转化生长因子-β和Smad4在绝经过渡期大鼠卵巢颗粒细胞中的表达

目的 探讨转化生长因子-β（TGF-β）、Smad4 在绝经过渡期大鼠卵巢颗粒细胞中的表达，分析其与卵巢功能衰退的关系。 方法 雌性SD大鼠分为对照组（C组，6月龄，阴道涂片筛选，n=9）、绝经过渡期组（MT组，12~14月龄，阴道涂片筛选，n=8），大鼠麻醉处死后迅速取出卵巢，采用机械分离方法释放卵泡颗粒细胞，于CO₂培养箱中培养，利用免疫细胞化学法检测促卵泡刺激素受体（FSHR）蛋白的表达，鉴定卵巢颗粒细胞；免疫细胞化学法检测TGF-β、Smad4蛋白在两组卵巢颗粒细胞的表达；采用Real-time PCR的方法检测各组颗粒细胞中TGF-β、Smad4 mRNA的表达。 结果 免疫细胞化学显示分离培养的颗粒细胞纯度＞95%，MT组TGF-β、Smad4蛋白表达水平低于对照组（P<0.05）；Real-time PCR结果显示，两组均有TGF-β、Smad4 mRNA的表达，MT组TGF-β、Smad4 mRNA表达水平显著低于对照组（P<0.05）。 结论 TGF-β、Smad4参与绝经过渡期大鼠卵巢功能的衰退过程，绝经过渡期大鼠功能衰退的部分原因可能与卵巢颗粒细胞中TGF-β、Smad4 的表达降低有关。

Expression of transforming growth factor-β and Smad4 in ovarian granulosa cells in menopausal transitional rat

Objective To investigate the expression of transforming growth factor beta(TGF-β) and Smad4 in ovarian granulosa cells of rats during menopausal transition and to analyze their relationship with the decline of ovarian function. Methods Female SD rats were divided into control group (group C, 6 months old, vaginal smear screening, n=9) and menopausal transition(MT) group (MT group, 12-14 months old, vaginal smear screening, n=8), rats were sacrificed quickly after ovariectomy. The ovarian granulosa cells were released by mechanical separation method, and cultured in CO₂ incubator. The expression of follicle-stimulating hormone receptor(FSHR) protein was observed by immunocytochemistry to identify ovarian granulosa cells. Immunohistochemistry was used to detect the expression of TGF-β and Smad4 protein in ovarian granulosa cells and images were collected and analyzed; The expression of TGF-β and Smad4 mRNA in granulosa cells was detected by Real-time PCR. Results Immunocytochemistry showed that the purity of granulosa cells was > 95%. The intensities of immunocytochemical positive staining for TGF-β and Smad4 in MT group was lower than those in normal group (P<0.05).Real-time PCR showed that both TGF-β and Smad4 mRNA were expressed. The amount of Real-time PCR for TGF-β and Smad4 mRNA was significantly lower than those of the control group (P<0.05). Conclusion TGF-β and Smad4 may be involved in the decline of ovarian function in rats with menopausal transition. The causes of the functional decline in postmenopausal rats may be partly related to the decrease of TGF-β and Smad4 expression in ovarian granulosa cells.