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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 黃念祖 | zh_TW |
| dc.contributor.advisor | Nien-Tsu Huang | en |
| dc.contributor.author | 熊彥程 | zh_TW |
| dc.contributor.author | Yen-Cheng Hsiung | en |
| dc.date.accessioned | 2024-08-08T16:18:16Z | - |
| dc.date.available | 2024-08-09 | - |
| dc.date.copyright | 2024-08-08 | - |
| dc.date.issued | 2024 | - |
| dc.date.submitted | 2024-07-30 | - |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/93804 | - |
| dc.description.abstract | 心血管疾病是全球最大死因之一，研究心血管的各式反應變得十分重要。本研究中我們開發出一個微流道系統用以建立得以模擬人體內的氧氣及剪應力環境。我們利用四個平行但寬度不同的微流道製造出來的剪應力分別為2.5, 5, 10, 20dyne/cm2，同時我們也一個建立了一個介於2.8 %-12 % 的氧氣濃度梯度。我們在這些微流道中培養人類臍靜脈血管內皮細胞，使這些內皮細胞同時受到剪應力以及氧氣梯度的刺激模擬在人體內的條件。我們使用相位差顯微鏡以及螢光染劑來分析細胞的型態、活性氧化物種以及一氧化氮的胞內表現。我們初步發現細胞沿著流體方向排列的速度較不存在缺氧條件的細胞快，我們更發現間歇性缺氧卻沒有如同缺氧促進細胞轉向的效果。而活性氧化物的分泌量則在一小時的實驗中受到缺氧而有所抑制，最後在六小時的實驗中，一氧化氮表現量並沒有發現明顯變化。這項研究有助於了解內皮細胞在各式心血管疾病、腫瘤以及人體運動時的反應，開發的微流道平台更有助於後人運用研究更複雜的生理條件。 | zh_TW |
| dc.description.abstract | Cardiovascular diseases are the leading cause of death worldwide, making the study of various cardiovascular responses critically important. In this study, we developed a microfluidic system designed to simulate the oxygen and shear stress environments within the human body. Using four parallel microchannels of different widths, we generated shear stresses of 2.5, 5, 10, and 20 dyne/cm2. Additionally, we established an oxygen concentration gradient ranging from 2.8 % to 12 %. Human umbilical vein endothelial cells were cultured within these channels, exposing them to both shear stress and the oxygen gradient to simulate in vivo conditions. We employed phase-contrast microscopy and fluorescent dyes to analyze cell morphology, reactive oxygen species, and nitric oxide production. Preliminary findings revealed that in hypoxic conditions, cell alignment along the flow direction was faster compared to normoxic conditions. We also found that cyclic hypoxia did not enhance cell reorientation. ROS production was suppressed under hypoxia within one hour, while NO production showed no significant changes over a six-hour period. This study enhances our understanding of endothelial cell responses in various cardiovascular diseases, tumors, and during physical exercise. Furthermore, this developed platform can also be a powerful tool for researchers studying complicated physiological conditions. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2024-08-08T16:18:16Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2024-08-08T16:18:16Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | Verification Letter from the Oral Examination Committee i
Acknowledgements ii 摘要iii Abstract iv Contents v List of Figures ix List of Tables xv Denotation xvi Chapter 1 Introduction 1 1.1 ROS in endothelial cells . . . . . . . . . . . . . . . . . . . . . . . . 1 1.1.1 Source of ROS . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 1.1.2 ROS signaling in endothelial cells . . . . . . . . . . . . . . . . . . 2 1.1.3 ROS related to vascular diseases . . . . . . . . . . . . . . . . . . . 4 1.2 Hypoxia effect on endothelial cells . . . . . . . . . . . . . . . . . . . 4 1.2.1 HIF-1α and HIF-2α . . . . . . . . . . . . . . . . . . . . . . . . . . 5 1.2.2 Hypoxia effect on ROS and NO production . . . . . . . . . . . . . 6 1.3 Shear stress effect on endothelial cells . . . . . . . . . . . . . . . . . 10 1.3.1 Shear stress on the regulation of ROS and NO . . . . . . . . . . . . 10 1.3.2 Shear stress on morphology . . . . . . . . . . . . . . . . . . . . . . 12 1.4 The combined effect of hypoxia and shear stress . . . . . . . . . . . 14 1.5 Literature review . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 1.5.1 Traditional methods to generate shear stress . . . . . . . . . . . . . 15 1.5.2 Methods to study shear stress in microfluidics . . . . . . . . . . . . 16 1.6 Generating hypoxia in microfluidics . . . . . . . . . . . . . . . . . . 18 1.6.1 Using gases as oxygen scavengers . . . . . . . . . . . . . . . . . . 18 1.6.2 Using chemical reactions as oxygen scavengers . . . . . . . . . . . 21 1.6.3 Using cells as oxygen scavengers . . . . . . . . . . . . . . . . . . . 21 1.6.4 Using CoCl2 as hypoxia inducers . . . . . . . . . . . . . . . . . . . 22 1.7 Motivation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24 Chapter 2 Theory 25 2.1 Hypoxic condition in microfluidic device . . . . . . . . . . . . . . . 25 Chapter 3 Materials and methods 31 3.1 The microfluidic chip . . . . . . . . . . . . . . . . . . . . . . . . . . 