本文關(guān)鍵詞： 缺血缺氧 AQP4基因沉默 磁共振成像 血氣生化　出處：《大連醫(yī)科大學(xué)》2012年碩士論文　論文類型：學(xué)位論文

【摘要】：目的：新生兒缺氧缺血性腦病(hypoxic-ischemic encephalopathy，HIE)或稱為缺氧缺血性腦損傷(HIBD)主要是由圍產(chǎn)期窒息缺氧所引致的新生兒腦損傷疾病，窒息所致的早期缺血缺氧性腦水腫是本病的主要發(fā)病機(jī)制。臨床發(fā)病較為常見，凡能引起新生兒窒息的因素，如胎兒宮內(nèi)窘迫，高位產(chǎn)鉗術(shù)，產(chǎn)傷等，都可引起HIE，該病是造成新生兒傷殘的主要疾病。根據(jù)病情分為輕、中、重型，輕者預(yù)后良好，無后遺癥殘留；重者由于中樞神經(jīng)系統(tǒng)嚴(yán)重缺血缺氧，可導(dǎo)致新生兒早期死亡或產(chǎn)生永久性的神經(jīng)損害。本研究以注射過AQP4siRNA干擾和對照序列的新生豬為實驗對象，模擬早期新生兒HIBD模型，通過觀察比較兩組動物在臨床癥狀體征、影像學(xué)表現(xiàn)和病理學(xué)表現(xiàn)的差異，來探討AQP4基因沉默技術(shù)在早期新生豬缺血缺氧性腦損傷中的潛在價值，以期為新生兒HIBD疾病的診斷和治療提供新的理論依據(jù)和方向。 方法：將32只健康清潔的新生約克種豬隨機(jī)分為實驗組和對照組，通過腦脊液循環(huán)途徑給藥，分別鞘注等量的AQP4siRNA干擾序列和對照序列混合溶液。經(jīng)過24h的腦內(nèi)轉(zhuǎn)染后，對實驗組和對照組新生豬進(jìn)行相同程度的缺血缺氧處理，模擬早期新生兒HIBD模型。通過對比兩組動物缺血缺氧后各時點在早期神經(jīng)行為學(xué)、血氣生化酶學(xué)指標(biāo)、MR分子影像學(xué)和腦神經(jīng)組織的早期病理學(xué)表現(xiàn)的差異，運用柱狀圖或堆積條圖對兩組動物的神經(jīng)行為學(xué)評分和血氣生化酶學(xué)指標(biāo)進(jìn)行直觀的比較，并采用T檢驗或非參數(shù)檢驗對各時點評分和血氣生化指標(biāo)進(jìn)行兩兩比較，采用重復(fù)測量方差分析和T檢驗的方法對兩組動物各時點DWI圖像的ADC值進(jìn)行比較，同時采用免疫組織化學(xué)法（SP法）檢測兩組新生豬腦內(nèi)AQP4蛋白的表達(dá)水平并進(jìn)行陽性細(xì)胞計數(shù)，判斷兩組的差異是否具有顯著性（P0.05）。所用數(shù)據(jù)均采用SPSS16.0統(tǒng)計軟件包進(jìn)行統(tǒng)計分析。 結(jié)果：實驗組（即注射AQP4siRNA干擾序列組）和對照組（即注射AQP4siRNA對照序列組）新生豬在自主呼吸、皮膚黏膜顏色和神經(jīng)系統(tǒng)癥狀上均出現(xiàn)不同程度的異常，但實驗組在自主規(guī)律呼吸和正常皮膚黏膜顏色的恢復(fù)更快，在意識、顱神經(jīng)、反射、動作及合作協(xié)調(diào)等神經(jīng)功能的反應(yīng)表現(xiàn)更佳；且對照組在HI后2h、3h、6h的神經(jīng)行為學(xué)評分分別為7.200±0.789、10.000±1.700、13.400±1.174，明顯低于實驗組。實驗組HI后12h的PCO2、PO2、LDH分別為34.77±9.40mmHg、85.90±18.93mmHg、586.22±111.58U/L，對照組該時點各指標(biāo)值分別為45.94±12.52mmHg、65.40±23.13mmHg、813.10±325.30U/L，兩者的差異具有統(tǒng)計學(xué)意義（P0.05）。在DWI圖像上，隨著時間延長，對照組各時點異常高信號區(qū)出現(xiàn)范圍和強(qiáng)度較實驗組明顯，兩組各時點ADC值存在顯著性差異（P0.05）；將兩組動物的海馬、額頂區(qū)、側(cè)腦室旁區(qū)的石蠟切片進(jìn)行SP免疫組化染色并采用DAB顯色后，兩組的水通道蛋白4被不同程度著色，實驗組各時點AQP4蛋白表達(dá)陽性的細(xì)胞數(shù)明顯較對照組少（P0.05或0.01），且海馬、額頂區(qū)的AQP4蛋白表達(dá)比側(cè)腦室旁區(qū)多。兩組動物AQP4蛋白表達(dá)的變化趨勢大致相似，且與ADC值和DWI圖像各時點的變化情況相對應(yīng)。 結(jié)論：①AQP4基因沉默技術(shù)能顯著降低HIBD模型中新生豬腦組織早期水通道蛋白4的表達(dá)，，緩解臨床缺氧癥狀和體征，減輕腦組織水腫的程度，提示在新生兒缺血缺氧性腦損傷疾病中，水通道蛋白4可作為一個潛在的治療靶點。②DWI結(jié)合ADC圖/值可以較好的評價HIE/HIBD腦水腫的程度。
[Abstract]:Objective: neonatal hypoxic ischemic encephalopathy (hypoxic-ischemic, encephalopathy, HIE) or hypoxic ischemic brain damage (HIBD) is the main disease of neonatal brain injury caused by perinatal asphyxia and hypoxia, early hypoxic ischemic brain edema caused by asphyxia is the main pathogenesis of the disease. Clinical disease is common, which can cause factors of neonatal asphyxia, such as fetal distress, high forceps, birth trauma, can cause HIE, the disease is the main disease causing neonatal disability. According to the condition is divided into light, heavy, light, good prognosis, no residual sequelae; severe ischemia due to severe CNS hypoxia, can lead to nerve damage in early neonatal death or have permanent. In this study, neonatal pigs injected AQP4siRNA interference and the control sequence as the experimental object, the simulation of early neonatal HIBD model, through observation and comparison The difference between two groups of animals in clinical symptoms and signs, imaging findings and pathological findings is to explore the potential value of AQP4 gene silencing in early neonatal swine hypoxic ischemic brain damage, so as to provide new theoretical basis and direction for the diagnosis and treatment of neonatal HIBD disease.
