本文選題：Hippo通路　切入點：腫瘤起始細胞　出處：《浙江大學》2016年博士論文　論文類型：學位論文

【摘要】：免疫系統(tǒng)和炎癥在腫瘤發(fā)生的每個階段都有著很重要的作用。在已形成的腫瘤里,浸潤的炎細胞通過與腫瘤細胞相互作用,改變腫瘤微環(huán)境從而影響腫瘤發(fā)展。腫瘤里的各種炎細胞及其與腫瘤細胞的相互作用是設(shè)計有效免疫治療的理論基礎(chǔ)。肝臟腫瘤在中國和世界范圍內(nèi)對健康都是一個巨大的威脅。然而由于肝癌遺傳背景的復雜性至今還缺少準確模擬肝臟腫瘤的小鼠遺傳學模型。人肝臟腫瘤的發(fā)生往往伴隨著炎癥反應,然而在炎性微環(huán)境里產(chǎn)生的腫瘤起始細胞是否主動改造免疫微環(huán)境尚不清楚。Hippo信號通路是近年來發(fā)現(xiàn)的調(diào)控器官大小的關(guān)鍵信號途徑,它的異常與癌癥發(fā)生特別是肝癌密切相關(guān)。為了在體內(nèi)研究炎癥細胞對肝臟腫瘤起始細胞的影響,特別是由Hippo通路失調(diào)引起的肝臟腫瘤,我建立了一個流體動力學注射和轉(zhuǎn)座子結(jié)合的小鼠肝癌模型。利用這種模型,在成體小鼠肝實質(zhì)細胞內(nèi)穩(wěn)定過表達激活型的Hippo通路效應分子YAP,在4個月內(nèi)就可以強烈誘導小鼠肝臟腫瘤生成。我們發(fā)現(xiàn)YAP的抑制蛋白VGLL4可以有效抑制YAP引起的腫瘤,而YAP的下游分子,miR-130a可以通過靶向VGLL4增強YAP誘導的肝臟腫癌形成。這一研究揭示了 Hippo信號通路調(diào)控腫瘤發(fā)生的正反饋回路。我意外地發(fā)現(xiàn)在YAP誘導腫瘤生成的早期,甚至是單個細胞的腫瘤起始細胞階段就能很強地招募巨噬細胞。這些巨噬細胞具有腫瘤相關(guān)巨噬細胞的性質(zhì)。敲除Hippo通路的激酶Mst1/2或Lats1/2從而激活內(nèi)源性YAP也可以招募巨噬細胞。在腫瘤起始細胞里,YAP可以上調(diào)趨化因子Ccl2和Csf1的表達來招募巨噬細胞。YAP通過TEAD對Ccl2實現(xiàn)直接調(diào)控。通過敲減Ccl2和Csf1不僅有效阻止巨噬細胞的招募,還能導致腫瘤起始細胞被免疫清除。這種清除依賴于適應性免疫系統(tǒng),并且通過p53依賴的途徑。腫瘤起始細胞的清除最終導致了 YAP不能誘導腫瘤生成。在人肝癌的癌前病變中和其他原癌基因活化型AKT和EGFR誘導形成的腫瘤起始細胞里,YAP的活化也伴隨著巨噬細胞的招募。以上結(jié)果首先建立了一種快速、靈活,與人類疾病相關(guān)的肝癌小鼠模型,可廣泛用于研究肝癌信號轉(zhuǎn)導途徑以及肝癌細胞與微環(huán)境的相互作用。更重要的是本研究揭示了 Hippo通路在肝臟腫瘤形成過程中的一種新的非細胞自身作用的機制,即可以通過調(diào)控腫瘤微環(huán)境從而促進腫瘤發(fā)生。并且我們發(fā)現(xiàn)腫瘤起始細胞早在單個細胞階段就可以通過主動招募巨噬細胞來塑造免疫抑制性微環(huán)境。這提示了一種新的免疫預防的方法,即通過抑制YAP活性或腫瘤起始細胞相關(guān)巨噬細胞達到清除腫瘤起始細胞預防腫瘤發(fā)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.
【學位授予單位】：浙江大學
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R735.7

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