高纯度子宫内膜细胞分离和体外培养技术及其应用

目的：分离、纯化子宫内膜间质细胞（ESC）和腺上皮细胞，建立体外ESC蜕膜化模型。方法：选择正常育龄妇女的新鲜子宫内膜组织，经多酶消化制成细胞悬液，分离、纯化间质细胞和腺上皮细胞；分别培养至细胞聚合成片，随机分组，用等渗浓度的甾体激素诱导ESC体外蜕膜化改变。结果：分离的间质细胞其纯度达95%以上，其细胞活力达92%——95%。在此体外培养系统中，ESC对生理剂量激素的应答出现与活体相仿的蜕膜化改变。结论：利用此技术可在细胞和分子水平上研究子宫内膜的各种功能及其调节。

Application and technique in the separation and culture of highly purified endometrial cells in vitro

Objectives:Purification of the epithelial and stromal cells from human normal endometrium; and induction of decidual changes in cultured stromal cells by physiological doses of ovarian steroids. Methods:By biopsy, specimens were histologically normal proliferative or secretary endometrium. Separated cells were obtained using enzymatic digestion in conjunction with physical separation procedures. Epithelial and stromal cells were purified and cultured as monolayers in RPMI 1640 medium containing antibiotics, 25 mol/L HEPES, and 10% fetal bovine serum. Stromal cells had been treated with physiological doses of steroids after monolayers had become confluent. Then, the hormonal effects on the morphological changes, assessed with phase contrast microscopy and on the production of prolactin(PRL) were measured. Results: The morphology of the two main endometrial cell types in culture is depicted in the figure. Isolated populations of endometrial stromal cells(ESC) exhibited over 95% homogeneity, ascertained immunocytochemically; Treatment with physiological doses of estradiol(E 2), testosterone(T) and progesterone(P) resulted in an increased PRL and differentiation into decidual cells that are bigger and rounding and have the morphology of hormone secreting cells. During the process of decidualization, stromal cells change from fibroblastic spindles into polygonal cells and begin to express specific decidual protein PRL. Conclusions:We have developed an in vitro system in which human ESC respond to physiological doses of ovarian steroids with morphology, and biochemical changes that are characteristic of decidualization in vivo .Therefore, this culture system provides an in vitro model for decidualization.