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標題: | 開發用於研究提高間質液壓對於細胞環境彈性影響之三維細胞培養平台 Development of a Three-Dimensional Cell Culture Platform for Studying Interstitum Elasticity Induced by Increased Interstitial Fluid Pressure |
作者: | Ching-Wei Lan 藍慶瑋 |
指導教授: | 郭柏齡 |
關鍵字: | 三維細胞培養,剪切波彈性影像,間質液壓,胞外間質硬度,膠原蛋白, 3D cell culture,shear wave elasticity imaging,interstitial fluid pressure,ECM stiffness,collagen, |
出版年 : | 2018 |
學位: | 碩士 |
摘要: | 細胞與其生長基質建構該微環境的硬度,在細胞生長過程可透過感受外在硬度而朝向不同的發展行為與表現,例如影響幹細胞分化和癌症轉移等。先前研究中也證實硬度在二維環境的重要性,然而在三維環境中細胞行為與其在二維又有所差異,除此之外三維的環境也較貼近細胞於生物體內生長的情形,故在此篇研究中我們將細胞培養於一種主要成分由一型膠原蛋白所組成的細胞培養支架達到三維培養的目標,並設計一種由聚乙烯對酞酸脂(超鋼龍,PETP)材料所組合的流道裝置,透過將含有細胞培養的膠體置於該裝置,利用流道的特殊設計對於細胞施加流體壓力以及產生間質流,並使用20MHz的超音波探頭聚焦送入聲場輻射力,再透過40MHz的探頭同步監測剪切波傳遞,進行超音波剪切彈性影像測量,該系統有著非侵入式、非接觸式測量以及即時監控的優點。我們預計挑選目標細胞並模擬其所在組織環境的流體壓力,探討水壓造成細胞對於環境硬度的影響之關聯性:如纖維母細胞與傷口癒合或是肝細胞受壓後代謝程度的變化等,流體壓力與組織環境硬度以及細胞行為總是有相當程度的關係但卻很難去量化或是確定從屬關係,透過該研究,期望可以進一步的了解三者之間的關係。我們發現到3T3纖維母細胞在液壓環境培養兩天後,會使得細胞內的α-平滑肌肌動蛋白(α-smooth muscle actin, α-SMA)表現量上升,進而造成細胞外微環境的收縮,另外也透過轉化生長因子-β激酶(Transforming growth factor beta,TGF-β)抑制劑發現該現象能夠被削弱,故推測間質液壓是透過轉化生長因子-β受體途徑(TGF-β pathway)來提升α-SMA之表現量,並給予細胞更強的收縮力來改變胞外基質硬度。 Microenvironment stiffness was formed by cell and ECM, and cells could sense the local stiffness and present different behavior and performance during the development, such as stem cell differentiation or cancer metastasis and so on. Previous studies have suggested that stiffness plays an important role in 2D environment; however, we have seen differences in cell behavior between 2D and 3D culture environment. In general, cells cultured in vitro 3D models are more similar to the real environment in organism. In this study, we used the collagen type I for cell culturing to construct the 3D culture model, and developed a 3D culture platform which made of polyethylene terephthalate polyester (PETF). We placed the cell-populated collagen gel into the culture chamber and applied fluid pressure and pressure drop to generate the interstitial fluid flow by a novel design of the platform. Next, we generated the ultrasound radiation force by 20 MHz push transducer and monitor the shear wave propagation by 40 MHz image transducer to process the shear wave elasticity imaging measurement which has an advantage in non-invasive, non-contact and real-time measurement. We would select the target cell and imitate its physiological environment to investigate relationship between fluid pressure and ECM stiffness, such as fibroblasts in wound healing or the variation of metabolic efficiency in hepatocytes under pressure conditions. There appears to be a causality among the fluid pressure, interstitium elasticity and cell behavior and we tried to get more information about them thought our experiments. We found the increased expression of α-SMA in 3T3 fibroblast under the fluid pressure condition for 2 days culture, and reduced this trend by blocking the TGF-β receptor. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/69893 |
DOI: | 10.6342/NTU201800562 |
全文授權: | 有償授權 |
顯示於系所單位: | 生醫電子與資訊學研究所 |
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