不同浓度去氢表雄酮对小鼠蜕膜化子宫内膜基质细胞增殖及 Dtprp mRNA 表达的影响

摘要

Objective To observe the effects of different concentrations of dehydroepiandrosterone ( DHEA) on prolif-eration and decidual prolactin-related protein ( Dtprp) mRNA expression of decidualized endometrial stromal cells in mice . Methods Primary endometrial stromal cells were isolated and cultured in vitro from mice on day 4 of pregnancy .The cells were divided into the experimental group 1, group 2, group 3, group 4, model group and control group .The experimental groups and the model group were treated with estradiol ( E2 ) and progesterone ( P4 ) to induce in vitro decidualization .The experimental groups 1, 2, 3 and 4 were exposed to 0.1, 1, 10 and 50μmol/L DHEA, respectively.The control group was not treated with E2, P4 and DHEA.The expression of Dtprp mRNA was detected by real-time PCR after 72-hour culture. Morphological changes were observed under inverted microscope after 24 and 48 h.The cells of the experimental groups and model group were cultured in 96-well plates except the blank wells .The cell proliferation ability was measured using a cell counting kit-8 after 72 h.Results The relative expression of Dtprp mRNA in the experimental groups 1, 2, 3 and 4 was 0.978 0 ±0.258 0, 0.764 0 ±0.135 0, 0.250 0 ±0.022 0 and 0.013 0 ±0.000 0, respectively;while that in the model group and control group was 1.000 0 ±0.000 0 and 0.000 2 ±0.000 0.The expression of Dtprp mRNA in both group 3 and group 4 was lower than that of the model group (all P<0.05).Significant difference was found between the experi-mental group 2 and group 4 (P<0.05).The cells in the control group showed a fibroblast-like appearance.Cells in the model group enlarged and had an increased amount of cytoplasm .The appearance of cells in the experimental group 1 and group 2 was similar to that of the model group .Cells in the experimental group 3 and 4 became polygonal with less cyto-plasm.The corrected absorbance of the experimental groups 1, 2, 3, 4, model group and backgroud absorbance were 1.222 8 ±0.038 2, 0.790 5 ±0.207 1, 0.592 5 ±0.229 8, 0.265 8 ±0.037 4, 1.149 7 ±0.106 2 and 0.087 1 ± 0.006 3, respectively.Significant difference was found between experimental groups 3, 4 and the model group, between experimental group 1 and groups 2, 3 and 4, between experimental group 2 and group 4 (all P<0.05).Conclusion The proliferation and Dtprp mRNA expression of mouse endometrial stromal cells are not significantly affected by 0.1 and 1μmol/L DHEA, while 10 and 50 μmol/L DHEA inhibits cell proliferation and down-regulates Dtprp mRNA expression .%目的：观察不同浓度去氢表雄酮（ DHEA）对小鼠蜕膜化子宫内膜基质细胞增殖及蜕膜催乳素相关蛋白（ Dtprp） mRNA表达的影响。方法体外分离培养妊娠第4天小鼠的子宫内膜基质细胞。将细胞分为实验1组、实验2组、实验3组、实验4组、模型组、对照组；各实验组与模型组以E2、P4诱导蜕膜化，实验1、2、3、4组分别加入0．1、1、10、50μmol／L的DHEA，模型组不加DHEA，对照组培养液不给予E2、P4及DHEA；培养72 h后采用real-time PCR法检测细胞中的Dtprp mRNA，分别于培养24、48 h后倒置显微镜下观察细胞形态变化。取各实验组、模型组的细胞悬液接种于96孔板，调零组不接种细胞；培养72 h后检测细胞增殖能力。结果实验1、2、3、4组细胞中Dtprp mRNA相对表达量分别为0．9780±0．2580、0．7640±0．1350、0．2500±0．0220、0．0130±0．0000，模型组与对照组分别为1．0000±0．0000、0．0002±0．0000；实验3、4组Dtprp mRNA相对表达量与模型组相比，P均＜0．05；实验2组与实验4组相比，P＜0．05。对照组细胞呈成纤维细胞样；模型组细胞形态更饱满，胞质增多；实验1、2组细胞形态与模型组近似，实验3、4组细胞呈多角形，胞质减少。实验1、2、3、4组细胞增殖能力分别为1．2228±0．0382、0．7905±0．2071、0．5925±0．2298、0．2658±0．0374，模型组为1．1497±0．1062，调零组校正A值为0．0871±0．0063；实验3、4组与模型组相比，P均＜0．05；实验1组与实验2、3、4组相比，P均＜0．05；实验2组与实验4组相比，P＜0．05。结论0．1、1μmol／LDHEA对蜕膜化子宫内膜基质细胞增殖及DtprpmRNA表达无明显影响，而10、50μmol／L DHEA可抑制细胞增殖，并下调Dtprp mRNA表达。