【摘要】：目的:許多臨床研究表明,慢性應(yīng)激是引發(fā)人類生殖功能障礙的重要因素。本研究采用慢性輕度不可預(yù)見性應(yīng)激與孤養(yǎng)相結(jié)合的方法建立慢性應(yīng)激大鼠模型,探索慢性應(yīng)激對(duì)大鼠生殖行為的影響,并從形態(tài)學(xué)角度檢驗(yàn)睪丸生精細(xì)胞和間質(zhì)細(xì)胞在慢性應(yīng)激作用下的形態(tài)學(xué)改變狀況。以此探討慢性應(yīng)激條件下,生殖行為和生殖功能改變的機(jī)制問題。 方法:16只雄性SD大鼠隨機(jī)分為對(duì)照組和模型組(n=8),模型組采用慢性輕度不可預(yù)見性應(yīng)激加孤養(yǎng)刺激大鼠,連續(xù)進(jìn)行35天,構(gòu)建模型,對(duì)照組正常飼養(yǎng)大鼠。整個(gè)實(shí)驗(yàn)過程中連續(xù)檢測(cè)大鼠體重,結(jié)束后使用糖水偏愛實(shí)驗(yàn)和曠場(chǎng)實(shí)驗(yàn)檢測(cè)大鼠的情緒狀態(tài),以確認(rèn)造模成功。接近行為的檢測(cè)使用Agmo提出的雌鼠偏愛的實(shí)驗(yàn)方法,用雌鼠偏愛分?jǐn)?shù)和雌/雄區(qū)停留時(shí)間反映大鼠的接近行為。行為檢測(cè)結(jié)束后犧牲大鼠,取睪丸組織石蠟包埋,HE染色后光學(xué)顯微鏡下觀察睪丸間質(zhì)和生精小管的形態(tài),測(cè)量間質(zhì)細(xì)胞面積、數(shù)量;測(cè)量生精小管直徑、生精上皮厚度及一定區(qū)域內(nèi)細(xì)胞數(shù),并作統(tǒng)計(jì)分析。與此同時(shí),樹脂包埋,超薄切片后,在透射電子顯微鏡下觀察對(duì)照組、模型組睪丸間質(zhì)細(xì)胞和生精細(xì)胞的細(xì)胞結(jié)構(gòu)和細(xì)胞器形態(tài),并對(duì)比發(fā)現(xiàn)其中的差異。 結(jié)果:(1)慢性應(yīng)激對(duì)情緒狀態(tài)的影響:在慢性應(yīng)激作用下從14天起模型組大鼠體重顯著少于對(duì)照組,并持續(xù)至35天。造模結(jié)束后與對(duì)照組相比,模型組大鼠糖水偏愛分?jǐn)?shù)減少(p0.01),曠場(chǎng)實(shí)驗(yàn)中行進(jìn)距離、站立次數(shù)顯著減少(p0.05,p0.01),中央格停留時(shí)間顯著延長(zhǎng)(p0.05),過中央格次數(shù)、糞便粒數(shù)顯著增多(p0.01,p0.05)。顯示動(dòng)物模型制作成功。 (2)慢性應(yīng)激對(duì)接近行為的影響:模型組自身對(duì)比,雌鼠處于發(fā)情期或發(fā)情間期對(duì)模型組雄鼠的雌鼠偏愛分?jǐn)?shù)不影響;對(duì)雌/雄鼠區(qū)停留時(shí)間無影響。與對(duì)照組對(duì)比,雌鼠為發(fā)情間期時(shí),模型組雌鼠偏愛分?jǐn)?shù)顯著減小(p0.001);模型組雄鼠雌鼠區(qū)停留時(shí)間顯著減少(p0.01),雄鼠區(qū)停留時(shí)間顯著增加(p0.01)。 (3)慢性應(yīng)激對(duì)睪丸影響的光鏡檢測(cè)結(jié)果:與對(duì)照組相比,模型組大鼠生精小管管腔內(nèi)的精子明顯稀少,模型組生精小管短軸平均直徑(p0.01)、生精上皮平均厚度(p0.01)和生精細(xì)胞數(shù)(p0.001)均小于對(duì)照組;睪丸間質(zhì)面積(p0.01)、單個(gè)間質(zhì)細(xì)胞面積(p0.01)睪丸間質(zhì)細(xì)胞數(shù)(p0.01)均小于對(duì)照組,且統(tǒng)計(jì)結(jié)果有顯著性意義。 (4)慢性應(yīng)激對(duì)睪丸影響的電鏡檢測(cè)結(jié)果:與對(duì)照組正常細(xì)胞相比,模型組精原細(xì)胞細(xì)胞質(zhì)與細(xì)胞核間出現(xiàn)空泡;精母細(xì)胞內(nèi)質(zhì)網(wǎng)擴(kuò)張,細(xì)胞質(zhì)出現(xiàn)空泡,線粒體腫脹;精子細(xì)胞內(nèi)質(zhì)網(wǎng)腫脹甚至形成空泡。精子鞭毛結(jié)構(gòu)紊亂,外周致密纖維出現(xiàn)斷裂和空泡。間質(zhì)細(xì)胞細(xì)胞核出現(xiàn)深凹陷,染色質(zhì)聚集,出現(xiàn)細(xì)胞凋亡現(xiàn)象。 結(jié)論:慢性應(yīng)激環(huán)境可以導(dǎo)致雄性大鼠發(fā)生生殖行為抑制;并且引發(fā)睪丸間質(zhì)細(xì)胞損傷和凋亡,以及生精細(xì)胞的受損。慢性應(yīng)激導(dǎo)致大鼠生殖行為的減退可能與睪丸間質(zhì)細(xì)胞受損,睪酮水平下降有關(guān)。
[Abstract]:Objective: Many clinical studies have shown that chronic stress is an important factor to induce human reproductive dysfunction. In this study, the model of chronic stress rats was established by the combination of chronic mild unpredictability and isolation, and the effects of chronic stress on the reproductive behavior of rats were explored, and the morphological changes of the spermatogenic cells and the interstitial cells under the effects of chronic stress were examined from the morphological angle. In this paper, the mechanism of the change of reproductive and reproductive function under the condition of chronic stress is discussed. Methods:16 male SD rats were randomly divided into control group and model group (n = 8). Rat body weight was detected continuously throughout the course of the experiment. After the end of the experiment, the experimental and open-field experiment was used to test the emotional state of the rats, so as to confirm the model. The test method of the female mouse preference proposed by the Agmo for the detection of the proximity behavior, using the female mouse preference score and the female/ male region residence time to reflect the proximity of the rat In order to sacrifice the rat after the end of the behavior detection, the testis tissue was embedded in paraffin, and the morphology of the testis and the seminiferous tubules was observed under the optical microscope