本文關(guān)鍵詞： 糖原合成酶激酶3β 腦衰反應(yīng)調(diào)節(jié)蛋白2 形覺剝奪性弱視 大鼠 視皮層　出處：《南華大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

【摘要】：目的:研究糖原合成酶激酶3β(Glycogen synthase kinase 3β,GSK-3β)和腦衰反應(yīng)調(diào)節(jié)蛋白2(Collapsin response mediator protein 2,CRMP-2)在形覺剝奪性弱視大鼠視皮層中的表達,探索其在弱視發(fā)病機制中的意義。方法:1.出生14天的健康大鼠64只,隨機選擇32只大鼠行單側(cè)眼瞼縫合術(shù)作形覺剝奪性弱視組,另外32只大鼠不做任何處理成為正常組。根據(jù)大鼠出生的天數(shù),在21天時將形覺剝奪性弱視組和正常組大鼠用10%水合氯醛麻醉后行P-VEP(Pattern Visual Evoked Potential)檢測,記錄P100波的潛伏期和波幅值,判斷單眼形覺剝奪性弱視模型是否建立成功。2.根據(jù)大鼠出生的天數(shù),分別在14天、21天、45天、120天4個時間點處死形覺剝奪性弱視組和正常組大鼠,每小個時間點處死大鼠各8只。所有處死的大鼠均取其大腦視皮層組織,用4%多聚甲醛PBS溶液固定后,封蠟行組織切片。采用HE(Hematoxylin Eosin)染色觀察形覺剝奪性弱視組和正常組大鼠視皮層神經(jīng)元細胞的形態(tài);然后采用sp免疫組化法檢測形覺剝奪性弱視組和正常組大鼠視皮層4個時間點gsk-3β、crmp-2免疫陽性神經(jīng)元的表達,并使用圖像分析系統(tǒng)測定gsk-3β、crmp-2免疫陽性神經(jīng)元表達的平均光密度值(aod值)并進行統(tǒng)計學(xué)分析。結(jié)果:1.p-vep檢測:與正常組相比,形覺剝奪性弱視組大鼠p100波的潛伏期明顯延長(p0.05),振幅明顯降低(p0.05),提示單側(cè)眼瞼縫合7天后,可成功建立單眼形覺剝奪性弱視大鼠模型。2.he染色:正常組大鼠視皮層神經(jīng)元細胞輪廓清晰,可清楚顯示各神經(jīng)元細胞的形態(tài),細胞分布較密集,形覺剝奪性弱視組大鼠視皮層神經(jīng)元分布稀松,且排列紊亂,無法清晰辨別細胞形態(tài),胞體變小,核固縮,呈嗜堿性,部分核溶解。3.免疫組化結(jié)果:1)gsk-3β在大鼠視皮層存在表達。正常組大鼠視皮層gsk-3β的表達在14天,21天,45天,120天各時間點之間比較,各時間點之間均無明顯差異(p0.05)。形覺剝奪性弱視組大鼠視皮層gsk-3β的表達在14天,21天,45天,120天各時間點之間比較,除45天與120天比較無差異(p0.05)外,其余各時間點之間兩兩比較均有差異(p0.05)。與正常組大鼠相比,形覺剝奪性弱視大鼠視皮層gsk-3β的表達在14天時無明顯差異(p0.05),但在21天,45天,120天時均升高(p0.05)。2)crmp-2在大鼠視皮層中存在表達。正常組大鼠視皮層crmp-2的表達在14天,21天,45天,120天各時間點之間比較,除45天與120天比較無差異(p0.05)外,其余各時間點之間兩兩比較均有差異(p0.05)。形覺剝奪性弱視組大鼠視皮層crmp-2的表達在14天,21天,45天,120天各時間點之間比較,除45天和120天(p0.05)外,其余各時間點之間兩兩比較均有差異均有統(tǒng)計學(xué)意義(p0.05)。與正常組大鼠相比,形覺剝奪性弱視大鼠視皮層crmp-2的表達在14天時無明顯差異(p0.05),但在21天,45天,120天時均降低(p0.05)。3)大鼠視皮層gsk-3β和crmp-2表達的相關(guān)性:在正常組大鼠視皮層中g(shù)sk-3β和crmp-2兩者之間無明顯相關(guān)性(r=-0.142,p0.05),而在形覺剝奪性弱視組大鼠視皮層中,兩者呈負相關(guān)性(r=-0.838,p0.05)。結(jié)論:1.在大鼠視覺發(fā)育關(guān)鍵期的早期,行單側(cè)眼瞼縫合術(shù)7天后可以成功建立單眼形覺剝奪性弱視大鼠模型。2.在正常大鼠視覺發(fā)育關(guān)鍵期內(nèi),gsk-3β和crmp-2在大鼠大腦視皮層均有表達,說明二者有可能參與大鼠視覺發(fā)育的過程。3.在形覺剝奪性弱視大鼠視皮層中,gsk-3β的表達隨著剝奪時間的延長逐漸升高,45天時達到峰值,而crmp-2的表達隨著剝奪時間延長而逐漸下降,45天時達到低值,兩者呈負相關(guān)性,提示在視覺發(fā)育的可塑性期內(nèi),gsk-3β和crmp-2可能通過某種調(diào)節(jié)機制參與弱視的發(fā)生發(fā)展過程。
[Abstract]:Objective: To study the influence of glycogen synthase kinase 3 (Glycogen synthase kinase 3 beta, beta GSK-3) and collapsin response mediator protein 2 (Collapsin response mediator protein 2, CRMP-2) in the perception of visual expression in the cortex of deprived amblyopia rats, and explore its significance in the pathogenesis of amblyopia. Methods: 1. 