子宫内膜细胞分离培养方法的改良及其雌二醇分泌水平评价

Improvement on Isolation and Culture of Human Eutopic Endometrial Cells and Evaluation on the Production of Estradiol in Isolated and Incorporated Cultural Endometrial Cells Respectively

本研究对现有子宫内膜腺上皮和基质细胞的培养方法加以改良,并对分离与混合培养的子宫内膜细胞分泌的雌二醇水平进行比较,探讨子宫内膜异位症(简称内异症)子宫内膜局部高浓度雌二醇的生成来源。通过高浓度胶原酶和DNase联合消化、筛网过滤及贴壁纯化法,分离纯化25例子宫内膜腺上皮和基质细胞,通过形态学观察和细胞免疫荧光法鉴定,22例分离成功的内膜细胞中腺上皮细胞和基质细胞的纯度均在95%以上。电化学发光免疫法检测内膜细胞培养液上清液中的雌二醇水平,结果显示,内异症在位内膜细胞混合培养组的分泌水平最高,明显高于其它各组(P<0.01),而经过分离培养后的内异症在位内膜腺上皮细胞组明显高于基质细胞组(P<0.01)。在非内异症内膜细胞的混合和分离培养组中,细胞分泌雌二醇的水平极低,接近最低检测值。显示内异症患者从其发病的源头——"在位内膜"就已表现出高分泌雌激素的特征。

A better yield of isolated and purified human endometrial glandular epithelial cells and stromal cells was obtained by a modified culture procedure, and the level of estradiol production in their cultural supernanant was measured by electrochemiluminescence immunoassay in order to explore the histoloical origin of high concentration of estradiol in local endometrium of endometriosis. Digestion with high concentrated collagenase and DNase, filtration and adhesion purification were used to isolate, purify human endometrial glandular epithelial cells and stromal cells in 25 samples. The endometrial cell culture was successfully established in 22 of 25 endometrium samples which was observed under the inverted microscope. Cytokeratin and vimentin were detected by indirect immunofluescence staining to identify the glandular epithelial cells and stromal cells respectively, and the purities of the cells were both higher than 95%. The level of estradiol production in the incorporated cultural eutopic endometrial cells group was higher than that of any other group （P〈0.01）, and the level of glandular epithelial cells was higher than that of stromal cells in two isolated groups （P〈0.01）, Both incorporated and isolated endometrial cells without endometriosis showed an extremely low concentration of estradiol production which was almost up to the limit of detection. These findings suggest that eutopic endometrium of patients shares alterations with the ectopic tissue at a higher concentration of estrogen production.