TLR4信号通路参与人结肠癌细胞免疫逃逸机制的研究

发布时间：2018-01-04 16:23

本文关键词：TLR4信号通路参与人结肠癌细胞免疫逃逸机制的研究　出处：《武汉大学》2009年博士论文　论文类型：学位论文

【摘要】：慢性感染和炎症可以导致肿瘤的发生、发展,但其机制并不太清楚。许多研究发现在此过程中天然免疫系统发挥了重要的作用,而在天然免疫过程中起决定性作用的Toll样受体(Toll-like receptors, TLRs)家族在肿瘤中的作用也越来越受到人们的重视。 TLRs最早在果蝇中发现,起初人们认为其对节肢动物胚胎发育时的倾向性具有重要作用。随着研究的进一步进展,人们发现其具有对抗病原微生物感染的能力。1997年Medzhitov等发现了人类的第一个TLR(后来被命名为TLR4)。接着,人类的其他TLRs相继被发现。迄今为止,在人类已经发现了11个TLRs成员,对它们相应的配体也有所了解。TLRs主要表达于免疫细胞尤其是巨噬细胞和树突状细胞表面,通过广泛地特异性地识别病原体相关的分子模式(pathogen-associated molecular patterns, PAMPs),偶联信号传导途径,激活免疫细胞分泌大量炎性细胞因子和趋化因子,发挥机体的抗病原微生物感染能力,构成机体免疫反应的第一道防线。TLRs不仅能激活天然免疫,而且也为激活获得性免疫提供共刺激信号,是天然免疫与获得性免疫两者之间的桥梁。因此,TLRs的功能及其作用信号转导机制的研究具有重要的理论意义和应用价值。当人们在研究免疫细胞表面TLRs的结构、功能及信号转导途径的时候,却发现多种肿瘤细胞表面也表达多种TLRs,并且可能与肿瘤细胞的免疫逃逸相关。本课题主要目的是检测TLR4在人结肠癌细胞的表达,检测结肠癌细胞表达的TLR4是否参与结肠癌细胞免疫逃逸及其可能的机制。我们选取了人结肠癌细胞HT-29及SW480,发现TLR4在nRNA与蛋白水平均有表达。接下来,我们用TLR4外源性配体LPS作为激活剂,观察TLR4信号转导通路是否活化以及活化后细胞生物学活性的改变。结果表明,LPS刺激HT-29与SW480细胞后,NF-κB及MAPK信号转导通路被激活,而且IL-8分泌水平增加,表明HT-29与SW480表达的TLR4具有功能活性,能被LPS激活。同时HT-29与SW480可以自发分泌一定水平的TGF-β、VEGF,LPS可通过TLR4增强TGF-β、VEGF及IL-8的分泌。进一步研究发现,LPS刺激对HT-29与SW480细胞的增殖无影响,但对TRAIL诱导的细胞凋亡有耐受作用。表明TLR4信号活化可以促进人结肠癌细胞逃逸机体的免疫攻击,有利于肿瘤的发生发展。总之,本研究发现人结肠癌细胞HT-29与SW480表达TLR4, TLR4配体LPS刺激可以促进人结肠癌细胞分泌免疫抑制性细胞因子TGF-β、VEGF及IL-8,并提高结肠癌细胞对TRAIL诱导的凋亡的耐受作用,这一作用可能与NF-κB信号通路被激活有关。本实验结果将有助于了解TLR4信号途径活化后在结肠癌细胞中的作用,对TLR4信号在肿瘤生物学和肿瘤免疫学的功能增添了新的认识,并为结肠癌的治疗提供了新靶点。
[Abstract]:Chronic infection and inflammation can lead to tumorigenesis and progression, but its mechanism is not very clear. Many studies have found that innate immune system plays an important role in this process. Toll-like receptors, a Toll like receptor that plays a decisive role in the innate immune process. The role of TLRs family in tumor has been paid more and more attention. TLRs was first found in Drosophila, initially thought to play an important role in the developmental tendency of arthropod embryos. In 1997, Medzhitov et al discovered the first human TLRs (later named TLR4). Other human TLRs have been discovered. So far, 11 TLRs members have been discovered in humans. It is also known that the corresponding ligands. TLRs are mainly expressed on the surface of immune cells, especially macrophages and dendritic cells. Pathogen-associated molecular patterns (PAMPs) are widely and specifically identified by pathogen-associated molecular patterns. The coupling signal transduction pathway activates the immune cells to secrete a large number of inflammatory cytokines and chemokines to exert the ability of anti-pathogenic microorganism infection. TLRs can not only activate innate immunity, but also provide costimulatory signal for activation of acquired immunity, which is the bridge between innate immunity and acquired immunity. The study of the function and signal transduction mechanism of TLRs has important theoretical significance and application value. When people are studying the structure, function and signal transduction pathway of TLRs on immune cells, it is found that many kinds of TLRs are also expressed on the surface of many tumor cells. The main purpose of this study is to detect the expression of TLR4 in human colon cancer cells. To determine whether the TLR4 expressed in colon cancer cells is involved in the immune escape of colon cancer cells and its possible mechanism. We selected human colon cancer cells HT-29 and SW480 and found that TLR4 was expressed at both nRNA and protein levels. Then we used TLR4 exogenous ligand LPS as activator. The activation of TLR4 signal transduction pathway and the changes of cell biological activity after activation were observed. The results showed that LPS-stimulated HT-29 and SW480 cells. NF- 魏 B and MAPK signal transduction pathway were activated, and the level of IL-8 secretion increased, indicating that TLR4 expressed by HT-29 and SW480 had functional activity. At the same time, HT-29 and SW480 could spontaneously secrete a certain level of TGF- 尾. VEGF could enhance TGF- 尾 through TLR4. The secretion of VEGF and IL-8. Further study showed that LPS-stimulated proliferation of HT-29 and SW480 cells had no effect. The results showed that the activation of TLR4 signal could promote the immune attack of human colon cancer cells, which was beneficial to the occurrence and development of tumor. In conclusion, we found that human colon cancer cells HT-29 and SW480 express TLR4. TLR4 ligand LPS stimulation can promote the secretion of immunosuppressive cytokines TGF- 尾 -VEGF and IL-8 in human colon cancer cells. And to improve the tolerance of colon cancer cells to TRAIL induced apoptosis. This effect may be related to the activation of NF- 魏 B signaling pathway. Our results may be helpful to understand the role of TLR4 signaling pathway activation in colon cancer cells. New understanding of the function of TLR4 signal in tumor biology and tumor immunology has been added, and a new target for the treatment of colon cancer has been provided.
【学位授予单位】：武汉大学
【学位级别】：博士
【学位授予年份】：2009
【分类号】：R392;R735.35

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