Notch-Pax9通路在酒精相关性口腔癌中的作用研究

发布时间：2018-03-09 01:17

  本文选题：口腔癌　切入点：酒精　出处：《首都医科大学》2017年博士论文　论文类型：学位论文

【摘要】：口腔癌是世界第六大癌症,近几年,包括口腔癌在内的上消化道恶性肿瘤的发病率在发展中国家呈上升趋势。流行病学研究表明饮酒是口腔癌危险因素之一,口腔癌的危险性及死亡率与饮酒呈剂量依赖。本实验通过临床样本研究和体内外实验发现在酒精相关性口腔癌中Pax9和Notch信号通路表达抑制,且两者间存在相关性。Notch-Pax9通路可能在酒精相关性口腔癌发展中发挥一定作用。1.Pax9和Notch通路在酒精相关性口腔癌中的表达临床样本研究:利用Tissue Array,免疫组化染色检测Pax9在口腔鳞癌和食道鳞癌中的阳性表达;从Duke大学获取人类食道鳞癌标本,通过免疫组化染色比较正常组织与食道鳞癌中Pax9和Notch通路下游基因的表达含量,以及饮酒者和非饮酒者组织中Pax9表达含量;利用GEO数据库分析正常组织与口腔鳞癌及食道鳞癌中Pax9和Notch通路下游基因的表达含量;各实验结果均显示相较于配对正常组织,口腔鳞癌及食道鳞癌样本中Pax9表达下降或缺失,Notch通路的下游效应分子Hes1表达呈现相同的下降趋势,与Pax9表达具有相关性。并且相较于非饮酒者,饮酒者的食道鳞癌组织中Pax9和Hes1表达量进一步降低。体外培养人上消化道鳞状上皮细胞KYSE510,给予不同浓度的乙醇处理,通过Western blotting和基因芯片技术检测酒精对上消化道上皮细胞Pax9和Notch信号通路的影响。实验结果显示酒精可以抑制Pax9和Notch信号通路的表达,并呈剂量依赖性。2.Pax9和Notch信号通路的相关性研究利用Ch IP实验检测小鼠舌,食道及前胃中Pax9和Notch信号通路的相关性,结果提示Pax9可能为Notch通路下游基因。建立Notch通路转录因子RBPJ基因敲除小鼠模型,结果显示Pax9在小鼠舌,食道和前胃上皮中均表达降低,与Notch通路下游效应基因表达一致,提示Pax9受Notch信号通路调控,可能为Notch信号通路的下游基因。用Notch抑制剂DBZ和Notch激活剂Jagged1分别处理KYSE510,检测Pax9表达变化。结果显示Pax9与Notch通路的表达趋势一致。并且酒精可以拮抗Notch激活剂Jagged1对Notch-Pax9的激活作用。3.Pax9敲除对小鼠实验性口腔癌的作用研究建立Pax9基因敲除小鼠模型,利用免疫组化染色和荧光免疫组化染色检测小鼠食道上皮细胞增殖能力变化。结果显示Pax9基因敲除促进小鼠食道上皮细胞增殖,干扰细胞分化。给予Pax9基因敲除小鼠NMBA,研究Pax9敲除是否影响小鼠口腔癌及食道癌发生率。实验结果显示Pax9敲除后可以促进NMBA诱导的小鼠口腔癌和食道癌的发生率。综上所述,Notch信号通路和Pax9在酒精相关性口腔癌和食道癌中表达下调,Pax9可能受Notch信号通路调控,为Notch信号通路的下游基因。Pax9基因敲除会促进小鼠食道上皮细胞增殖,干扰分化功能,促进小鼠口腔癌和食道癌的发生率。Notch-Pax9表达抑制可能是酒精相关性口腔癌的分子机制之一。
[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.
【学位授予单位】：首都医科大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R739.8

【相似文献】

相关期刊论文 前10条

1 张燕;郑志z，

本文编号：1586369