巨噬细胞在肝脏肿瘤起始细胞存活和肝癌发生中的关键作用研究

发布时间：2018-03-21 09:47

本文选题：Hippo通路　切入点：肿瘤起始细胞　出处：《浙江大学》2016年博士论文　论文类型：学位论文

【摘要】：免疫系统和炎症在肿瘤发生的每个阶段都有着很重要的作用。在已形成的肿瘤里,浸润的炎细胞通过与肿瘤细胞相互作用,改变肿瘤微环境从而影响肿瘤发展。肿瘤里的各种炎细胞及其与肿瘤细胞的相互作用是设计有效免疫治疗的理论基础。肝脏肿瘤在中国和世界范围内对健康都是一个巨大的威胁。然而由于肝癌遗传背景的复杂性至今还缺少准确模拟肝脏肿瘤的小鼠遗传学模型。人肝脏肿瘤的发生往往伴随着炎症反应,然而在炎性微环境里产生的肿瘤起始细胞是否主动改造免疫微环境尚不清楚。Hippo信号通路是近年来发现的调控器官大小的关键信号途径,它的异常与癌症发生特别是肝癌密切相关。为了在体内研究炎症细胞对肝脏肿瘤起始细胞的影响,特别是由Hippo通路失调引起的肝脏肿瘤,我建立了一个流体动力学注射和转座子结合的小鼠肝癌模型。利用这种模型,在成体小鼠肝实质细胞内稳定过表达激活型的Hippo通路效应分子YAP,在4个月内就可以强烈诱导小鼠肝脏肿瘤生成。我们发现YAP的抑制蛋白VGLL4可以有效抑制YAP引起的肿瘤,而YAP的下游分子,miR-130a可以通过靶向VGLL4增强YAP诱导的肝脏肿癌形成。这一研究揭示了 Hippo信号通路调控肿瘤发生的正反馈回路。我意外地发现在YAP诱导肿瘤生成的早期,甚至是单个细胞的肿瘤起始细胞阶段就能很强地招募巨噬细胞。这些巨噬细胞具有肿瘤相关巨噬细胞的性质。敲除Hippo通路的激酶Mst1/2或Lats1/2从而激活内源性YAP也可以招募巨噬细胞。在肿瘤起始细胞里,YAP可以上调趋化因子Ccl2和Csf1的表达来招募巨噬细胞。YAP通过TEAD对Ccl2实现直接调控。通过敲减Ccl2和Csf1不仅有效阻止巨噬细胞的招募,还能导致肿瘤起始细胞被免疫清除。这种清除依赖于适应性免疫系统,并且通过p53依赖的途径。肿瘤起始细胞的清除最终导致了 YAP不能诱导肿瘤生成。在人肝癌的癌前病变中和其他原癌基因活化型AKT和EGFR诱导形成的肿瘤起始细胞里,YAP的活化也伴随着巨噬细胞的招募。以上结果首先建立了一种快速、灵活,与人类疾病相关的肝癌小鼠模型,可广泛用于研究肝癌信号转导途径以及肝癌细胞与微环境的相互作用。更重要的是本研究揭示了 Hippo通路在肝脏肿瘤形成过程中的一种新的非细胞自身作用的机制,即可以通过调控肿瘤微环境从而促进肿瘤发生。并且我们发现肿瘤起始细胞早在单个细胞阶段就可以通过主动招募巨噬细胞来塑造免疫抑制性微环境。这提示了一种新的免疫预防的方法,即通过抑制YAP活性或肿瘤起始细胞相关巨噬细胞达到清除肿瘤起始细胞预防肿瘤发生的效果。
[Abstract]:The immune system and inflammation play an important role in every stage of tumorigenesis. In formed tumors, infiltrating inflammatory cells interact with tumor cells. Changing the microenvironment of the tumor affects the development of the tumor. Various inflammatory cells in the tumor and their interaction with the tumor cells are the theoretical basis for the design of effective immunotherapy. Liver neoplasms are considered to be healthy in both China and the world. However, due to the complexity of the genetic background of liver cancer, there is still no genetic model of mice that can accurately simulate the liver tumor. The occurrence of human liver tumor is often accompanied by inflammatory reaction. However, it is not clear whether tumor initiation cells in inflammatory microenvironment actively modify the immune microenvironment. Hippo signaling pathway is the key signal pathway to regulate organ size found in recent years. In order to study the effect of inflammatory cells on liver tumor initiation cells in vivo, especially liver tumors caused by Hippo pathway imbalance, I have established a hydrodynamic injection and transposon binding model of liver cancer in mice. In adult mouse liver parenchyma cells, Yap, a stable overexpressed Hippo pathway effector molecule, can strongly induce tumor formation in mouse liver within 4 months. We found that VGLL4, the inhibitory protein of YAP, can effectively inhibit tumor induced by YAP. The downstream molecule of YAP, miR-130a, can enhance liver tumorigenesis induced by YAP by targeting VGLL4. This study reveals that the Hippo signaling pathway regulates the positive feedback circuit in tumorigenesis. I accidentally found that in the early stage of YAP induced tumorigenesis, Even the tumor initiation phase of a single cell can strongly recruit macrophages. These macrophages have the properties of tumor-associated macrophages. Knockout kinase Mst1/2 or Lats1/2 of the Hippo pathway can also activate endogenous YAP. The expression of chemokines Ccl2 and Csf1 can be upregulated in tumor initiation cells, which can directly regulate Ccl2 by TEAD. Not only can Ccl2 and Csf1 inhibit the recruitment of macrophages, but YAP can not only inhibit the recruitment of macrophages, but also increase the expression of Csf1. It can also cause tumor initiation cells to be immunized. This clearance depends on the adaptive immune system. And through the p53 dependent pathway, the clearance of tumor initiation cells resulted in the failure of YAP to induce tumorigenesis. In precancerous lesions of human hepatocellular carcinoma and other proto-oncogene-activated AKT and EGFR induced tumor initiation cells. The activation of Yap is accompanied by the recruitment of macrophages. A flexible mouse model of liver cancer associated with human disease, It can be widely used to study the signal transduction pathway of liver cancer and the interaction between hepatoma cells and microenvironment. More importantly, this study reveals a new mechanism of non-cellular self-action of Hippo pathway in the process of liver tumor formation. That is, by regulating the tumor microenvironment to promote tumorigenesis, and we have found that tumor initiation cells can create immunosuppressive microenvironments by actively recruiting macrophages as early as a single cell stage. A new method of immune prevention, That is, by inhibiting YAP activity or tumor initiation cell associated macrophages, the tumor initiation cells can be eliminated to prevent tumorigenesis.
【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R735.7

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