孕激素及间质细胞培养液对原代培养内膜腺上皮HOXA11基因表达的影响

目的:探讨间质细胞是否参与了孕激素对子宫内膜腺上皮的调控,及其初步的作用机制。方法:将增生期子宫内膜间质细胞经激素处理后进行培养,提取培养液。用浓度为30%的提取培养液对腺上皮细胞进行原代培养,当细胞生长融合时,加入孕酮或孕雌激素培养4h、24h。提取细胞总RNA,用半定量RT-PCR方法检测腺上皮细胞HOXA11基因表达量。结果:当内膜腺上皮细胞中含有30%经孕激素处理的间质细胞培养液时,加入孕激素或孕、雌激素后其HOXA11基因,在培养4h时表达量有下降趋势;24h时,表达量下降明显;而用RU486预处理后再加入孕激素或雌孕激素,腺上皮细胞HOXA11基因表达量与对照组无差异;当上皮细胞中含有30%经RU486预处理后,再加入孕激素处理的间质细胞培养液时,孕激素或孕、雌激素对内膜腺上皮HOXA11表达的负调控作用在4h时消失;24h时,转为正调控(HOXA11基因表达量增加)。结论:孕激素对内膜腺上皮HOXA11基因的负调控作用需要问质细胞分泌的孕激素依赖因子的参与,而且由间质细胞和内膜腺上皮中的孕激素受体共同介导完成这一负调控作用。

The Effect of Progesterone on HOXA11 Expression of Endometrial Epithelial Cells Primary Cultured with Stromal Cells Culture Medium

Objective: To examine whether the stromal cells took part in the process when HOXA11 gene expression was down-regulated by progesterone in endometrial epithelial cells in vivo. Method: The stromal cells were cultured in proliferative stage after the treatment of hormones then the stromal cells culture medium (SCM) was collected. The endometrial epithelial cells were incubated with SCM at concentrations of 30%, when cell fusion appeared, progesterone or progesterone with Mifepristone was added to culture for 4 hours and 24 hours, and total RNA was isolated from epithelial cells for semi-quantitative RT-PCR. Furthermore, the effect of progesterone and Mifepristone (RU486) on HOXA11 gene expression in endometrial epithelium with SCM was examined, and the alteration of HOXA11 gene at different time (the four hours, the twenty-four hours) was investigated. Result: In vitro, progesterone diminished HOXA11 gene expression in endometrial epithelium that was incubated in minimum essential medium(MEM) with 30% SCM treated with progesterone for 4 hours and 24 hours. But this change disappeared after adding RU486 before the treatment in endometrial epithelial cells. On the contrary, HOXA11 expression was stimulated by progesterone in the epithelial cells, which was incubated for 24 hours in MEM with 30% SCM that had been added RU486 before treatment with progesterone. Moreover, adding RU486 before progesterone in endometrial epithelium, it was unchangeable that HOXA11 gene expressed in endometrial epithelial cells which were incubated for 24 hours in MEM, diluted with 30% SCM treated with RU486 and progesterone. Conclusion: The stromal cells were necessary in the process when progesterone down-regulated the expression of HOXA11 gene in endometrial epithelium in midluteal. Meanwhile, the effect of progesterone on HOXA11 gene expression was mediated by PR both in the stromal and epithelial cells.