31 3.1.1 Microfluidic chip design . . . . . . . . . . . . . . . . . . . . . . . 31 3.1.2 Microfluidic chip fabrication process . . . . . . . . . . . . . . . . . 33 3.2 Cell culture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 3.2.1 Cell passage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 3.2.2 Cell freezing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 3.2.3 Cell seeding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 3.3 Hypoxic condition generation . . . . . . . . . . . . . . . . . . . . . 36 3.4 Oxygen level measurement in microfluidic devices . . . . . . . . . . 36 3.4.1 Fluorescence lifetime imaging microscopy . . . . . . . . . . . . . . 36 3.4.2 Oxygen sensor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37 3.5 System setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39 3.6 Cell imaging . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39 3.7 ROS measurement in cells . . . . . . . . . . . . . . . . . . . . . . . 40 3.8 NO measurement in cells . . . . . . . . . . . . . . . . . . . . . . . . 40 3.9 Image processing . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40 3.9.1 Bright-field image processing . . . . . . . . . . . . . . . . . . . . . 40 3.9.2 Fluorescence image processing . . . . . . . . . . . . . . . . . . . . 44 Chapter 4 Results and Discussion 45 4.1 Simulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45 4.1.1 Basic fluid characteristics . . . . . . . . . . . . . . . . . . . . . . . 45 4.1.2 Oxygen microenvironment establishment . . . . . . . . . . . . . . 46 4.1.2.1 Constant oxygen gradient . . . . . . . . . . . . . . . . 47 4.1.2.2 Cyclic oxygen gradient . . . . . . . . . . . . . . . . . 49 4.2 Cell morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52 4.2.1 Orientation changing with very short shear stress stimulation . . . . 53 4.2.2 Cell morphology after stimulation . . . . . . . . . . . . . . . . . . 54 4.3 ROS production . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60 4.3.1 ROS staining testing . . . . . . . . . . . . . . . . . . . . . . . . . 60 4.3.2 ROS production after continuous hypoxia and shear stress . . . . . . 63 4.4 NO production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64 4.4.1 NO staining testing . . . . . . . . . . . . . . . . . . . . . . . . . . 64 4.4.2 NO production after continuous hypoxia and shear stress . . . . . . 66 4.5 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70 4.5.1 Chip design revisit . . . . . . . . . . . . . . . . . . . . . . . . . . 70 4.5.2 Hypoxic condition generation . . . . . . . . . . . . . . . . . . . . . 71 4.5.3 Morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72 4.5.4 ROS production . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73 4.5.5 NO production . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73 Chapter 5 Conclusions 74 Chapter 6 Future Work 76 References 79 Appendix A — Cell Morphology 86 A.1 Scatter plots . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86 | - |
| dc.language.iso | en | - |
| dc.subject | 活性氧化物種 | zh_TW |
| dc.subject | 一氧化氮 | zh_TW |
| dc.subject | 人臍靜脈內皮細胞 | zh_TW |
| dc.subject | 剪應力 | zh_TW |
| dc.subject | 缺氧 | zh_TW |
| dc.subject | 微流體 | zh_TW |
| dc.subject | reactive oxygen species | en |
| dc.subject | hypoxia | en |
| dc.subject | shear stress | en |
| dc.subject | nitric oxide | en |
| dc.subject | human umbilical vein endothelial cell | en |
| dc.subject | microfluidic | en |
| dc.title | 開發微流道系統晶片分析剪應力與缺氧刺激對於內皮細胞產生活性氧化物與一氧化氮之影響 | zh_TW |
| dc.title | Development of a Microfluidic System to Study the Reactive Oxygen Species and Nitric Oxide Dynamics of Endothelial Cells under Simultaneous Stimulation of Shear Stress and Hypoxia | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 112-2 | - |
| dc.description.degree | 碩士 | - |
| dc.contributor.oralexamcommittee | 郭柏齡;楊東霖;董奕鍾 | zh_TW |
| dc.contributor.oralexamcommittee | Po-Ling kuo;Tony Yang;Yi-Chung Tung | en |
| dc.subject.keyword | 微流體,人臍靜脈內皮細胞,活性氧化物種,一氧化氮,剪應力,缺氧, | zh_TW |
| dc.subject.keyword | microfluidic,human umbilical vein endothelial cell,reactive oxygen species,nitric oxide,shear stress,hypoxia, | en |
| dc.relation.page | 94 | - |
| dc.identifier.doi | 10.6342/NTU202402333 | - |
| dc.rights.note | 同意授權(全球公開) | - |
| dc.date.accepted | 2024-08-01 | - |
| dc.contributor.author-college | 電機資訊學院 | - |
| dc.contributor.author-dept | 生醫電子與資訊學研究所 | - |
| 顯示於系所單位： | 生醫電子與資訊學研究所 | |
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| ntu-112-2.pdf | 30.45 MB | Adobe PDF | 檢視/開啟 |
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