臺灣博碩士論文加值系統

本論文探討血液流體動力學與內皮細胞增生的關係，實驗藉由自製的脈動式流體產生系統，結合聚二甲基矽氧烷(PDMS)等軟性材料，製作脈動平行板流體腔(flow chamber)，產生剪應力範圍3~25 、流量範圍約0.1~1.6ml/sec、雷諾數約20~210。另外藉由PDMS A劑與B劑混合的比例不同製作各種硬度的PDMS(E=1.76~0.072 MPa)，並在其上培養內皮細胞，探討內皮細胞在不同硬度的軟性材質上時的增生條件，比較流體刺激前與刺激後的差異。

實驗所需的內皮細胞來自胎兒臍帶的靜脈(HUVEC)，由台大醫院產房取得，每次實驗的細胞皆用多條臍帶混合，以減少生物體上的誤差，實驗結果顯示未受流體刺激的內皮細胞，在硬度不同的聚二甲基矽氧烷(PDMS)上增生量均不相同，硬度越大增生量越高 (E=1.76 MPa，增生量15.4% ; E=0.072MPa，增生量8.8%)，但受到流體刺激後，硬度高與硬度低均有顯著的增生效果 (E=1.76 MPa，增生量44.8% ; E=0.072MPa，增生量44.5%)， 另外提高脈動流體剪應力的時間梯度時，增生量亦增加(E=1.76 MPa，增生量56.8%)，而流體Womersley number的大小似乎也對內皮細胞增生扮演重要的角色。

This study discussed the effect of hemodynamics on the proliferation of endothelial cells. We designed the pulsatile parallel plate flow chamber which combined PDMS polymer on the chamber bottom. Shear stress range were 3-25 dyne/cm2、flow rate were 0.1-1.6 ml/sec、Re numbers were 20-210. By using different mixture ratio of sylgard 184(A) and curing agent (B), the PDMS polymer can made with different stiffness, the resulted Young’s moduli were about 0.072-1.76 MPa. The endothelial cells were cultured on the PDMS polymer to study the cell proliferation under no flow condition and excited flow with different membrane stiffness.

Human umbilical vein endothelial cells (HUVEC) were obtained from NTUH. to To reduce individual bias the endothelial cells were collected and mixed from more than one umbilical cords in all experiments. Results showed that the endothelial cells proliferation under no flow conditions increases on high stiffness PDMS (E=1.76 MPa, 15.4%; E=0.072MPa, 8.8%). Under excited flow, the cell proliferation for both the high surface-stiffness and low surface-stiffness increased remarkably. (E=1.76 MPa, 44.8%; E=0.072MPa, 44.5%). Besides, the enhanced temporal gradient in shear stress also increases the cell proliferation. It is therefore concluded that the Womersley number seemed to play an important rule on the endothelial cells proliferation.

誌謝..............................................................................................................................Ⅰ
中文摘要......................................................................................................................Ⅱ
英文摘要......................................................................................................................Ⅲ
內文目錄......................................................................................................................Ⅳ
圖目錄..........................................................................................................................Ⅵ
表目錄..........................................................................................................................Ⅸ

第一章 序論................................................................................................................1
1-1 前言.........................................................................................................................1
1-2 研究動機與目的.....................................................................................................2
1-3 文獻回顧.................................................................................................................3
1-3-1 平行板流體腔之相關設計........................................................................3
1-3-2 內皮細胞增生與剪應力之相關研究.......................................................4
1-4 論文架構.................................................................................................................6
第二章 研究方法與設備..........................................................................................10
2-1 脈動式流體系統................................................................................................10
2-1-1 平行板流體腔之製作方法與設計參數.................................................10
2-1-2 流量波之計算.........................................................................................10
2-1-3 剪應力大小之計算.................................................................................11
2-1-4 生理範圍之流體力學相關參數之比對.................................................14
2-1-5 PDMS制模方法與厚度量測..................................................................15
2-1-6 PDMS楊氏模數之量測方法..................................................................................16
2-2 PVDF感測器原理..............................................................................................18
2-2-1 壓電材料之特性.....................................................................................18
2-2-2 PVDF壓電感測器製作方法 .................................................................................19
2-2-3 電荷放大器與電壓放大器.....................................................................19
2-2-4 壓電係數之校正.....................................................................................25
2-2-5 低通濾波器、與60Hz帶阻濾波器.......................................................25
2-2-6 PVDF量測流體之正向應力及剪應力與理論值比對..........................26
2-2-7 LabView資料擷取..................................................................................26
2-3 內皮細胞培養方法與體外溫控螢光顯微系統............................................................29
2-3-1 倒立式螢光顯微鏡與細胞體外培養系統之架構..................................29
2-3-2 內皮細胞培養與生物實驗之流程.........................................................29
2-3-3 內皮細胞增生與定量方法.....................................................................30
第三章 結果與討論..................................................................................................40
3-1 PDMS 楊氏模數之量測結果............................................................................40
3-2 內皮細胞附著狀況之探討.................................................................................42
3-2-1 Vitrogen對細胞附著之影響...................................................................42
3-2-2 流體作用後對細胞貼附之影響..............................................................43
3-3 細胞增生之探討.................................................................................................44
3-3-1 BrdU添加時間與細胞增生量之探討....................................................44
3-3-2 無流體作用下覆蓋PDMS之硬度對細胞增生的影響.........................44
3-3-3 流體作用下覆蓋PDMS的厚度對細胞增生的影響.............................45
3-4-4 有流體作用下覆蓋PDMS之硬度對細胞增生的影響.........................47
第四章 結論及未來研究建議..................................................................................57
4-1 結論.........................................................................................................57
4-2 未來研究建議.........................................................................................59
參考文獻......................................................................................................................60

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