after HE staining, the area and the number of the interstitial cells were measured, the diameter of the spermatogenic tube, the thickness of the spermatogenic epithelium and the cells in a certain region were measured. Count and make statistics In the meantime, the cell structure and organelle morphology of the testis and the spermatogenic cells in the control group and the model group were observed under the transmission electron microscope at the same time, and the difference was found in the model group. The results were as follows: (1) The effect of chronic stress on the emotional state: The weight of rats in the model group was significantly lower than that of the control group under the effect of chronic stress. Compared with the control group, the preference score of sugar water in the model group decreased (p0.01) and the number of standing times was significantly reduced (p0.05, p0.01), and the time of the central cell was significantly prolonged (p0.05), the number of cells in the central cell and the number of the grains per day (p0.01, p0). 05). Display animal model system to be successful. (2) The effect of chronic stress on the approach behavior: the model group itself vs. the female rats in the estrus or oestrus interval did not affect the female preference score of the male rats in the model group; the female/ male mouse area was stopped There was no effect on the time. In contrast to the control group, the preference score of the female rats in the model group was significantly decreased (p0.001), while the residence time of the female rats in the model group was significantly decreased (p0.01), and the residence time of the male rats was significantly increased (p (3) The results of light microscopy on the effects of chronic stress on the testis: compared with the control group, the sperm in the cavity of the model group was significantly less than that of the control group, and the average diameter of the short axis of the seminiferous tubule in the model group was less than that of the control group. (p0.01), the average thickness of the spermatogenic epithelium (p0.01) and the number of spermatogenic cells (p0.01) were all smaller than that of the control group, the interstitial area of the testis (p0.01), the area of the individual interstitial cells (p0.01) and the number of the testis interstitial cells (p0.01) were all smaller than that of the control group, and the statistical junction The effect of chronic stress on the testis was found to be significant. (4) The results of the electron microscopic examination of the effect of chronic stress on the testis: the vacuoles between the cytoplasm and the nucleus of the spermatogonia were observed in the model group compared with the normal cells in the control group, and the endoplasmic reticulum of the spermatocytes expanded and the cytoplasm was out. vacuolation The swelling of the net and even the formation of vacuoles. The structure of the flagellum of the sperm is disordered and the outer Breakage and vacuolation of the fibers. The nucleus of the interstitial cells has a deep depression, and the chromatin is concentrated. Conclusion: The chronic stress environment can lead to the inhibition of the reproductive behavior of the male rats. The death, and the damage to the spermatogenic cells. Chronic stress can lead to a decrease in the reproduction behavior of the rat and the interstitial cells of the testis.
【學(xué)位授予單位】：浙江理工大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類號(hào)】：R363

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