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研究生： 翁上雯
Wong, Shang-Wen.
論文名稱： 以系統生物學方法和大數據資料庫探勘分析正常、發育和再生肝臟之間的基因和表觀遺傳差異細胞網路機制
Investigation of Genetic-and-Epigenetic Networks for Differential Cellular Mechanisms between Normal, Developing and Regenerating Livers via Systems Biology Method and Big Data Mining
指導教授： 陳博現
Chen, Bor-Sen
口試委員: 詹鴻霖
Chan, Hong-Lin
王慧菁
Wang, Hui-Ching
蘇士哲
Sue, Shih-Che
王禹超
Wang, Yu-Chao
學位類別： 碩士
Master
系所名稱： 電機資訊學院 - 電機工程學系
Department of Electrical Engineering
論文出版年： 2017
畢業學年度： 105
語文別： 英文
論文頁數： 86
中文關鍵詞： 肝臟 、再生 、發育 、肝臟發育 、造血 、系統生物學 、大數據探勘 、表觀遺傳 、網路生物標記 、多分子藥物
外文關鍵詞： liver, regeneration, development, hepatogenesis, hematopoiesis, systems biology, big data analysis, epigenetics, network biomarkers, multi-molecule drug
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    • 肝臟是具有較高再生能力的器官，在出生前後對於維持體內平衡和健康發揮重要作用。出生前肝臟發育（HG）和部分肝臟切除（PHx）後的肝臟再生（LR）的遺基因調控機制已經有深入的研究；然而，關於肝臟在肝臟發育和肝臟再生期間為了適應快速的變化所進行地表觀遺傳修飾的機制所知甚少。首先，我們使用系統生物學和大數據探勘，透過發育、正常和肝臟部分切除後再生的肝臟的轉錄組和甲基化來建構全基因組基因及表觀遺傳網路（GWGEN）。接著，我們對3個肝臟條件的GWGEN使用主成分網絡投影（PNP）來提取核心GWGEN。最後，對核心GWGEN進行分析比較，以探究肝臟發育和肝臟再生的重要信號傳導途徑和表觀遺傳修飾機制。我們觀察到細胞外信號被傳導到轉錄因子（TFs），使其目標基因受到調節，從而誘導負責肝臟發育的細胞機制：AR的磷酸化將造血前驅細胞招募到發育中的肝臟，並且ZBTB8A的磷酸化、DNA甲基化活化的MAF和GATA1能促進造血細胞系分化。此外，我們觀察到通過TF ZEB1磷酸化的表觀遺傳修飾對內質網壓力的監控機制。在正常肝臟中，表觀遺傳修飾引發多倍體機制，包括肝細胞中IGF1R、IGF1和ATXN3的磷酸化以及EGR1的DNA甲基化。已經觀察到多倍體機制有利於肝臟的再生。在肝臟再生過程中，表觀遺傳調控對於部分肝臟切除所引起的微環境的反應具有重要作用。在再生的肝臟中，TF ETS1的乙酰化可以觸發SERPINE1在部分肝臟切除後建立最早期的微環境。透過FOXO3的乙酰化和磷酸化，來使損傷的肝細胞自我凋亡；NFATC1和NFATC2也可以產生細胞激素以招募免疫相關細胞，並移除細胞碎片。MDM2的磷酸化和SUMO化正調控AP-1複合物（FOSL2和JUN）的表現，並促進肝細胞增殖以恢復肝臟質量和功能；通過基因調控和DNA甲基化活化MIR21可以維持肝臟再生的瞬態環境。最後，我們提出了一種藥物設計流程，來設計一種多分子藥物來標靶到我們探究肝臟發育和肝臟再生的基因及表觀遺傳機制後提出的特定生物標誌物上，以促進肝臟再生。

