E-cadherin基因稳定转染小鼠胚胎干细胞及其对分化细胞黏附能力的影响

目的 转染上皮型钙黏蛋白(E-cadherin)基因入小鼠胚胎干(ES)细胞,观察其对分化细胞间黏附能力变化的影响.方法 构建E-cadherin的真核表达载体pEGFP-E-cadherin,转染小鼠ES细胞并进行体外分化.利用逆转录-聚合酶链反应(RT-PCR)等检测E-cadherin在分化系统中的动态表达,同时观察分化细胞间黏附能力的变化情况.结果 基因转染的ES细胞分化后第1～17天稳定表达E-cadherin,而普通ES细胞分化后则表达逐渐降低.稳定表达E-cadherin的分化细胞间黏附能力明显增强并在19 d时仍能保持致密的细胞连接和多层生长状态,普通ES细胞则由拟胚体(EB)结构逐渐分化为松散的细胞群落.结论 稳定表达E-cadherin的ES细胞分化后,细胞间黏附能力明显增强并保持多层细胞生长状态,这对于将ES细胞分化研究提升到组织水平具有积极的意义.

E-cadherin stable transfection into mouse ES cells and its effects on adhesion ability of differentiated cells in vitro

Objective To stably transfect E-eadhefin into embryonic stem cells (ESCs), and observe the impact of E-cadherin on adhesion ability of differentiated cells. Methods BALB/e mouse ESCs were infected with recombinant plasmid EGFP-E-cadherin. E-cadhefin dynamic expression in E-cadherinESCs differentiation system was detected by using RT-PCR and ICC. The appearances of cell-cell adhesion in the E-cadherin-ESCs differentiating system was observed dynamically under a microscope. Results The exogenous E-cadhefin gene was successfully transferred into BALB/e mouse ESCs and expressed stably. In E-cadherin-ESCs differentiation system, E-cadhefin-mRNA was expressed stably during the differentiating period from day 1 to day 17. But in un-infeeted ESCs differentiation system,E-cadherin-mRNA was not expressed till on day 5 and expressed weakly along with the differentiating time. At last, the E-cadhefin-ESCs formed pronounced cell aggregates and cuboidal morphology, whereas un-infected ESCs remained more spread out and corded in morphology. Conclusion Transferring exogenous E-cadherin gene into mouse ESCs could enhance the cell adhesion and maintain multi-layer form in ESCs differentiation system.