本文選題：卵巢功能衰退 + POF��； 參考：《河北北方學院》2017年碩士論文

【摘要】：卵巢功能衰退是指由于女性卵巢的生殖功能和內分泌功能下降導致月經紊亂、生育力下降及低雌激素癥狀的發(fā)生,也可稱為卵巢儲備功能下降(Decreasing Ovarian Reservation,DOR),包括圍絕經期卵巢功能衰退和卵巢早衰(premature ovarian failure,POF)。卵巢功能衰退意味著女性開始進入衰老,嚴重影響著女性生殖健康及預期健康生命質量。因此探尋卵巢衰老中內在分子機制,尋找靶向目的蛋白改善卵巢功能從而延緩卵巢功能衰退具有重大意義。卵巢功能衰退存在極大的個體差異性,提示卵巢內微環(huán)境起著重要作用,本課題以絕經過渡期雌性大鼠為研究對象,著眼于與卵巢功能密切相關的轉化生長因子-β(trans-forming growth factorβ,TGF-β)信號通路中的細胞內信號轉導蛋白Smad蛋白,探究在卵巢功能衰退中受體激活型Smad蛋白Smad2、Smad3及抑制型Smad蛋白Smad7的蛋白水平及基因水平的表達變化,初步研究Smad蛋白與卵巢功能衰退的關系。本課題選取陰道涂片篩選法驗證的12月齡絕經過渡期雌性SD(Sprague-Dawley)大鼠28只,,隨機分成絕經過渡期組(MT)、雌二醇治療絕經過渡期組(E2),每組14只,另選14只6月齡雌性大鼠作為性成熟對照組(C)。E2治療組皮下注射17-β雌二醇(0.2mg/kg,1次/3天),MT組及C組皮下注射同等劑量的芝麻油溶劑6周后取雙側卵巢,計算卵巢指數、行HE染色觀察卵巢形態(tài)和卵泡數量計數評估各組卵巢功能,通過實時熒光定量PCR法檢測Smad2,Smad3,Smad7基因在各組卵巢組織中的mRNA表達差異,免疫組織化學方法檢測Smad2、P-Smad2、Smad3、P-Smad3、Smad7蛋白的在各組卵巢中的定位和表達變化。根據卵巢指數及形態(tài)學計算各級卵泡數量結果發(fā)現MT組及E2組卵巢功能較C組降低,MT組卵巢功能為最低,結果有統(tǒng)計學意義(P0.05)。RT-q PCR結果顯示MT組、E2組Smad2 mRNA、Smad3mRNA表達水平顯著低于C組(P0.01),E2組Smad2、Smad3 mRNA水平顯著高于MT組(P0.05);Smad7 mRNA水平在MT組、E2組中顯著高于C組(P0.01),E2組低于MT組(P0.05)。免疫組織化學結果表明Smad2、Smad3、P-Smad2、P-Smad3及Smad7蛋白在顆粒細胞、卵母細胞及卵泡膜細胞中均有表達,三組間有表達差異。Smad2,Smad3蛋白水平在C組、E2組、MT組中呈逐漸下降趨勢,具有統(tǒng)計學意義(P0.05),相同P-Smad2、P-Smad3蛋白水平呈相同趨勢,結果無統(tǒng)計學意義;而Smad7蛋白水平在C組、E2組、MT組呈升高趨勢,具有統(tǒng)計學意義(P0.05)。通過對以上結果分析得出,Smad2、Smad3、Smad7在卵巢卵泡細胞中廣泛表達,對卵巢功能起調控作用。在卵巢功能衰退過程中,無論從蛋白水平及基因水平,均有Smad2,Smad3,Smad7參與,Smad2、Smad3起正向調控,Smad7起負向調控作用。卵巢功能衰退與Smad2,Smad3表達下調,Smad7表達上調有關。
[Abstract]:The decline of ovarian function refers to the disorder of menstruation, fertility decline and the occurrence of low estrogen symptoms due to the decline of reproductive and endocrine functions of female ovaries. Also known as ovarian reserve decline, decreasing Ovarian reservation, including peri-menopausal ovarian dysfunction and premature ovarian failure. The decline of ovarian function means that women begin to age, which seriously affects their reproductive health and the quality of life of their expected health. Therefore, it is of great significance to explore the intrinsic molecular mechanism of ovarian senescence and to find target protein to improve ovarian function and delay ovarian function decline. There are great individual differences in ovarian dysfunction, suggesting that the microenvironment plays an important role in the study of menopausal transition female rats. We focus on the intracellular signal transduction protein Smad protein in transforming growth factor 尾 TGF- 尾 signaling pathway, which is closely related to ovarian function. To investigate the expression of receptor activated Smad protein Smad2Smad3 and inhibitory Smad protein Smad7 in ovarian dysfunction, and to explore the relationship between Smad protein and ovarian dysfunction. In this study, 28 postmenopausal female SD Sprague-Dawley rats verified by vaginal smear screening were randomly divided into menopausal transition group (MTO) and estradiol group (14 rats in each group). Another 14 6-month-old female rats were selected as the control group of sexual maturation. The ovarias were obtained by subcutaneous injection of 17- 尾 estradiol 0.2mg / kg of estradiol 0.2mg / kg in group C and group C by subcutaneous injection of the same dose of sesame oil solvent for 6 weeks, and the ovarian index was calculated, and the ovarian index was calculated by subcutaneous injection of 17- 尾 estradiol 0.2mg / kg of estradiol. The morphology of ovary and the number of follicles were observed by HE staining to evaluate the ovarian function in each group. The difference of mRNA expression of Smad2 Smad3 Smad7 gene in ovarian tissues was detected by real-time fluorescence quantitative PCR method. Immunohistochemical method was used to detect the localization and expression of Smad2T P-Smad3P- Smad3 Smad7 protein in the ovaries of each group. According to the ovarian index and morphology, the ovarian function of MT and E2 group was lower than that of C group, and the ovarian function of MT group was the lowest. Results the results showed that the expression of Smad2 mRNA-Smad3 mRNA in MT group was significantly lower than that in group C (P 0.01). The level of Smad3 mRNA was significantly higher in MT group than that in MT group. The level of Smad7 mRNA in MT group was significantly higher than that in group C (P 0.01) and that in group C was significantly higher than that in group C (P 0.01). The results of immunohistochemistry showed that Smad2Smad3 and Smad7 protein were expressed in granulosa cells, oocytes and follicular cells, and the Smad2 Smad3 protein level decreased gradually in group C and E 2 group, and the expression of Smad3 protein was decreased gradually in group C and E 2 group, and the expression of Smad2 Smad3 protein was decreased gradually in group C and E _ 2 group, and the expression of Smad2 Smad3 protein was decreased gradually in E _ 2 group and T _ T group. The level of P-Smad2, P-Smad3 protein showed the same trend, but the level of Smad7 protein increased in group C (P 0.05). Through the analysis of the above results, it is concluded that Smad2, Smad3, Smad7 are widely expressed in ovarian follicular cells and play a regulatory role in ovarian function. In the course of ovarian function decline, Smad2, Smad3 and Smad7 play a negative role in the positive regulation of Smad7 both in protein level and gene level. The decline of ovarian function was associated with down-regulation of Smad3 expression and up-regulation of Smad7 expression.
【學位授予單位】：河北北方學院
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R711.75

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