【摘要】：第一部分構(gòu)建經(jīng)口主動吸煙動物模型[目的]吸煙對呼吸系統(tǒng)、消化系統(tǒng)、生殖系統(tǒng)、感官系統(tǒng)等多系統(tǒng)造成危害,可導致各種疾病的發(fā)生、發(fā)展。為了研究煙草與人類疾病的相關性,吸煙相關疾病模型的建立及完善越來越受到人們的關注,目前吸煙相關疾病模型主要包括臨床樣本吸煙模型、體外吸煙細胞模型和吸煙動物模型三類。但迄今為止,有關主動吸煙動物模型方面的研究鮮少,經(jīng)口主動吸煙動物模型更未見國內(nèi)外文獻報道。本實驗的目的是構(gòu)建經(jīng)口主動吸煙動物模型。[方法]Wistar大鼠20只,主動吸煙組和非吸煙組各10只,雌雄各半。主動吸煙組大鼠采用主動吸煙裝置,建立大鼠經(jīng)口主動吸煙模型。主動吸煙組大鼠每只連續(xù)經(jīng)口吸煙3個月,每日兩次,每次0.5小時。非吸煙組大鼠由經(jīng)口裝置吸空氣,時間和頻率與主動吸煙組相同。觀察大鼠吸煙成癮性、口腔黏膜外觀的改變、神經(jīng)行為學改變和體重的變化等。組化技術(shù)(HE染色)檢查口腔黏膜的組織病理改變。[結(jié)果]主動吸煙組大鼠約2周時間吸煙成癮,3個月時大體觀可見口腔黏膜上皮顏色變?yōu)榛野咨?質(zhì)地粗糙,彈性降低。經(jīng)統(tǒng)計學分析,體重較非吸煙組顯著降低�？谇徽衬づR床大體觀及組織病理學檢查顯示,主動吸煙組大鼠口腔黏膜發(fā)生白斑,發(fā)生率為75%。[結(jié)論]本實驗構(gòu)建了經(jīng)口主動吸煙動物(大鼠)模型,主動吸煙造成大鼠口腔黏膜大體和病理學改變,誘導口腔白斑的發(fā)生。第二部分 主動吸煙對大鼠口腔黏膜上皮細胞NOD1信號通路的影響[目的]口腔黏膜上皮細胞中NOD1信號通路是口腔黏膜固有免疫的重要組成部分,經(jīng)本課題組前期體外實驗和人體組織標本研究發(fā)現(xiàn),吸煙可影響NOD1信號通路關鍵蛋白的表達,提示吸煙可抑制NOD1通路,降低口腔黏膜局部免疫�；谇捌谘芯�,本實驗在構(gòu)建經(jīng)口主動吸煙動物模型的基礎上,研究經(jīng)口主動吸煙對口腔黏膜上皮細胞NOD1信號通路的影響。[方法]通過免疫印跡和免疫組化技術(shù)及圖像分析法,對主動吸煙組與非吸煙組大鼠頰粘膜上皮細胞NOD1、RIP2、NF-κB、P-NF-κB (phospho-NF-κB,磷酸化NF-κB)蛋白表達進行檢測,比較兩組表達水平的差異。[結(jié)果]相對于非吸煙組,主動吸煙組NOD1和NF-κB的蛋白表達水平下降,RIP2、和P-NF-κB的蛋白表達水平上升。[結(jié)論]主動吸煙可導致口腔黏膜上皮細胞中NOD1信號通路中關鍵蛋白的改變,結(jié)果顯示,主動吸煙對口腔黏膜上皮細胞中NOD1信號通路產(chǎn)生了影響。
[Abstract]:The first part is to construct the animal model of active smoking through mouth [objective] smoking causes harm to respiratory system, digestive system, reproductive system, sensory system and so on, which can lead to the occurrence and development of various diseases. In order to study the relationship between tobacco and human diseases, more and more attention has been paid to the establishment and improvement of smoking-related disease models. At present, the models of smoking-related diseases mainly include the smoking models of clinical samples. In vitro smoking cell model and smoking animal model. But so far, there are few studies on active smoking animal models, and there is no domestic and foreign literature on active smoking animal models. The purpose of this study was to establish an animal model of active smoking via mouth. [methods] Twenty Wistar rats were divided into active smoking group (n = 10) and non smoking group (n = 10). The active smoking model was established by using active smoking device in the active smoking group. Rats in active smoking group smoked twice a day for 0.5 hours twice a day for 3 months. The time and frequency of inhaling air in non-smoking group was the same as that in active smoking group. The changes of smoking addiction, oral mucosa appearance, neurobehavioral changes and body weight were observed. The histopathological changes of oral mucosa were examined by HE staining. [results] in the active smoking group, the rats were addicted to smoking for about 2 weeks. At 3 months, the color of oral mucosal epithelium became gray and white, the texture was rough, and the elasticity was decreased. By statistical analysis, the body weight was significantly lower than that of non-smoking group. The clinical gross and histopathological examination of oral mucosa showed that leukoplakia was found in oral mucosa of active smoking group (75%). [conclusion] the animal model of active oral smoking was established in this experiment. The oral mucosa gross and pathological changes were induced by active smoking, and the occurrence of oral leukoplakia was induced. Part II effects of active smoking on NOD1 signaling pathway in rat oral mucosal epithelial cells [objective] NOD1 signaling pathway is an important part of oral mucosal innate immunity in rats. Previous experiments in vitro and human tissue samples showed that smoking could affect the expression of key proteins in NOD1 signaling pathway, suggesting that smoking could inhibit NOD1 pathway and reduce local immunity of oral mucosa. Based on the previous studies, the effects of oral active smoking on NOD1 signaling pathway in oral epithelial cells were studied based on the establishment of oral active smoking animal model. [methods] NOD1,RIP2,NF- 魏 B P-NF- 魏 B (phospho-NF- 魏 B) in buccal mucosal epithelial cells of active smoking and non-smoking rats were studied by Western blot, immunohistochemistry and image analysis. The expression of phosphorylated NF- 魏 B was detected and the differences between the two groups were compared. [results] compared with non-smoking group, the expression of NOD1 and NF- 魏 B protein decreased in active smoking group, and the protein expression level of RIP2, and P-NF- 魏 B increased in active smoking group. [conclusion] active smoking can result in the change of key proteins in NOD1 signaling pathway in oral mucosal epithelial cells. The results show that active smoking has an effect on NOD1 signaling pathway in oral mucosal epithelial cells.
【學位授予單位】：南京大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R781

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