細胞沉積現象與多孔基質內細胞遷移行為之實驗與模擬研究; Experiment and Simulation Study of Cell Sedimentation and Cell Migration through Porous Constructs

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題名:	細胞沉積現象與多孔基質內細胞遷移行為之實驗與模擬研究;Experiment and Simulation Study of Cell Sedimentation and Cell Migration through Porous Constructs
作者:	鍾志昂;鄭宇哲;黃衍任
貢獻者:	機械工程學系
關鍵詞:	博登量測器;細胞遷移;數學模型;模擬;隨機漫步;化學趨向;Boyden chamber;cell migration;mathematic model;simulation;random walk;chemotaxis;機械工程類
日期:	2010-08-01
上傳時間:	2011-07-12 13:58:53 (UTC+8)
出版者:	行政院國家科學委員會
摘要:	細胞的運動對於生理組織的生長，傷口治癒 (wound healing)，發炎反應，病理癌細胞的散佈和轉移，以及對於組織工程體外培養細胞在支架的生長與分佈，都有重要的影響。細胞的運動不僅與細胞本身的特性有關，也會受外界環境的影響。為了瞭解細胞的運動方式與行為，學者設計不同的設備以量測細胞的運動，在各式量測細胞群體運動行為的方法中，博登量測器是相當便利也經常使用的裝置。博登量測器的基本構造是一個腔室，由一片多孔薄膜將之隔間為上下兩層，細胞懸浮液置於上腔室，細胞受重力作用沉積於薄膜表面，經過薄膜孔洞向下攀爬，下腔室可置放化學趨向因子，藉由擴散作用，化學趨向因子在多孔薄膜裡產生濃度梯度，吸引細胞產生化學趨向性移動。為了避免細胞增殖引起細胞數目判讀的誤差，博登量測實驗通常不超過10 小時。然而細胞沉積需要約 3 小時，而細胞沉積到薄膜表面，剛貼附在基質上時，也無法即刻產生適當的收縮力使細胞向前遷移，需經一段時間適應環境，細胞與基質的貼附力量才能逐漸增強到足以支撐細胞產生移動。然而在前人描述博登量測器實驗的模型中，並沒有考慮細胞沉積及細胞與基質貼附的影響，而是假設實驗一開始，所有細胞就已經完全沉積貼附在薄膜上，並以最大的移動速度向前遷移，因而對實驗結果的預測與判讀將產生誤差。本計畫擬對細胞在博登量測器的遷移行為進行詳實的理論與實驗探討，計畫內容分為三部分。第一部分是細胞沉積模型的建立，並以實際細胞的沈積實驗，測量細胞在沈積時的平均速度與濃度關係，以修正模型係數。第二部分是細胞與薄膜表面貼附適應的探討，觀測細胞貼附薄膜後開始運動的現象，量測細胞移動速度與細胞薄膜之間的貼附時間的關係曲線，以詳實評估細胞貼附基質後的移動行為。第三部分整合前述細胞沉積與細胞貼附適應模型，建立博登量測器的完整細胞遷移數學模型，撰寫程式進行模擬，並進行博登量測器實驗，比對理論與實驗結果。全部研究將在三年內完成，希冀經由理論與實驗的相互印證，提高對細胞遷移的知識與技術，建立細胞沉積與遷移的數學模型，用以量測計算細胞群體的遷移係數，幫助瞭解細胞於多孔支架的遷移行為，加速組織工程的進步。 Cell migration phenomena underlie many biological processes. Physiological examples include immune responses, angiogenesis, and cell movement into wounds for wound healing. On the pathological side, tumor cells moving into the circulatory system and causing metastasis is still a critical challenge for cancer treatment. On the curative side, tissue engineering is working to create tissue implants from cell-seeded polymers. To this end, understanding how cell motilities influence cell distribution and growth in engineered tissue implants is crucial to the development of functional tissue grafts. In order to study how cells migrate in response to their environmental cues, common techniques that can carry out in vitro cell locomotion tests cannot be ignored. Of the various test protocols, Boyden chamber assay has been applied over a wide range because it can provide a convenient means of determining cell diffusion coefficients as well as tactic motilities. Briefly speaking, a Boyden chamber is comprised of two wells separated by a porous membrane called the chamber filter. To perform chemotaxis tests, solutions of different chemoattractant concentrations are put in the upper and lower wells respectively to build up chemical gradients. Cell movements responding to chemical gradients are assessed by counting the number of cells that reach the membrane underside in some incubation time. From a practical point of view, the incubation period may last only hours, which is quite close to the time required for cells to settle upon the membrane. Therefore part of the cells would still remain in the suspension with other cells in the membrane. Furthermore, cells would not start moving as soon as they reach the membrane surface. Instead, cells need to adapt themselves to the substrates before commencing locomotion. As a result, mathematic modeling without considering the cell settling effect or attachment adaption is likely to induce errors in interpreting the experimental data. The proposed project aims to perform a comprehensive study on the cell migration using Boyden chambers. The project is divided into three major parts. The first part includes modeling and verification experiments on cell sedimentation. We are planning on building a mathematical description for cell settling through the suspension above the chamber membrane. Experiments will also be carried out by measuring cell settling speeds using time lapse videomicroscopy with image processing method. The second part aims to clarify the relationship between cell motility and attachment. By conducting experiments using time lapse videomicroscopy with image processing method, the correlation of cell moving speed with cell attachment time will be determined. The third part combines the previous two portions in an attempt to develop a comprehensive mathematical model as to interpret the entire process of cell movement in a Boyden chamber assay. Results from simulation will be compared with the Boyden tests. In vitro cell assays will be performed in collaboration with the Clinical Research Center of Cathay General Hospital. All these works are proposed to be finished within three years. The research results will be of important contribution to the advances of cell locomotion assays, and help improve tissue engineering progression. 研究期間：9908 ~ 10007
關聯:	財團法人國家實驗研究院科技政策研究與資訊中心
顯示於類別:	[機械工程學系] 研究計畫

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