本文選題：口腔癌　切入點：酒精　出處：《首都醫(yī)科大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：口腔癌是世界第六大癌癥,近幾年,包括口腔癌在內(nèi)的上消化道惡性腫瘤的發(fā)病率在發(fā)展中國家呈上升趨勢。流行病學(xué)研究表明飲酒是口腔癌危險因素之一,口腔癌的危險性及死亡率與飲酒呈劑量依賴。本實驗通過臨床樣本研究和體內(nèi)外實驗發(fā)現(xiàn)在酒精相關(guān)性口腔癌中Pax9和Notch信號通路表達(dá)抑制,且兩者間存在相關(guān)性。Notch-Pax9通路可能在酒精相關(guān)性口腔癌發(fā)展中發(fā)揮一定作用。1.Pax9和Notch通路在酒精相關(guān)性口腔癌中的表達(dá)臨床樣本研究:利用Tissue Array,免疫組化染色檢測Pax9在口腔鱗癌和食道鱗癌中的陽性表達(dá);從Duke大學(xué)獲取人類食道鱗癌標(biāo)本,通過免疫組化染色比較正常組織與食道鱗癌中Pax9和Notch通路下游基因的表達(dá)含量,以及飲酒者和非飲酒者組織中Pax9表達(dá)含量;利用GEO數(shù)據(jù)庫分析正常組織與口腔鱗癌及食道鱗癌中Pax9和Notch通路下游基因的表達(dá)含量;各實驗結(jié)果均顯示相較于配對正常組織,口腔鱗癌及食道鱗癌樣本中Pax9表達(dá)下降或缺失,Notch通路的下游效應(yīng)分子Hes1表達(dá)呈現(xiàn)相同的下降趨勢,與Pax9表達(dá)具有相關(guān)性。并且相較于非飲酒者,飲酒者的食道鱗癌組織中Pax9和Hes1表達(dá)量進一步降低。體外培養(yǎng)人上消化道鱗狀上皮細(xì)胞KYSE510,給予不同濃度的乙醇處理,通過Western blotting和基因芯片技術(shù)檢測酒精對上消化道上皮細(xì)胞Pax9和Notch信號通路的影響。實驗結(jié)果顯示酒精可以抑制Pax9和Notch信號通路的表達(dá),并呈劑量依賴性。2.Pax9和Notch信號通路的相關(guān)性研究利用Ch IP實驗檢測小鼠舌,食道及前胃中Pax9和Notch信號通路的相關(guān)性,結(jié)果提示Pax9可能為Notch通路下游基因。建立Notch通路轉(zhuǎn)錄因子RBPJ基因敲除小鼠模型,結(jié)果顯示Pax9在小鼠舌,食道和前胃上皮中均表達(dá)降低,與Notch通路下游效應(yīng)基因表達(dá)一致,提示Pax9受Notch信號通路調(diào)控,可能為Notch信號通路的下游基因。用Notch抑制劑DBZ和Notch激活劑Jagged1分別處理KYSE510,檢測Pax9表達(dá)變化。結(jié)果顯示Pax9與Notch通路的表達(dá)趨勢一致。并且酒精可以拮抗Notch激活劑Jagged1對Notch-Pax9的激活作用。3.Pax9敲除對小鼠實驗性口腔癌的作用研究建立Pax9基因敲除小鼠模型,利用免疫組化染色和熒光免疫組化染色檢測小鼠食道上皮細(xì)胞增殖能力變化。結(jié)果顯示Pax9基因敲除促進小鼠食道上皮細(xì)胞增殖,干擾細(xì)胞分化。給予Pax9基因敲除小鼠NMBA,研究Pax9敲除是否影響小鼠口腔癌及食道癌發(fā)生率。實驗結(jié)果顯示Pax9敲除后可以促進NMBA誘導(dǎo)的小鼠口腔癌和食道癌的發(fā)生率。綜上所述,Notch信號通路和Pax9在酒精相關(guān)性口腔癌和食道癌中表達(dá)下調(diào),Pax9可能受Notch信號通路調(diào)控,為Notch信號通路的下游基因。Pax9基因敲除會促進小鼠食道上皮細(xì)胞增殖,干擾分化功能,促進小鼠口腔癌和食道癌的發(fā)生率。Notch-Pax9表達(dá)抑制可能是酒精相關(guān)性口腔癌的分子機制之一。
[Abstract]:Oral cancer is one of the 6th largest cancers in the world. In recent years, the incidence of malignant tumors in the upper digestive tract, including oral cancer, is on the rise in developing countries. Epidemiological studies have shown that alcohol consumption is one of the risk factors for oral cancer. The risk and mortality of oral cancer were dose-dependent. In this study, the expression of Pax9 and Notch signal pathway was inhibited in alcoholic oral carcinoma by clinical and in vivo experiments. Notch-Pax9 pathway may play a role in the development of alcohol-related oral carcinoma. 1. The expression of Pax9 and Notch pathway in alcohol-related oral carcinoma: using Tissue Arrayand immunohistochemical staining to detect Pax9. Positive expression in oral squamous cell carcinoma and esophageal squamous cell carcinoma; Human esophageal squamous cell carcinoma (OSCC) samples were obtained from Duke University. The expression of Pax9 and Notch downstream genes in normal tissues and esophageal squamous cell carcinomas (OSCC) and Pax9 expression in drinkers and non-drinkers were compared by immunohistochemical staining. GEO database was used to analyze the downstream gene expression of Pax9 and Notch