Blot检测细胞迁移相关蛋白(E-钙黏蛋白、N-钙黏蛋白、波形蛋白)的表达量。[结果]最佳转染条件是质粒量3μg/孔,转染后48 h观察,此时转染效率可达(82.94±8.08)%。转染pEGFP-PD-L1质粒后,HeLa细胞的PD-L1过表达3.5倍,划痕融合率显著增高(p<0.01);波形蛋白和N-钙黏蛋白的表达量显著升高(p<0.05),E-钙黏蛋白表达量显著下降(p<0.05)。[结论]PD-L1过表达显著促进HeLa细胞迁移能力和上皮向间质转化水平。
[Objective]To investigate the effect of PD-L1 overexpression on HeLa cell migration.[Methods]PD-L1 overexpression plasmid（pEGFP-PD-L1） was constructed by molecular cloning technique and transfected into HeLa cell by PEI method.The optimized transfection condition was chosen by adjusting plasmid quantity（1 μg,3 μg,5 μg） and transfection time（24 h,36 h,48 h,72 h）.Cell fusion efficiency was detected by wound healing assay.Vimentin,E-cadherin,and N-cadherin proteins expression levels were detected by Western Blot assay.[Results] The optimized transfection condition was adding 3μg/well plasmid and 48 h transfection time.The highest transfection efficiency was（82.94 ± 8.08） %.After plasmid transfection,PD-L1 expression was 3.5 times higher than untransfected group and the wound area healing efficiency was significantly increased（p 〈 0.01）.Vimentin and N-cadherin protein expressions were significantly increased and E-cadherin protein expression was significantly decreased（p 〈 0.05）.[Conclusion] PD-L1 overexpression promotes HeLa cell migration and the level of epithelial-mesenchymal transition.
出处
《生物技术》
北大核心
2017年第6期557-562,共6页
Biotechnology
基金
国家自然科学基金项目("MRTF-A和STAT3调控乳腺癌EMT及机制研究"
No.31501149
"抗活性氧自由基的载白藜芦醇Ti O2纳米管促进骨修复研究"
No.31700824)
