p57与同源框基因HOXA10在子宫内膜基质细胞体外蜕膜化过程中的表达

本文旨在从mRNA和蛋白水平研究子宫内膜基质细胞(endometrial stromalcell,ESC)体外蜕膜化过程中p57和同源框基因HOXA10(homeobox A10 gene)的表达变化以及HOXA10的亚细胞定位,从而推测其在蜕膜化过程中的作用。本实验联合使用0.5mmol/L8-溴-cAMP和1×10?6mol/LMPA(medroxyprogesterone acetate)作用1、2、4d(D1、D2、D4)诱导ESC发生蜕膜化,相应时间点提取mRNA和蛋白质行半定量RT-PCR和免疫印迹,同时以2%低血清培养ESC1、4d作为对照(C1、C4)。用间接免疫荧光和基因转染的方法,观察蜕膜化过程中HOXA10的亚细胞定位。结果显示:(1)蜕膜化过程中HOXA10的表达进行性下降,D2开始与对照组(C4)比较具有显著性差异(P<0.05)。(2)相反,蜕膜化过程中p57的表达进行性上升,D2开始与对照组C4比较也有显著性差异(P<0.05)。(3)低血清培养ESC1、4d后,p57和HOXA10的表达没有显著性差异(P>0.05)。(4)蜕膜化过程中HOXA10始终定位于胞核,不发生胞浆胞核穿梭。以上观察结果表明:(1)p57的高表达是ESC脱离细胞周期走向分化的因素之一。(2)HOXA10的低表达可能是p57上调的原因之一。(3)孕激素受体(progesterone receptor,PR)途径参与了促进ESC脱离细胞周期而走向分化的过程。

Expression of p57 and homeobox A10 during decidualization of endometrial stromal cell in vitro.

In order to elucidate the function of homeobox A10 gene (HOXA10) and p57 during decidualization our present study was designed to observe the change of HOXA10 and p57 expression and subcellular localization of HOXA10 in the process of endometrial stromal cell (ESC) differentiation in vitro. Decidualization was induced by 0.5 mmol/L 8-Bromo-cAMP (8-Br-cAMP) together with 1x10(-6) mol/L medroxyprogesterone acetate (MPA). Expression of p57 and HOXA10 was detected by RT-PCR and Western blot after 1-day, 2-day, and 4-day treatment (D1, D2, D4). ESCs cultured in 2%FBS for 1 and 4 d were used as control (C1, C4). The location of HOXA10 was detected by indirect immunofluorescence and HOXA10-GFP transfection. The results are as follows: (1) The expression of HOXA10 decreased progressively during the course of decidualization, and showed significant difference compared to control group C4 after 2-day treatment (D2). (2) On the contrary, the expression of p57 increased progressively and also showed significant difference compared to the control group C4 after 2-day treatment (D2). (3) There was no significant change of HOXA10 and p57 expression after culturing ESCs in 2%FBS for 4 d (C1, C4). (4) HOXA10 located in the nucleus throughout the course. Cytoplasm and nucleus shuttle was not detected in the experiment. Our results suggest that the down-regulation of HOXA10 may contribute to the increase of p57 and that the up-regulation of p57 likely plays an important role in ESC differentiation. Progesterone receptor (PR) pathway may participate in promoting ESCs to exit cell cycle and enter differentiation.