pathway in normal tissue, oral squamous cell carcinoma and esophageal squamous cell carcinoma. In oral squamous cell carcinoma (OSCC) and esophageal squamous cell carcinoma (OSCC), the expression of Hes1, the downstream effector molecule of Notch pathway, decreased in the same trend, and was correlated with the expression of Pax9 in oral squamous cell carcinoma (OSCC) and esophageal squamous cell carcinoma (OSCC). The expression of Pax9 and Hes1 in esophageal squamous cell carcinoma (OSCC) was further decreased. KYSE510cells were cultured in vitro and treated with different concentrations of ethanol. The effects of alcohol on Pax9 and Notch signaling pathway in upper gastrointestinal epithelial cells were detected by Western blotting and gene chip technique. The results showed that alcohol could inhibit the expression of Pax9 and Notch signaling pathway. The correlation between Pax9 and Notch signaling pathway in tongue, esophagus and forestomach of mice was detected by Ch IP experiment. The results suggested that Pax9 might be the downstream gene of Notch pathway. The expression of Pax9 in tongue, esophagus and forestomach of mice was decreased, which was consistent with the downstream effect gene expression of Notch pathway. The results suggest that Pax9 is regulated by Notch signaling pathway. KYSE510 was treated with Notch inhibitor DBZ and Notch activator Jagged1 to detect the expression of Pax9. The results showed that the expression trend of Pax9 was the same as that of Notch pathway, and alcohol could antagonize the expression of Notch activator Jagged1. Activation of Notch-Pax9. 3. Effects of Pax9 knockout on experimental oral cancer in mice; The proliferative ability of mouse esophageal epithelial cells was detected by immunohistochemical staining and fluorescence immunohistochemical staining. The results showed that Pax9 gene knockout promoted the proliferation of mouse esophageal epithelial cells. Pax9 knockout mice were given Pax9 gene knockout mice to study whether Pax9 knockout could affect the incidence of oral and esophageal cancer in mice. The results showed that Pax9 knockout could promote the incidence of oral and esophageal cancer induced by NMBA in mice. In conclusion, the down-regulation of Pax9 and Notch signaling pathway and Pax9 expression in alcohol-related oral carcinoma and esophageal carcinoma may be regulated by Notch signaling pathway. Knockout of the downstream gene. Pax9 gene of Notch signaling pathway can promote the proliferation of mouse esophageal epithelial cells, interfere with differentiation function, and promote the inhibition of the expression of .Notch-Pax9 in mouse oral and esophageal carcinomas, which may be one of the molecular mechanisms of alcohol-related oral carcinoma.
【學(xué)位授予單位】：首都醫(yī)科大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R739.8

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