本文選題：嗎啡依賴　切入點(diǎn)：大腦皮層　出處：《河北醫(yī)科大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

【摘要】：目的:嗎啡具有明顯的鎮(zhèn)痛作用是眾所周知的,但它同時(shí)又具有很高的成癮性,近年來嗎啡的濫用呈明顯增高趨勢(shì),嚴(yán)重危害了人類身心健康,社會(huì)安定和諧。持續(xù)的嗎啡攝取對(duì)全身各系統(tǒng)均可造成不同程度的損害,其中對(duì)神經(jīng)系統(tǒng)的毒性損害最為顯著。長(zhǎng)期嗎啡依賴導(dǎo)致了包括大腦皮層在內(nèi)的較多腦區(qū)發(fā)生病理性改變,導(dǎo)致大腦發(fā)生急、慢性缺血,缺氧,從而引發(fā)神經(jīng)細(xì)胞和神經(jīng)纖維的一系列損傷性改變,目前國(guó)內(nèi)外對(duì)于嗎啡依賴的研究主要限于機(jī)能和代謝方面,病理形態(tài)方面的研究尚不夠詳細(xì)。迄今為止的研究雖然表明嗎啡依賴導(dǎo)致了神經(jīng)細(xì)胞的損傷,但是何種基因蛋白參與了神經(jīng)損傷、神經(jīng)保護(hù)及神經(jīng)結(jié)構(gòu)的重塑,目前國(guó)內(nèi)外尚無詳細(xì)報(bào)道。在參與調(diào)節(jié)神經(jīng)細(xì)胞生存、維持及損傷的眾多轉(zhuǎn)錄因子中,神經(jīng)細(xì)胞微管蛋白2(microtubeasso ciated protein2,MAP2),信號(hào)轉(zhuǎn)導(dǎo)及轉(zhuǎn)錄激活蛋白3(Signal Transduction and transcription protein-3,STAT-3),載脂蛋白酶E(apolipoprotein E,APOE),3-硝基酪氨酸(3-nitrotyrosine,3-NT)、C-fos在不同時(shí)程嗎啡依賴中的表達(dá)變化目前國(guó)內(nèi)外未見詳細(xì)報(bào)道,本研究目的旨在探明MAP2、STAT-3、APOE、3-NT及C-fos在不同時(shí)程嗎啡依賴中的表達(dá)狀態(tài),為嗎啡毒性作用導(dǎo)致的神經(jīng)損傷的機(jī)制研究提供病理形態(tài)學(xué)依據(jù)。方法:1本實(shí)驗(yàn)采用健康Wistar大鼠,體重為(300±20)g的健康雌性大鼠64只,分為空白對(duì)照組1周組、嗎啡依賴1周組,空白對(duì)照組2周組、嗎啡依賴2周組,空白對(duì)照組4周組、嗎啡依賴4周組,空白對(duì)照組6周組和嗎啡依賴6周組。嗎啡依賴4組大鼠按逐日遞增原則,背部皮下注射鹽酸嗎啡,每天二次(8:00,20:00),連續(xù)5天。劑量分別為10mg·kg-1,20mg·kg-1,30mg·kg-1,40mg·kg-1,50mg·kg-1。每次注射前均用75%酒精嚴(yán)格消毒,一次性注射器注射。5天后確認(rèn)嗎啡依賴形成后,嗎啡依賴1周組、2周組、4周組、6周組分別給予30mg·kg-1鹽酸嗎啡背部皮下注射,一天兩次(8:00,20:00),分別連續(xù)注射至1周、2周、4周、6周,建立四個(gè)不同時(shí)段的嗎啡依賴大鼠模型�？瞻讓�(duì)照1周組、2周組、4周組、6周組分別注射等劑量的生理鹽水,分別連續(xù)注射至1周、2周、4周、6周。模型建立后,腦組織經(jīng)固定、脫水、透明、包埋、連續(xù)切片等處理后待做各種染色使用。①蘇木精-伊紅(HE)染色觀察常規(guī)病理學(xué)改變。②硫堇特殊染色觀察神經(jīng)細(xì)胞尼氏體變化。③Fluoro-Jade B染色以及原位末端標(biāo)記法(TUNEL)檢測(cè)嗎啡導(dǎo)致的神經(jīng)細(xì)胞變性壞死及凋亡情況。④免疫組織化學(xué)染色觀察相關(guān)蛋白表達(dá)變化。2統(tǒng)計(jì)學(xué)分析:數(shù)據(jù)以“均數(shù)±標(biāo)準(zhǔn)差(Mean±SD)”表示,用SPSS 13.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)分析,各組均數(shù)的比較行單因素方差分析(ANOVA),用最小顯著差法(least significant difference,LSD)作兩兩比較,以P0.05為有顯著統(tǒng)計(jì)學(xué)差異。結(jié)果:1 HE染色結(jié)果正常對(duì)照組大鼠大腦皮層組織結(jié)構(gòu)清晰,各層神經(jīng)細(xì)胞排列整齊,未見明顯病理形態(tài)學(xué)改變;嗎啡依賴1周組神經(jīng)細(xì)胞無明顯改變,嗎啡依賴2周組小血管周圍間隙稍有增大;嗎啡依賴4周組可見少量神經(jīng)細(xì)胞固縮,神經(jīng)細(xì)胞水腫,小血管及神經(jīng)細(xì)胞周圍間隙增大,小膠質(zhì)細(xì)胞增生;嗎啡依賴6周組可見組織結(jié)構(gòu)疏松,神經(jīng)細(xì)胞水腫,部分神經(jīng)細(xì)胞固縮,小膠質(zhì)細(xì)胞增生明顯。2硫堇染色結(jié)果正常對(duì)照組可見神經(jīng)細(xì)胞尼氏體顆粒清晰可辨;嗎啡依賴1周組與2周組未見明顯改變;4周組尼氏體數(shù)量減少,結(jié)構(gòu)不清;6周組尼氏體消失,細(xì)胞固縮。3 Fluoro-Jade B染色觀察大鼠大腦皮層神經(jīng)細(xì)胞變性壞死情況正常對(duì)照組未見變性壞死陽(yáng)性細(xì)胞;嗎啡依賴1周組與2周組未見變性壞死陽(yáng)性細(xì)胞;4周組偶見變性壞死陽(yáng)性細(xì)胞;6周組變性壞死陽(yáng)性細(xì)胞明顯增多。4原位末端標(biāo)記法(TUNEL)結(jié)果嗎啡依賴各組均未檢測(cè)到凋亡細(xì)胞,表明大腦皮層神經(jīng)細(xì)胞在本實(shí)驗(yàn)設(shè)定的嗎啡依賴時(shí)程內(nèi)未發(fā)生明顯的細(xì)胞凋亡。