mTOR通路调节PKM2影响食管鳞癌细胞有氧糖酵解机制的研究

发布时间：2018-03-29 14:44

本文选题：食管鳞癌　切入点：PKM2　出处：《山西医科大学》2017年博士论文

【摘要】：食管鳞状细胞癌(ESCC)是一种地区发病率高、治愈率低、生存率低,且预后差的恶性肿瘤。其代谢、增殖与转移涉及众多信号通路。这些信号通路相互交叉连接,形成复杂的信号通路网,使得食管鳞癌的治疗效果并不如预期。文献表明,通过调节食管鳞癌组织的能量代谢来控制肿瘤细胞的生长,可以作为一种新的肿瘤治疗方法进行探索和研究,对开发新的治疗策略具有非常重要的意义。已有研究表明,不同于正常组织细胞,肿瘤组织细胞无论氧气充足还是缺氧,都采用糖酵解的方式获取能量,即有氧糖酵解。M2型丙酮酸激酶(PKM2)是肿瘤细胞有氧糖酵解过程中最后一步的关键酶,是它将磷酸烯醇式丙酮酸(PEP)分解为丙酮酸,从而使各种癌细胞获取能量。研究表明,PKM2不仅在肿瘤细胞代谢中发挥重要作用,而且在肿瘤细胞的增殖、转化以及预后中也是重要的信号分子,但是PKM2在肿瘤细胞中被调节的机制有待进一步研究。m TOR信号通路调节细胞功能,影响蛋白质的合成,在多种恶性肿瘤中发现其高度活跃。其下游信号分子可调节多种转录因子影响肿瘤细胞中关键酶的形成。在肝癌和肾脏肿瘤中发现,m TOR可以调节PKM2的表达。所以,设计本课题,观察在食管鳞癌中m TOR信号通路是否可以调节PKM2的表达,以及是否可以以此影响肿瘤细胞的能量代谢。首先通过病理切片的免疫组化染色确定在食管鳞癌中m TOR和PKM2的表达均上调。通过改变m TOR和PKM2的表达量来研究其对食管鳞癌细胞KYSE150能量代谢的影响,证明m TOR可以调节PKM2进而影响食管鳞癌细胞的有氧糖酵解,为开发食管鳞癌新的治疗策略,提供依据和思路。目的:1、研究PKM2对食管鳞癌细胞有氧糖酵解的影响。2、研究m TOR信号通路对食管鳞癌细胞有氧糖酵解的影响。3、研究在食管鳞癌细胞中m TOR通路调节PKM2影响有氧糖酵解代谢的机制。方法:第一部分:食管鳞癌患者组织样本及数据分析为验证食管鳞癌患者组织中PKM2及p-m TOR的表达情况,选取食管鳞癌及非肿瘤食管组织样本各30例,并充分收集患者的临床病理组织及资料进行免疫组化实验,检测PKM2及p-m TOR在食管鳞癌及非肿瘤食管组织中的表达情况。第二部分:PKM2的表达影响食管鳞癌细胞的有氧糖酵解1、检测PKM2下调对食管鳞癌细胞KYSE150葡萄糖消耗和乳酸生成的影响,并与非肿瘤食管组织进行对比。2、过表达食管鳞癌细胞系KYSE150细胞中的PKM2,检测PKM2过表达对食管鳞癌细胞葡萄糖消耗和乳酸生成情况比较的影响。第三部分:抑制或者活化m TOR信号通路对食管鳞癌细胞有氧糖酵解的影响1、m TOR抑制剂雷帕霉素作用于KYSE150细胞,观察其对m TOR信号通路以及对细胞有氧糖酵解的影响。2、下调m TOR抑制基因PTEN对m TOR信号通路以及细胞有氧糖酵解的影响。第四部分:在食管鳞癌KYSE150细胞系中,m TOR通路通过对PKM2的调节影响KYSE150细胞的有氧糖酵解1、m TOR抑制剂雷帕霉素作用于KYSE150细胞对PKM2表达量的影响。2、抑制基因PTEN抑制组对PKM2表达以及细胞糖酵解作用的影响。3、PKM2被敲低后,抑制PTEN基因(即活化m TOR通路)也不能上调食管鳞癌细胞的有氧糖酵解作用。结果:1、m TOR和PKM2在食管鳞癌组织中表达升高。2、PKM2在食管鳞癌细胞代谢中对有氧糖酵解有调节作用。3、m TOR在食管鳞癌细胞代谢中对有氧糖酵解有调节作用。4、在食管鳞癌细胞中m TOR通路通过调节PKM2影响有氧糖酵解。结论:本实验初步阐明PKM2和m TOR在食管鳞癌组织中均呈现高表达状态,m TOR在食管鳞癌细胞中可以调节PKM2,继而影响食管鳞癌细胞的有氧糖酵解,为食管鳞癌的治疗提供了新的策略。
[Abstract]:Esophageal squamous cell carcinoma (ESCC) is an area of high incidence, low cure rate, low survival rate and poor prognosis of malignant tumor. Its metabolism, proliferation and metastasis involves many signaling pathways. These pathways cross connect with each other, forming a complex network of signaling pathways, the therapeutic effect of esophageal squamous cell carcinoma is not as expected. Literature shows that, through the regulation of energy metabolism in human esophageal squamous cell carcinoma to control the growth of tumor cells, can be used as a new tumor treatment methods of exploration and research, has very important significance for the development of new therapeutic strategies. Studies have shown that different from normal cells, tumor cells regardless of sufficient oxygen or oxygen and use glycolysis way to obtain energy, namely the aerobic glycolysis of.M2 pyruvate kinase (PKM2) is a key enzyme in tumor cells of oxygen during glycolysis is the last step, it will be allyl phosphate Alcohol type pyruvate (PEP) decomposition of pyruvate, so that a variety of cancer cells to obtain energy. The results show that PKM2 not only play an important role in the metabolism of tumor cells, and the proliferation of tumor cells, but also an important signal molecule transformation and prognosis, but the mechanism of PKM2 is regulated in tumor cells for further study of.M TOR the signaling pathways that regulate cell function, affect protein synthesis, the highly active in a variety of malignant tumors. The downstream signaling molecules can regulate many transcription factors influence the formation of key enzymes in tumor cells. Found in liver cancer and kidney cancer, the expression of M TOR PKM2 can be adjusted. Therefore, the subject of the design, to observe the expression in esophageal squamous cell carcinoma m TOR signal pathway can regulate PKM2, and whether this can affect energy metabolism of tumor cells. The immunohistochemical staining of pathological