E-cadherin expression during the differentiation of human trophoblasts.
E-cadherin expression during the differentiation of human trophoblasts.
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发表时间：
1991-11
期刊：
影响因子：
4.6
通讯作者：
Christos Coutifaris;L. Kao;H. Sehdev;U. Chin;G. O. Babalola;O W Blaschuk;Jerome F. Strauss
中科院分区：
文献类型：
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作者：
Christos Coutifaris;L. Kao;H. Sehdev;U. Chin;G. O. Babalola;O W Blaschuk;Jerome F. Strauss
人类滋养层细胞的形态和功能分化最终形成终末分化的多核合胞滋养层。在培养中，分离的单核细胞滋养层细胞聚集，然后融合形成合胞体，重演体内过程。在本研究中，我们研究了钙（2+）依赖的细胞粘附分子（CAM），E-cadherin，在滋养层细胞形态分化过程中的表达。从人绒毛膜绒毛分离细胞滋养层细胞，通过分散进行中的培养物获得JEG-3和BeWo绒毛膜癌细胞，细胞滋养层细胞系，其在标准培养条件下不具有融合能力。培养物终止在定时间隔和E-钙粘蛋白进行了分析，免疫细胞化学和电子显微镜使用特定的抗体。此外，通过蛋白质印迹和北方印迹研究E-钙粘蛋白的表达。在细胞滋养层细胞的聚集过程中，E-钙粘蛋白定位于细胞表面的细胞-细胞接触点，并不能证明以下细胞融合。相反，在整个培养期间，它保留在聚集的JEG-3和BeWo细胞的表面上。蛋白质印迹分析显示，E-钙粘蛋白（120 × 10（3）Mr）的时间依赖性增加，这与正常细胞滋养层细胞和JEG-3细胞在24 h时的最大聚集体形成一致。随后观察到融合细胞滋养层细胞中E-钙粘蛋白的显著减少，因为合胞滋养层细胞成为培养中的主要细胞形式。与免疫组化观察一致，在非融合的JEG-3细胞中E-钙粘蛋白水平没有变化。北方印迹证明在96小时的培养中融合感受态细胞中4.5 kb转录物显著减少。正常非融合BeWo细胞暴露于1.5 mM 8-溴环AMP诱导细胞融合和合胞体形成。这个过程是伴随着从细胞表面的E-钙粘蛋白的免疫细胞化学和蛋白质印迹和平行减少的E-钙粘蛋白mRNA的丰度评估的消失。免疫中和实验中使用的抗血清直接对细胞外结构域的钙粘蛋白抑制合胞体形成正常滋养层细胞相比，抗血清对E-钙粘蛋白胞质尾，这对这些细胞的聚集和融合没有影响。我们的结论是，E-钙粘蛋白存在于一个动态的融合能力的细胞滋养层细胞，其基因表达的下调与细胞融合相一致。此外，这一过程似乎是环AMP介导的BeWo绒毛膜癌细胞。（400字处截断摘要）
The morphologic and functional differentiation of human trophoblast cells culminates in the formation of the terminally differentiated multinucleated syncytial trophoblast. In culture, isolated mononuclear cytotrophoblasts aggregate and then fuse to form syncytia, recapitulating the in vivo process. In the present studies, we investigated the expression of the Ca(2+)-dependent cell adhesion molecule (CAM), E-cadherin, during the morphologic differentiation of trophoblastic cells. Cytotrophoblasts were isolated from human chorionic villi, and JEG-3 and BeWo choriocarcinoma cells, cytotrophoblastic cell lines which under standard culture conditions are not fusion competent, were obtained by dispersion of ongoing cultures. Cultures were terminated at timed intervals and E-cadherin was analyzed by immunocytochemistry and electron microscopy using specific antibodies. In addition, E-cadherin expression was investigated by western and northern blotting. During the aggregation of cytotrophoblasts, E-cadherin was localized on the cell surface at points of cell-cell contact and could not be demonstrated following cellular fusion. In contrast, it remained on the surface of aggregated JEG-3 and BeWo cells throughout the duration of culture. Western blot analysis revealed a time-dependent increase in E-cadherin (120 x 10(3) Mr) which coincided with maximal aggregate formation at 24 h in both normal cytotrophoblasts and JEG-3 cells. A marked reduction of E-cadherin in fusing cytotrophoblasts was subsequently observed as syncytial trophoblasts became the predominant cellular form in culture. In agreement with the immunohistochemical observations, there was no change in E-cadherin levels in the non-fusing JEG-3 cells. Northern blotting demonstrated a significant reduction in the 4.5 kb transcript in fusion-competent cells over the 96 h of culture. Exposure of the normally non-fusing BeWo cells to 1.5 mM 8-bromo cyclic AMP induced cellular fusion and syncytium formation. This process was accompanied by a disappearance of E-cadherin from the cell surface as assessed by immunocytochemistry and western blotting and a parallel reduction in the abundance of the E-cadherin mRNA. Immunoneutralization experiments using an antiserum directed against the extracellular domain of cadherins inhibited syncytium formation in normal trophoblasts compared to an antiserum against the E-cadherin cytoplasmic tail, which had no effect upon aggregation and fusion of these cells. We conclude that E-cadherin exists in a dynamic state in fusion-competent cytotrophoblasts and that down regulation of its gene expression coincides with cellular fusion. In addition, this process appears to be cyclic AMP-mediated in BeWo choriocarcinoma cells.(ABSTRACT TRUNCATED AT 400 WORDS)