5免疫組織化學(xué)染色5.1 MAP2免疫組織化學(xué)染色結(jié)果正常對(duì)照組大鼠大腦皮層神經(jīng)細(xì)胞胞漿及突觸中MAP2呈強(qiáng)陽(yáng)性表達(dá),嗎啡依賴1周、2周、4周、6周陽(yáng)性表達(dá)隨嗎啡依賴時(shí)間的延長(zhǎng)呈下降趨勢(shì)。5.2 STAT-3、APOE免疫組織化學(xué)染色結(jié)果嗎啡依賴1周組與2周組大鼠大腦皮層神經(jīng)細(xì)胞STAT-3、APOE陽(yáng)性表達(dá)與正常對(duì)照組相比呈上升趨勢(shì),隨著嗎啡依賴時(shí)程的延長(zhǎng)4周、6周組陽(yáng)性表達(dá)又呈現(xiàn)了減少趨勢(shì)。5.3 3-NT免疫組織化學(xué)染色結(jié)果嗎啡依賴1周組與2周組大鼠大腦皮層神經(jīng)細(xì)胞3-NT陽(yáng)性表達(dá)與正常組相比未見明顯差異,4周、6周陽(yáng)性表達(dá)呈逐漸增高趨勢(shì)。5.4 C-fos免疫組化化學(xué)染色結(jié)果正常組未見陽(yáng)性表達(dá);嗎啡依賴1周組少量陽(yáng)性表達(dá);嗎啡依賴其余各組未見表達(dá)。結(jié)論:本研究成功建立了大鼠不同時(shí)程嗎啡依賴模型,通過常規(guī)組織病理學(xué)、組織化學(xué)特染及免疫組織化學(xué)觀察,得出以下結(jié)論:較長(zhǎng)時(shí)程的嗎啡依賴導(dǎo)致了大腦皮層神經(jīng)細(xì)胞的變性、壞死,MAP2、STAT-3、APOE、3-NT、C-fos參與了神經(jīng)細(xì)胞的變性壞死。
[Abstract]:Objective: morphine has obvious analgesic effect is as everyone knows, but it at the same time, which is highly addictive, in recent years, morphine abuse increased significantly, serious harm to human health, social stability and harmony. Sustained morphine intake cause different damage to the body of the system can be the toxic damage to nerve the system is the most significant. The long-term morphine dependence leads to more areas of the brain including the cerebral cortex, the pathological changes of the brain, leading to the occurrence of acute, chronic ischemia, hypoxia, which triggered a series of damage of nerve cell and nerve fiber changes at home and abroad for the study of morphine dependence is mainly limited to the aspects of function and metabolism. Study on the pathological aspects are not detailed. The research so far suggests that although morphine dependence leads to nerve cell damage, but what kind of gene protein involved in God After injury, neuroprotection and remodeling of neural structures, both at home and abroad have not been reported in detail. In the regulation of neuronal cell survival, maintenance and damage of many transcription factors, nerve cell microtubule protein 2 (microtubeasso ciated protein2, MAP2), signal transduction and transcription activator protein 3 (Signal Transduction and transcription protein-3, STAT-3). Apolipoprotein E (apolipoprotein E, APOE protease), 3- nitrotyrosine (3-nitrotyrosine, 3-NT), the expression of C-fos in different time dependent changes in Cheng Mafei, there is no detailed report, the purpose of this study is to explore MAP2, STAT-3, APOE, 3-NT and C-fos expression in different time dependent Cheng Mafei's pathology study on the mechanism of the morphological basis for nerve injury leading to morphine toxicity. Methods: in this experiment, 1 healthy Wistar rats, weight (300 + 20) g the health of 64 female rats, 鍒嗕負(fù)絀虹櫧瀵圭収緇，

本文編號(hào)：1594032