【摘要】：目的：初步探討(SD)雌性大鼠在長期力竭游泳運動導(dǎo)致動情周期抑制后卵巢相關(guān)激素的表達變化和基因表達差異,從受體和基因水平探討運動性月經(jīng)失調(diào)(AMI)的發(fā)生機制。方法：選用3月齡、健康且有正常動情周期的雌性SD大鼠22只,隨機分為造模組(n=12只)和空白對照組(n=10只)。空白對照組不做任何干預(yù),造模組每天進行力竭游泳訓練,每周游泳6天,休息1天,每天上午8時對兩組大鼠進行陰道涂片。當確定造模成功后處死大鼠,取卵巢進行一般形態(tài)學觀察、測量并制成組織切片,利用HE染色和免疫組織化學等方法觀察卵巢一般組織結(jié)構(gòu)、卵巢E2、ER、PR以及血清生殖激素等的表達變化,并采用RIA技術(shù)檢測血清FSH、LH、E2、P濃度；在前期試驗中通過RFDD-PCR技術(shù)建立兩組差異基因表達譜,并在其中選取與蛋白質(zhì)的降解密切相關(guān)的泛素蛋白酶系統(tǒng)的相關(guān)基因,包括Psmb5、Psmd1、Psnd8、Ube3a、Usp10、Usp9x等進行定量PCR驗證。結(jié)果：1.在訓練過程中,造模組所有大鼠逐漸出現(xiàn)動情周期紊亂,同時,空白對照組大鼠動情周期保持正常。2、HE染色結(jié)果顯示：在造模組,典型生長卵泡和新鮮黃體明顯減少,原始卵泡、閉鎖卵泡相對增加；而在空白對照組卵巢皮質(zhì)中,可見大量不同階段的卵泡,典型生長卵泡和新鮮黃體。3、卵巢E2、ER、PR免疫組化分析結(jié)果顯示：造模組顆粒細胞與黃體細胞E2表達的IOD比空白組顯著降低(P0.05)；與空白組相比,造模組卵巢黃體細胞中和顆粒細胞中ER表達的IOD顯著降低(P0.05)；與空白組相比,造模組顆粒細胞和黃體細胞中PR表達的IOD也顯著降低(P0.05)。4、血清生殖激素結(jié)果顯示：與空白組相比,造模組大鼠血清E2、P、FSH、LH水平顯著降低(P0.05),具有顯著性差異；5、經(jīng)過定量PCR驗證,Psmb5、Psmd1、 Psmd8、Ube3a、Usp10、Usp9x在模型組中過表達,與RFDD-PCR測得的結(jié)果一致。結(jié)論：1、長期力竭運動可引起雌性大鼠動情周期抑制,可為游泳運動項目女運動員運動性月經(jīng)失調(diào)的研究提供動物模型。2、長期力竭運動使大鼠卵巢形態(tài)學發(fā)生改變,進而抑制卵巢功能,使卵巢激素的表達發(fā)生改變,血清性激素水平下降。3、泛素蛋白酶系統(tǒng)中Psmb5、Psmd1、Psmd8、Ube3a、Usp10、Usp9x過表達,可能在長期力竭運動引起的雌性大鼠動情周期抑制中發(fā)揮重要的作用。
[Abstract]:Aim: to investigate the changes of ovarian related hormones and gene expression in (SD) female rats after prolonged exhaustive swimming induced estrous cycle inhibition. To explore the mechanism of (AMI) in exercise induced menstrual disorders at the receptor and gene levels. Methods: 22 healthy female SD rats were randomly divided into model group (n = 12) and blank control group (n = 10). The blank control group did not do any intervention, the model group was given exhaustive swimming training every day, swimming 6 days a week, resting for one day, and taking vaginal smear on the rats of the two groups at 8: 00 am every day. The rats were killed after the successful establishment of the model, the ovary was taken for general morphological observation, and the tissue sections were measured and made. The general structure of the ovary was observed by HE staining and immunohistochemistry, and the ovary E2ER-ER2 was observed. The expression of PR and serum reproductive hormone were detected by RIA technique. Two sets of differential gene expression profiles were established by RFDD-PCR technique in previous trials, and the genes related to the ubiquitin protease system, including Psmb5,Psmd1,Psnd8,Ube3a,Usp10, which were closely related to protein degradation, were selected. Usp9x et al were tested by quantitative PCR. Results: 1. During the course of training, all the rats in the model group showed estrous cycle disorder gradually, while the estrus cycle of the blank control group kept normal. 2 the results of HE staining showed that the typical growth follicles and fresh corpus luteum decreased obviously in the model group. Primordial follicle, atresia follicle relatively increased; In the control group, there were a large number of follicles at different stages, typical growth follicles and fresh corpus luteum. The results of PR immunohistochemical analysis showed that the expression of E2 in granulosa cells and luteal cells in model group was significantly lower than that in blank group (P0.05). Compared with the blank group, the IOD expression of ER in luteal cells and granulosa cells in the model group was significantly lower than that in the control group (P0.05). Compared with the blank group, the IOD expression of PR in granulosa cells and luteal cells in the model group was significantly lower than that in the blank group (P0.05). The level of LH was significantly decreased (P0.05). 5. The results of quantitative PCR showed that Psmb5,Psmd1, Psmd8,Ube3a,Usp10,Usp9x was overexpressed in the model group, which was consistent with the results obtained by RFDD-PCR. Conclusion: 1. Long-term exhaustive exercise can induce estrous cycle inhibition in female rats, and can provide an animal model for the study of exercise menstrual disorders of female athletes in swimming events. 2. Long-term exhaustive exercise can cause morphological changes in the ovary of rats. Furthermore, the ovarian function was inhibited, the expression of ovarian hormone was changed, the serum sex hormone level was decreased. 3. The overexpression of Psmb5,Psmd1,Psmd8,Ube3a,Usp10,Usp9x in the ubiquitin protease system. It may play an important role in the inhibition of estrous cycle induced by long-term exhaustive exercise in female rats.
【學位授予單位】：成都體育學院
【學位級別】：碩士
【學位授予年份】：2013
【分類號】：R87

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