14 days of birth 64 healthy rats, 32 rats were randomly selected for unilateral eyelid suture for deprivation amblyopia, another 32 rats without any treatment as normal group. According to the number of days the rats were born, and in 21 days will form deprivation amblyopia group and normal group rats 10% chloral hydrate anesthesia (P-VEP Pattern Visual Evoked Potential) detection, record the latency and amplitude of P100 wave value judgment of monocular deprivation amblyopia model is established successfully according to the number of days.2. rats were born, respectively in 14 days, 21 days, 45 days, 120 days and 4 at the time of death Form deprivation amblyopia group and normal group rats, each time the rats were sacrificed 8 rats each. All rats were sacrificed were removed from the visual cortex tissue, with 4% paraformaldehyde fixed PBS solution after wax for tissue sections. By HE (Hematoxylin Eosin) staining of form deprivation amblyopia group and normal group rat visual cortex neuronal morphology; then using SP immunohistochemical method to detect the form deprivation amblyopia group and normal group rat visual cortex at 4 time points of GSK-3 beta, the expression of crmp-2 immunoreactive neurons, and use image analysis system for determination of GSK-3 beta, the average optical density of crmp-2 immunoreactivity positive neurons value (AOD value) and analyzed statistically. Results: 1.p-vep detection: compared with the normal group, deprivation amblyopia rats significantly prolonged the latency of P100 wave (P0.05), the amplitude decreased (P0.05), suggesting that unilateral eyelid suture can be 7 days. The successful establishment of monocular deprivation amblyopia rats.2.he staining: normal rat visual cortical neuron outline clear, can clearly show the neuron cell morphology, cell distribution is dense, the perception of visual cortical neurons were distributed deprived amblyopia rats, and unable to clearly identify the disorder, cell morphology, cell the body becomes small, karyopyknosis, basophilic nucleus dissolved.3. immunohistochemical results: 1) GSK-3 beta in the visual cortex of rats. The expression of GSK-3 beta expression in the cerebral cortex as normal rats in 14 days, 21 days, 45 days, 120 days between each time point, each time some were not significantly different (P0.05). The expression of GSK-3 beta form sense visual cortex deprived amblyopia rats in 14 days, 21 days, 45 days, 120 days between each time point, in addition to 45 days compared with 120 days of no difference (P0.05), 22 were the differences between the other time points (P0.05) and positive. 甯哥粍澶ч紶鐩告瘮,褰㈣鍓ュず鎬у急瑙嗗ぇ榧犺鐨眰gsk-3尾鐨勮〃杈懼湪14澶╂椂鏃犳槑鏄懼樊寮，

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