人子宫内膜中存在一群具有克隆增殖能力的细胞

Cloning efficiency of human endometrial cells in vitro

背景:克隆形成能力是干细胞的重要特性,多项研究证明,子宫内膜在起源上具有单克隆性。目的:通过体外培养,检测人子宫内膜细胞克隆形成能力。方法:采用机械和酶消化相结合的方法分离出人子宫内膜细胞,培养腺上皮细胞和基质细胞,观察细胞形态及生长特性并检测波形蛋白和角蛋白的表达,采用流式细胞仪对细胞进行CD133,CD34,CD45,CD90,CD73及CD29表型鉴定;观察细胞克隆生长特点、测量细胞克隆大小和计算克隆形成率。同时取人子宫肌细胞培养作为对照。结果与结论:经过12d的原代培养后,腺上皮细胞和基质细胞均可形成大、小两种克隆。角蛋白表达阳性为腺上皮细胞,波形蛋白表达阳性为基质细胞。流式细胞术检测子宫内膜细胞表达CD90、CD29、CD73等表面标记,不表达CD34、CD45,CD133。腺上皮细胞、基质细胞大克隆和小克隆大小及形成率比较,差异均有显著性意义(P<0.05);人子宫肌细胞用有限稀释法原代培养无细胞克隆生长。说明子宫内膜细胞在体外培养中,腺上皮细胞和基质细胞大克隆增殖能力均高于小克隆。这些细胞可能是子宫内膜干细胞。

BACKGROUND： Colony forming ability is an important characteristic of stem cells. Many studies have shown that the endometrium has monoclonal characteristics. OBJECTIVE： To detect the cloning efficiency of human endometriat cells in vitro METHODS： Human endometrial cells were isolated and dissociated mechanically and enzymatically from human endometrium. Stromal and epithelial cells were separated by two series of filters. The cell morphology and growth characteristics were observed and the expressions of vimentin and keratin were detected. Flow cytometry was used to identify the expression of CD133, CD34, CD45, CD90, CD73 and CD29 markers in endometrial cells. The clone growth characteristics were observed, the size of clones was measured and the rate of cloning efficiency was calculated. Meanwhile. human uterine muscle cell was used as a control. RESULTS AND CONCLUSION： After primary cultured for 12 days, glandular epithelial cells and stromal cells were divided into large and small clones by size and growth characteristics. The gland clone was positive for keratin and the stromal clone was positive for vimentin by immunohistochemistry. Results from flow cytometry showed that endometrial cells were positive for CD29, CD90 and CD73 but negative for CD34, CD45 and CD133. There was significant difference of the sizes and cloning effiency of the large and little gland clones between glandular epithelial cells and stromal cells （P 〈 0.05）. Human uterine smooth muscle cells were primary cultured by limiting dilution method. The endometrial cells can form landular epithelial cells and stromal cells in vitro which included large and small clones. The proliferation efficiency of the large gland clone was higher than that of the small gland clone. These cells may be the endometrial stem cells.