Methods: 32 healthy and clean York newborn pigs were divided into experimental group and control group, administered through the cerebrospinal fluid circulation pathways, respectively AQP4siRNA interference sequence of intrathecal equivalent and the control sequence mixed solution. After 24h in the brain after transfection, the experimental group and the control group of neonatal pigs with the same degree of ischemia and hypoxia simulation, early neonatal HIBD model. By comparing the two groups of animal after hypoxic ischemia at each time point in the early neurological behavior, blood biochemical index, pathological differences in early MR molecular imaging and brain imaging, using the histogram or stacked bar chart on neurobehavioral scores of the two groups of animal and blood biochemistry index for direct comparison, and the comparison of 22 T test or non parametric test on the score and blood biochemical index at each time point, by using repeated measures ANOVA and T test method On the two animal groups at each time point DWI image ADC value, at the same time by immunohistochemical method (SP method) for detection of AQP4 protein positive cells of two groups of newborn piglets in expression level, to determine whether the difference between the two groups was significant (P0.05). The data were collected for statistical analysis by SPSS16.0 the statistical software package.
Results: the experimental group (i.e. injection AQP4siRNA interference sequence group) and control group (i.e. control group injected with AQP4siRNA sequence) of newborn pigs in spontaneous breathing, skin and mucosa color and neurological symptoms were abnormal in different degrees, breathing in the independent law and normal skin mucosa color but the experimental group recovered faster, in consciousness. Cranial nerve, reflection, action and coordination etc. neural function response to better performance; and the control group at HI after 2h, 3h, 6h, neurobehavioral scores were 7.200 + 0.789,10.000 + 1.700,13.400 + 1.174, significantly lower than the experimental group. The experimental group PCO2, 12h HI PO2, LDH = 34.77 + 9.40mmHg, 85.90 + 18.93mmHg, 586.22 + 111.58U/L group, the time of each index = 45.94 + 12.52mmHg control, 65.40 + 23.13mmHg, 813.10 + 325.30U/L, the difference was statistically significant (P0.05). In the DWI image, with the prolongation of time, The control group at each time point of abnormal high signal area range and intensity was significantly higher than the experimental group, there were significant differences between the two groups at each time point ADC value (P0.05); the two group animal hippocampus, frontal and parietal areas, paraffin periventricular area for immunohistochemical staining of SP and the DAB color, water channel protein two group of 4 different degrees of coloring, experimental group AQP4 protein expression positive cell number was significantly less than that of the control group (P0.05 or 0.01), and the hippocampus, frontal and parietal areas, the expression of AQP4 protein in the periventricular region. More than two groups of animal protein AQP4 expression showed similar changing trends, changes and with ADC and DWI images corresponding to each time point.
Conclusion: the expression of AQP4 gene silencing technology can significantly reduce the early water channel protein in neonatal brain tissue in HIBD model 4, relieve the clinical symptoms and signs of hypoxia, reduce brain edema, suggesting that in neonatal hypoxic ischemic brain injury disease, aquaporin 4 can be used as a potential therapeutic target. DWI combined with ADC map / value can better evaluate HIE/HIBD brain edema.

