CHARACTERISTICS OF TROPHOBLAST CELLS MIGRATING FROM FIRST TRIMESTER CHORIONIC VILLUS EXPLANTS AND PROPAGATED IN CULTURE
CHARACTERISTICS OF TROPHOBLAST CELLS MIGRATING FROM FIRST TRIMESTER CHORIONIC VILLUS EXPLANTS AND PROPAGATED IN CULTURE
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DOI：
10.1016/0143-4004(95)90100-0
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发表时间：
1995-07-01
期刊：
影响因子：
3.8
通讯作者：
LALA, PK
中科院分区：
文献类型：
--
作者：
IRVING, JA;LYSIAK, JJ;LALA, PK
我们开发了一种从机械衍生的绒毛碎片的初级外植体中生长出来的纯早期妊娠人滋养层细胞的培养方法(Yagel等人，1989；Graham等人，1992)。我们现在已经广泛地描述了这些细胞在其最初生长和长期培养中的特征，采用了各种标记和技术，如下所述。通过使用上皮细胞(细胞角蛋白)和间充质细胞(波形蛋白)的双重免疫荧光标记，我们将绒毛迁移细胞群鉴定为纯滋养细胞(39%的副生体)或滋养细胞和成纤维细胞的混合物(61%)。用抗细胞角蛋白抗体和一组其他初级抗血清通过双重标记免疫染色进一步对纯滋养细胞突起进行表型鉴定，发现这些细胞表现出多种原位绒毛外侵袭性滋养细胞的标志：胰岛素样生长因子(IGF)-II、NDOG-5、增殖细胞核抗原(PCNA)、人类白细胞抗原框架抗原(W6/32)以及一组不同的整合素，包括α1、α3、α5、αv和β(1)亚单位以及α(V)β(3)/β(5)Vitonectin受体。它们的整合素α(6)和β(4)亚基均为阴性。在IV型胶原凝胶上生长的外植体的免疫金电子显微镜显示，单核滋养层细胞和多核细胞产生常规和癌胚型纤维连接蛋白。免疫标记、流式细胞仪和免疫沉淀显示，在长期培养中，这种表型特征完全保持不变；因此，从早孕绒毛外植体中迁移出来的滋养层细胞及其繁殖的后代属于胎盘侵袭性绒毛外滋养层细胞。
We developed a method of propagating pure first trimester human trophoblast cells growing out of primary explants of mechanically derived chorionic villus fragments (Yagel et al, 1989; Graham et al, 1992). We have now extensively characterized these cells during their initial outgrowth and in long-term culture, employing a variety of markers and techniques as outlined below. By double label immunofluorescence using epithelial (cytokeratin) and mesenchymal (vimentin) cell markers, we identified the chorionic villus migrant cell populations as pure trophoblast (39 per cent of outgrowths) or a mixture of trophoblast and fibroblast (61 per cent). Further phenotyping of the pure trophoblast outgrowths by double label immunostaining using anti-cytokeratin antibody and a panel of other primary antisera revealed that these cells exhibit a variety of markers characteristic of extravillous invasive trophoblast cells in situ: insulin-like growth factor (IGF)-II, NDOG-5, proliferating cell nuclear antigen (PCNA), human leucocyte antigen framework antigen (W6/32) and a distinct set of integrins including alpha 1, alpha 3, alpha 5, alpha v and beta(1) subunits and alpha(v) beta(3)/beta(5) vitonectin receptor. They were negative for alpha(6) and beta(4) integrin subunits. Immunogold electron microscopy of explants grown on type IV collagen gel revealed the production of conventional and oncofetal types of fibronectin by mononucleate tropholast cells and human placental lactogen by multinucleate cells. immunolabelling, flow cytometry and immunoprecipitation revealed that this phenotypic profile was retained with complete fidelity in the long-term culture; thus, trophoblasts migrating out of first trimester chorionic villus explants and their propagated progeny belong to the invasive extravillous trophoblast of the placenta.