BeWo细胞模型的建立与验证.doc

  BeWo 细胞模型的建立与验证# * 5 10 15 20  摘要：目的:研究建立 BeWo 细胞模型的方法，并验证其生物学特性。方法:将 BeWo 细胞以 1×105cells /cm2 接种到 Transwell 通透性支持物上，培养 5 d 时用透射电镜观察细胞的形态 学特点，测定跨膜电阻、标志性低-高分子量标志物荧光素和异硫氰酸荧光素标记的葡聚糖 （FD70S）的转运率；RT-PCR 法检测 BeWo 细胞中 P-糖蛋白（P-gp）/ MDR I mRNA 的表 达，并以罗丹明为 P-gp 探针底物，评价胎盘转运体 P-gp 的外排功能。结果:BeWo 细胞单层 生长 4d 时，形成均匀、致密的单层细胞，出现紧密连接，跨膜电阻达到 60?/cm2；3h 内荧 光素和 FD70S 的转运率分别为（9.82±1.66）%和（0.24±0.04）%。BeWo 细胞存在 P-gp mRNA 表达，维拉帕米抑制 P-gp 外排功能后，罗丹明 Papp(AP→BL)值由（5.79±0.36）×10-6cm/s 增大为（9.79±0.13）×10-6cm/s，表明 BeWo 细胞模型的 P-gp 外排功能良好。结论:本研究条 件下所建立的 BeWo 细胞模型具备与胎盘滋养层细胞相似的形态学特点，转运功能完备,可 以作为研究胎盘物质转运机制的体外模型。 关键词：BeWo 细胞；转运；胎盘 中图分类号：R714.7 Establishment and validation of human trophoblast cell line BeWo Guo Jie1, Song Dianrong1, Han Fang2, Zhang Wei1, Wang Yanan1, Wang Yuhua1 (1. Department of Gynecology,the Second Hospital Affiliated to Tianjin University of Traditional Chinese Medicine, TianJin 300150; 25  Abstract: Objective 2. Linfen People's Hospital, ShanXi LinFen 041000) To investigate the establishment of BeWo cell monolayer model, and to identify BeWo cell biological characteristics. Methods BeWo cell was cultivated in Transwell plate inserts with1×105cells/cm2 cell density in standard procedures for 5d. The morphological and growth characters of the monolayer were observed by transmission electron microscope. The 30 35 40 compactification of the monolayer was determined by the transepithelial electrical resistance (TEER) and the permeability of low and high–molecular-weight fluorescent makers (fluorescein and fluorescein-labeled dextran T70) were used to investigate the permeability of the model. The expression of P-glycoprotein mRNA in BeWo cell was detected by RT-PCR, and the efflux function of P-glycoprotein was evaluated by the Rh123, a probe substrate of P-glycoprotein. Results BeWo cell monolayer became well-proportioned and compact after 4 d growth and tight conjunctions were formed between cells. TEER value reached 60?/cm2, and the relative rates of fluorescein and FD70