【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R-332;R722.1

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 石向群,楊金升,張志琳,王運良,包仕堯;水通道蛋白-4在正常大鼠腦內(nèi)的分布研究[J];中華神經(jīng)醫(yī)學(xué)雜志;2004年04期

2 周增杰,曾存芝;腦梗死患者血清CPK、LDH水平的臨床意義探討[J];神經(jīng)疾病與精神衛(wèi)生;2003年04期

3 彭雪華,孫善全;水通道蛋白-4在大鼠大腦中的定位研究[J];解剖學(xué)報;2004年02期

4 程英,劉國瑞,黃映宏,關(guān)計添,郭岳霖;新生豬缺氧缺血性腦病早期DWI影像表現(xiàn)的實驗研究[J];中國臨床醫(yī)學(xué)影像雜志;2004年10期

5 張琳;袁芳;;大鼠腦外傷誘導(dǎo)水通道蛋白4表達(dá)增強(qiáng)加重腦水腫[J];首都醫(yī)科大學(xué)學(xué)報;2008年06期

6 付雪梅;封志純;;RNA干擾介導(dǎo)新生大鼠星形膠質(zhì)細(xì)胞水通道蛋白-4基因沉默[J];中國循證兒科雜志;2006年04期

7 張均田;腦缺血、葡萄糖/能量代謝障礙與神經(jīng)退行性疾病[J];中國藥理學(xué)通報;2000年03期

8 孟繁崢,饒明俐;大鼠局灶性腦缺血水通道蛋白4的變化[J];中風(fēng)與神經(jīng)疾病雜志;2003年03期

9 魯宏,孫善全;水通道蛋白-4在急性腦缺血組織中的表達(dá)與MR擴(kuò)散加權(quán)成像的相關(guān)性研究[J];中華放射學(xué)雜志;2003年06期

10 魯宏,熊仁平,胡惠,趙建農(nóng),曾燕,余聰,淦偉,李杰,謝微波,倪衛(wèi)國,呂發(fā)金,程相晨;水通道蛋白-4在腦缺血半暗帶組織中的表達(dá)[J];中華放射學(xué)雜志;2005年06期

相關(guān)博士學(xué)位論文 前1條

1 王曉明;新生兒缺氧缺血性腦病的MR分子影像學(xué)的臨床與基礎(chǔ)研究[D];中國醫(yī)科大學(xué);2004年

本文編號：1476428