sections To determine the expression of M TOR and PKM2 were up-regulated in esophageal squamous cell carcinoma. The expression change of M TOR and PKM2 to study its effect on esophageal squamous cell carcinoma KYSE150 cell energy metabolism, TOR proved that M can regulate PKM2 and esophageal squamous cell carcinoma of aerobic glycolysis, as a new strategy of development of esophageal squamous cell carcinoma treated with the basis and ideas. Objective: 1. Study on PKM2 effect of aerobic glycolysis of.2 for esophageal squamous cell carcinoma, m TOR signal pathway effects of aerobic glycolysis of.3 for esophageal squamous cell carcinoma, research in esophageal squamous cell carcinoma m TOR pathway in regulation of PKM2 can influence the mechanism of oxygen glycolytic metabolism. Methods: the first part: the sample and data analysis for patients with esophageal squamous cell carcinoma tissue expression validation of esophageal squamous cell carcinoma PKM2 and P-M TOR, selection of esophageal squamous cell carcinoma and non esophageal tumor tissue samples of the 30 cases, and the full collection of clinical and pathological tissues of patients Data and immunohistochemical experiments, to detect the expression of PKM2 and P-M TOR in esophageal squamous cell carcinoma and non tumor tissues. The second part: the effect of PKM2 expression in esophageal squamous cell carcinoma of aerobic glycolysis 1, detection of PKM2 downward effect on esophageal squamous cell carcinoma cell line KYSE150 glucose consumption and lactic acid, and compared.2 and esophageal tissue non tumor, overexpression of KYSE150 cells in esophageal squamous cell carcinoma PKM2 cell line, detect the effects of PKM2 overexpression on esophageal squamous cell carcinoma cell line comparison of glucose consumption and lactic acid production situation. The third part: the inhibition or activation of M TOR signaling pathway in esophageal squamous cell carcinoma cells effect of aerobic glycolysis inhibitor 1, m TOR the effect of rapamycin on KYSE150 cells and observe the M and TOR signal pathway on cell effect of aerobic glycolysis of.2, down-regulation of M TOR gene PTEN aerobic glycolysis effect on M cells and TOR signal pathway Ring. The fourth part: in esophageal squamous cell carcinoma cell line KYSE150, to regulate the PKM2 effect of KYSE150 cells in aerobic glycolysis 1 through the m TOR pathway, m TOR inhibitor rapamycin in KYSE150 cells the expression of.2 on the PKM2 effect of.3 suppressor gene PTEN inhibited group solution effect on PKM2 expression and cell sugar fermentation, knockdown of PKM2 inhibited PTEN gene (i.e., activation of M TOR pathway in esophageal squamous cell carcinoma) also can not increase the aerobic glycolysis. Results: 1 the expression of M, TOR and PKM2 in esophageal squamous cell carcinoma increased.2, PKM2 in esophageal squamous cell carcinoma on the metabolism of aerobic glycolysis is the role of.3 in regulation of M, TOR in esophageal squamous cell carcinoma on the metabolism of aerobic glycolysis of the regulatory role of.4 in esophageal squamous cell carcinoma m TOR pathway by regulating the PKM2 effect of aerobic glycolysis. Conclusion: the PKM2 and m TOR were preliminarily showed high expression in esophageal squamous cell carcinoma. State of state, m TOR can regulate PKM2 in esophageal squamous cell carcinoma cells, and then influence the aerobic glycolysis of esophageal squamous cell carcinoma cells, which provides a new strategy for the treatment of esophageal squamous cell carcinoma.

【学位授予单位】：山西医科大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R735.1

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