雄激素对卵巢颗粒细胞性激素结合球蛋白表达的调节

发布时间：2018-03-13 04:38

本文选题：高雄激素血症　切入点：雄激素受体　出处：《医学研究生学报》2017年05期 　论文类型：期刊论文

【摘要】：目的雄激素信号通路参与调控卵泡早期生长发育以及卵泡闭锁过程,而性激素结合球蛋白是调控卵巢局部雄激素水平的重要因素。文中探讨雄激素对卵巢颗粒细胞中性激素结合球蛋白(SHBG)表达的影响。方法 1培养人卵巢颗粒癌细胞株(KGN),分别给予含0 nmol/L DHT,500 nmol/L DHT,500 nmol/L DHT+60μmol/L flutamide的细胞培养液,提取细胞蛋白。用Western blot、免疫荧光等方法检测SHBG的表达。2脱氢表雄酮(DHEA)诱导高雄激素血症模型,12只雌性SD大鼠随机数字表法分为HA组和对照组,每组6只。HA组每只皮下注射DHEA 6 mg/(100 g·d)溶于0.2 m L实验级大豆油,对照组给予等体积的大豆油。连续注射35 d后腹腔注射5%水合氯醛麻醉各组大鼠,下腔静脉取各组大鼠血液,ELISA方法检测各组血清中SHBG的水平;剪切各组大鼠卵巢和部分肝,免疫组化方法检测卵巢颗粒细胞中SHBG的表达,Western blot方法检测肝雄激素受体(AR)及SHBG表达情况。结果与0nmol/L DHT处理24h AR表达(1.06±0.03)相比,300、400、500nmol/L DHT处理后(1.06±0.02、1.61±0.11、2.38±0.14)均升高(P0.05);SHBG与AR表达变化一致,随DHT浓度梯度增加表达升高,但在500 nmol/L DHT处理24 h后SHBG表达升高显著(P0.01)。与0 nmol/L DHT比较,500 nmol/L DHT处理后AR蛋白、SHBG蛋白表达均升高(P0.01);与500 nmol/L DHT比较,500 nmol/L DHT+60μmol/L flutamide处理后AR蛋白、SHBG蛋白表达均降低(P0.05)。免疫荧光结果表明,与0 nmol/L DHT相比,DHT促进AR的表达,同时加入flutamide可明显抑制AR的表达。SHBG的表达与AR相一致。卵巢组织HE染色结果表明HA组与对照组相比卵巢形态发生改变,表现为多囊卵巢;卵巢免疫组化结果表明SHBG在HA组中表达高于对照组。HA组血清SHBG表达低于对照组[(2.41±0.14)vs(4.80±0.35),P0.01]。HA组中肝SHBG表达表达低于对照组,而AR蛋白表达高于对照组(P0.05)。结论雄激素信号通路激活后SHBG在卵巢颗粒细胞癌细胞以及大鼠卵巢颗粒细胞中表达升高。
[Abstract]:Objective androgen signaling pathway is involved in early follicular growth and development and follicular atresia. The effect of androgen on the expression of SHBG in ovarian granulosa cells was studied. Methods 1. The cell culture medium containing 0 nmol/L DHT 500 nmol/L nmol/L DHT 60 渭 mol/L flutamide was used. Cell proteins were extracted. 12 female SD rats were randomly divided into HA group and control group by Western blot2 and immunofluorescence. 2. Dehydroepiandrosterone (DHEA) induced hyperandrogenemia model. 6 rats in each group were injected subcutaneously with DHEA 6 mg/(100 g 路d) dissolved in 0.2ml soybean oil. The control group was given soybean oil of the same volume. After 35 days of continuous injection, 5% chloral hydrate was injected intraperitoneally to anesthetized rats in each group. The levels of SHBG in serum of each group were detected by Elisa, and the ovary and partial liver of each group were cut off. The expression of SHBG and SHBG in granulosa cells were detected by immunohistochemical method. Results compared with 0 nmol / L DHT treatment for 24 h, the expression of AR was 1.06 卤0.03). Compared with 0 nmol / L DHT, the expression of AR was 1.06 卤0.02nmol / L DHT (1.06 卤0.02nmol / L) and 1.06 卤0.02nmol / L DHT + 0.112.38 卤0.14). The expression increased with the increase of DHT concentration gradient. However, after treatment with 500 nmol/L DHT for 24 h, the expression of SHBG increased significantly (P0.01G). Compared with 0 nmol/L DHT, the expression of AR protein of nmol/L DHT was significantly higher than that of 500 nmol/L DHT, and the expression of AR protein was significantly decreased after treatment of 500 nmol/L DHT with 500 nmol/L DHT 60 渭 mol/L flutamide. The results of immunofluorescence showed that, after treatment with 500 nmol/L DHT, the expression of SHBG was significantly higher than that of 0 nmol/L DHT. Compared with 0 nmol/L DHT, the expression of AR was enhanced by flutamide, and the expression of SHBG was significantly inhibited by flutamide. The results of HE staining in ovarian tissue showed that the morphology of ovary in HA group was changed compared with that in control group, which showed polycystic ovary. The expression of SHBG in HA group was higher than that in control group. The expression of serum SHBG in HA group was lower than that in control group [2.41 卤0.14 vs 4.80 卤0.35 P0.01] .The expression of liver SHBG in HA group was lower than that in control group. The expression of AR protein was higher than that of the control group P0.050.Conclusion the expression of SHBG in ovarian granulosa cell carcinoma and rat ovarian granulosa cells is increased after androgen signaling pathway activation.
【作者单位】：南京大学医学院江苏省医学分子技术重点实验室;
【基金】：国家自然科学基金(81471422)
【分类号】：R711.75

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