滋养层细胞原代体外培养体系的建立  

Establishment of in Vitro Culture System of Human Archae-generation Trophoblasts

作　　者：刘惠萍[1] 王若光[2] 李春梅[2] 李宾玲[1] 曾润清[1] 秦明春[1] 易鹏飞[1] 

机构地区：[1]湖南中医学院2003级,硕士研究生湖南长沙410007 [2]湖南中医学院中西医结合系

出　　处：《实用预防医学》2006年第5期1109-1111,共3页Practical Preventive Medicine

基　　金：国家自然科学基金(30472227;30000225)

摘　　要：目的 改善体外分离、培养人早孕绒毛细胞滋养层细胞（cytotrophoblast CTB）的方法，并对其进行鉴定，探讨CTB的一些生物学特性。方法 取人工流产6～8周妊娠绒毛，机械法分离，复合酶消化，差速贴壁法纯化细胞后继续培养。倒置显微镜观察其大体形态及体外转化，HE染色观察其胞核情况，免疫细胞化学观察细胞的细胞角蛋白18（cytokeratin CK18）、波形蛋白（vimentin Vim）和人胎盘催乳素（human placental lactogenh PL）的表达。流式细胞仪测定其细胞周期。结果分离培养的CTB呈不规则多角形片状，单层铺展生长。所有的细胞都表达CK18，Vim只在部分细胞表达，hPL免疫反应阳性物质在胞浆表达。细胞周期显示约有58％的细胞处于G0／G1期。结论 成功分离培养了人早孕绒毛CTB，方法简便易行，获得的细胞形态单一、生长稳定，为进一步研究CTB在妊娠生理和妊娠免疫耐受中的作用提供了实验基础。Objective To improve the methods for isolation and culture of human cytotrophoblasts （CTBs） from early pregnancy villi, to identify and investigate its partial biological properties. Methods Human villi from 6 - 8 weeks legal voluntary interrupted pregnancy were mechanically isolated, complex enzyme - digested, and purified by different speeding adherence. The cellular morphology was observed by inverted phase contrast microscope and the nucleus and cytoplasm were observed by haematoxylin and eosin stain （HE）. Immunocytochemistry was used to examine the expression of cytokeratin 18 （OK18）, vimentin （Vim） and human placental lactogen （hPL）. Flow cytometer was used to observe the cell cycle. Results The cultured cells appeared as patchy spreading and irregular polygonal with big oviform nucleus and transparent cytoplasm. OK18 expressed in all the cells cultured and Vim expressed only in some of the cells, hPL immunoreactive positive substances were distributed in cytoplasm. Cell cycle revealed that 58 % cells were at G0/G1 phase. Conclusion We successfully cultured the human cytotrophoblasts isolated from early pregnancy villi and obtained monomorphic cells with stable growth, which provided a useful experimental model for the studies of the roles of CTBs in gravidity physiology and immune tolerance.

关 键 词：滋养层细胞 免疫细胞化学 细胞培养 生物学特性 

分 类 号：R-331[医药卫生]