    The liver is an organ with high regenerative capacity and plays an important role in the maintenance of homeostasis and health before and after birth. The genetic mechanisms of prenatal hepatogenesis (HG) and liver regeneration (LR) after partial hepatectomy (PHx) had been thoroughly investigated; however, less is known about the mechanisms of epigenetic modifications which liver has performed to adapt to the rapidly changing circumstance during hepatogenesis and liver regeneration. First, we used systems biology and big data mining to construct whole genome-wide genetic-and-epigenetic networks (GWGENs) via the transcriptomes and methylomes of human developing, normal and regenerating liver after PHx. Then we extracted the core GWGENs by applying principal network projection (PNP) respectively on GWGENs of 3 hepatic conditions. Finally, the core GWGENs are analyzed and compared to investigate the significant signal transduction pathways and epigenetic modification mechanisms of hepatogenesis and liver regeneration. We observed extracellular signals were transduced to transcription factors (TFs), leading to the regulation of their target genes, in turn inducing cellular mechanisms that are responsible for hepatogenesis: phosphorylation of AR recruits hematopoietic progenitor stem cell into developing liver, and phosphorylation of ZBTB8A and DNA methylation activation of MAF and GATA1 promote hematopoietic cell line differentiation. Moreover, we observed the surveillance mechanism by the epigenetic modification of phosphorylation on TF ZEB1 to react to ER stress. In the human normal liver, polyploidy mechanism is initiated by epigenetic modification including the phosphorylation of IGF1R, IGF1, ATXN3 and DNA methylation of EGR1 in hepatocyte. The polyploidy mechanism is observed to benefit liver regeneration. During early stage of human liver regeneration, epigenetic regulations play an important role to respond to the microenvironment caused by PHx. In the regenerating liver, acetylation of TF ETS1 could trigger SERPINE1 to establish microenvironment at the earliest phase after PHx. Injured hepatocytes are removed by apoptosis due to the acetylation and phosphorylation of FOXO3; also NFATC1 and NFATC2 could produce cytokines to recruit immune-related cells and remove cell debris. Phosphorylation and sumoylation of MDM2 upregulating AP-1 complex (FOSL2, JUN) could promote hepatocytes proliferation to regain liver mass and function; activation of MIR21 by gene regulations and DNA methylation could sustain the transient environment for liver regeneration. Finally, we proposed a drug design procedure to design a multi-molecule drug targeting on specific biomarkers we investigated from the genetic-and-epigenetic mechanisms of hepatogenesis and liver regeneration to promote liver regeneration.

    中文摘要 I Abstract II Contents V List of Tables VII List of Figures VII List of Abbreviations VIII 1. Introduction 1 2. Materials and Methods 5 2.1 Overview of the construction processes of real GWGENs of 3 hepatic conditions 5 2.2 Big data mining and data preprocessing 6 2.3 Dynamic models of the candidate GWGEN for developing, normal and regenerating livers 8 2.4 System identification method of the dynamic models of candidate GWGEN 11 2.5 System order detection scheme of the dynamic models of GWGEN 17 2.6 Core network identification of the real GWGENs by applying the PNP method 19 3. Results and Discussion 23 3.1 Construction of the real GWGENs of the developing, normal and regenerating livers (Figures S1, S2 and S3). 23 3.2 Construction of core GWGENs of the developing, normal and regenerating livers (Figures S4, S5, and S6). 23 3.3 Core genetic-and-epigenetic mechanism during hepatogenesis (HG) (Figures 2 and 3) 25 3.3 (a) Cytoskeleton remodeling recruits hematopoietic progenitor cells (Figure 3(a)) 25 3.3 (b) Hematopoiesis cell line differentiation via the perception of ion concentration changes (Figure 3(b)) 26 3.3 (c) Tight surveillance control of hepatogenesis (Figure 3(c)) 28 3.3 (d) Polyploidy of hepatocyte in normal liver (Figure 3(d)) 28 3.4 Core genetic-and-epigenetic mechanism during liver regeneration (LR) (Figures 4 and 5) 31 3.4 (a) Establishing microenvironment for liver regeneration by acetylation activation of ETS1: phosphorylation, glycosylation, and deubiquitination activation of bipotential progenitor cell (FOXL1) to differentiate into hepatocytes and cholangiocytes (Figure 5(a)) 31 3.4 (b) T cell lineage development and differentiation in liver by GATA3; Apoptosis of injured hepatocytes by acetylation and phosphorylation of FOXO3; production of cytokines by NFATC1 and NFATC2 to recruit immune-related cells and remove cell debris (Figure 5(b)) 33 3.4 (c) Phosphorylation and sumoylation of AP-1 complex (FOSL2, JUN) promotes hepatocytes proliferation; activation of MIR21 by GRs and DNA methylation sustain the environment for liver regeneration (Figure 5(c)) 36 3.5 The overview of genetic-and-epigenetic mechanisms of developing liver (hepatogenesis), normal liver, and regenerating liver (liver regeneration) 40 3.6 Specific biomarkers for liver regeneration and multi-molecule drug design 44 4. Conclusions 46 Tables 48 Figures 66 References 83

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