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系統識別號 U0026-2406201517565800
論文名稱(中文) 細胞骨架纖維硬度的空間分布決定細胞移動的模式
論文名稱(英文) Spatial Distribution of Filament Elasticity Determines the Migratory Behavior of a Cell
校院名稱 成功大學
系所名稱(中) 基礎醫學研究所
系所名稱(英) Institute of Basic Medical Sciences
學年度 103
學期 2
出版年 104
研究生(中文) 韓逸成
研究生(英文) Hans I-Chen Harn
學號 S58981230
學位類別 博士
語文別 英文
論文頁數 110頁
口試委員 指導教授-湯銘哲
口試委員-蔣輯武
口試委員-吳佳慶
口試委員-鄭兆珉
口試委員-林耿慧
口試委員-王仰高
中文關鍵字 原子力顯微鏡  細胞骨架  機械力傳導  細胞遷移  蟹足腫  力學 
英文關鍵字 AFM  actin cytoskeleton  mechanotransduction  migration  keloid  force 
學科別分類
中文摘要 一個細胞的物理性質是由許多細胞內和外的因子所緊密的調控。任何伴隨著型態改變的細胞反應都需要精密的調控以平衡結構上的重整所產生的力量,而這些動態調整皆由細胞骨架所掌控。Actin纖維是產生細胞內部力量最基本的基石,同時它也透過focal adhesion complex傳輸外來機械刺激進入細胞內。在細胞遷移的過程當中,細胞不只經歷分子層次上的變化也有即時的機械性調整。另外,在不同的遷移模式當中,力量和纖維彈性如何在空間上被分布與調控仍尚被未了解。這些問題需要高階測量儀器的進展才能夠探討生物樣本從組織到細胞,再從細胞進入次細胞層級的物理性質。在緒論當中,我回顧了當今最新測量生物組織或細胞樣本的技術,不同器官和細胞之機械訊息傳導,不同種細胞骨架在細胞機械訊息傳遞中所扮演的角色,以及細胞遷移中細胞骨架的動態。在第三章中,我用原子力顯微鏡建立了一個測量方法以顯示1)非遷移中細胞只會產生一種纖維彈性,2)細胞在方向性遷移過程中會有空間分布性的產生兩種纖維彈性,3)病態細胞在空間分布上隨機產生兩種不同纖維彈性會遷移但不維持其方向性。在第四章中,我把這個模式延伸去探討蟹足腫纖維細胞的遷移本性,並顯示纖維彈性結合力量的生成皆須透過focal adhesion kinase所扮演的關鍵角色。這個論文給予actin纖維彈性嶄新的觀點,並展現它們在不同細胞遷移的行為模式當中如何做出物理性質的變化。
英文摘要 The mechanical properties of a cell is tightly regulated by multiple facets of intracellular processes and external factors. Any cellular response leading to morphological changes is highly tuned to balance the force generated from structural organization, provided by the cytoskeleton. Actin filaments serve as the backbone of intracellular force, and transduce external mechanical signal via focal adhesion complex into the cell. During migration, cells not only undergo molecular changes but also rapid mechanical modulation. Furthermore, how are the force and filament elasticity spatially regulated in different migratory behavior still remain unclear. These questions require advances in measuring devices to explore the biomechanical properties of biological samples from tissue to cellular and sub-cellular level. In the introduction, I reviewed the cutting edge technologies used today to measure biological samples on tissue and cellular levels, the mechanotransduction of different organ systems and the cell, the role of different cytoskeletons in the mechanobiology of the cell, and the cytoskeletal dynamics of cell migration. In chapter 3, I established a measuring methodology using atomic force microscopy to show 1) non-migratory cells only generated one type of filament elasticity, 2) cells generating spatially distributed two types of filament elasticity showed directional migration, and 3) pathologic cells that autonomously generated two types of filament elasticity without spatial distribution were actively migrating non-directionally. In chapter 4, I extended this model and explored the migrating nature of keloid fibroblasts, in which the coupling of filament elasticity and force generation through focal adhesion kinase play a vital role. The dissertation provides a whole new perspective to the elasticity of actin filaments, and how they are differentiated in different modes of migratory behavior.
論文目次 Abstract ----------- 3
Abstract in Chinese --------- 4
Acknowledgement ---------- 5
Contents ---------- 7
Figure of Contents--------- 9
Abbreviations --------- 11
Chapter 1: Introduction --------- 12
1.1 Measuring devices for biological samples -----13
1.2 Mechanotransduction of the cell ------ 18
1.3 Mechanotransduction in different organ systems --- 22
1.4 Cytoskeletons and cell elasticity ------ 30
1.5 Cell polarity and cell migration ----- 38
Chapter 2: Materials and Methods ------- 42
2.1 Cell culture -------- 42
2.2 Primary culture of keloid fibroblasts ----- 42
2.3 Atomic force microscopy------ 42
2.4 Living cell imaging ------- 43
2.5 Elasticity measurements ------- 43
2.6 Preparation of polyacrylamide gel ------ 44
2.7 Actin-RFP transfection ------ 44
2.8 Confocal images and three-dimensional reconstruction --- 44
2.9 Fibroblast wound heading assay ------ 45
2.10 Inhibitor treatment -------- 45
2.11 Microfabricated-post array detector ------ 45
2.12 Calculating force generation on mPAD ----- 46
2.13 Statistics --------- 47
Chapter 3: Spatial distribution of filament elasticity determines the migratory behavior of a cell ---------- 48
3.1 Abstract -------- 48
3.2 Introduction --------- 49
3.3 Results --------- 53
3.4 Discussion -------- 60
Chapter 4: Mechanical coupling of cytoskeletal elasticity and force generation is crucial for understanding the migrating nature of keloid fibroblasts - 62
4.1 Abstract -------- 62
4.2 Introduction --------- 63
4.3 Results --------- 65
4.4 Discussion --------- 68
Chapter 5: Summary and prospect ------- 70
References ---------- 72
Figures ---------- 94
Curriculum Vitae --------110
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