腫瘤晶片之微流體平台開發

Abstract

在此研究中開發一種腫瘤晶片(Tumor-on-a-chip)，藉由血管新生的技術，培養出可模擬體內腫瘤之體外微組織模型，以期提供可透過功能性微血管網路向腫瘤投以抗癌藥物，進行藥物篩選的研究。為了在體外實現腫瘤組織微環境，我們分別透過對流、擴散流體效應設計出了兩種可培養三維微組織的微流體平台，通過微流體系統對微腔室控制靜水壓力，氧氣張力及養分梯度做精準的控制，並找出各種培養參數如:間質滲流流速、coating時間、培養時間、氧氣濃度等不同條件下對於微血管組織培養的影響，最終我們成功在擴散質傳效應的流場中培養出三維的微組織。由實驗中發現，在以擴散主導的質傳機制下，可以成功促進血管生成(vasculogenesis)，達到35~45%血管面積比的微血管網路，並在缺氧環境(5%O2)下成功使癌組織缺氧，促使癌細胞分泌生長因子，在生長因子(VEGF)的濃度梯度下，發生腫瘤血管新生(tumor angiogenesis)，為了達成血管功能，並使血管與外部流道連接，我們透過在微流道coating一層纖維蛋白及內皮細胞，成功連接腔室內血管。由實驗結果證實，所發展的腫瘤晶片系統在培養17天之後的血管新生長度比起培養14天多了87.2%的長度，可達到1304.98μm，而在血管分支數上，培養17天(7.3 branches/vessel)也比起14天(3 branches/vessel)達到超過兩倍的分支數量，成功形成具微血管的組織結構。 透過本研究所開發出的腫瘤晶片，我們希望能在未來建立一標準的體外腫瘤模型，可以更接近體內由原來的微血管新生至腫瘤的機制，有效重現體內微環境，並通過此微流體晶片模擬體內複雜交互作用下的奈米藥物輸送，最終達到根據不同患者的篩藥結果來設計個人化的藥物治療平台，並加速前期藥物的篩選時程以及不同癌細胞間的研究。

In this study, a Tumor-on-a-chip device is developed. The goal of this project is to stimulate vasculogenic and angiogenic processes on a dual chamber device to develop a vascularized tumor model for the study of anti-cancer drugs. To develop a tumor tissue microenvironment in vitro, we design two microfluidic platforms that can develop three-dimensional microtissues using convective and diffusive mass transports. The microfluidic system controls the hydrostatic pressure, oxygen tension, and nutrient gradients inside the dual microtissue chambers. Optimization is conducted to identify culturing parameters, including interstitial flow rate, coating time, culture time, and oxygen tension. Based on the experimental study and finite element analysis, we successfully develop microvasculature in a vasculogenic chamber and induce angiogenic sprouting into adjacent tumor chamber. It is found that using diffusion-dominated mass transport, vasculogenesis can be successfully stimulated and reach 35% to 45% of the vessel area ratio. Furthermore, incubating the device in 5% oxygen incubator, SW480 tumor cells and fibroblast coculture can secret angiogenic factors. These angiogenic factor forms a concentration gradient between the dual chambers and stimulate angiogenic sprouting from vascularized tissue in the adjacent chamber. To achieve functional blood vessels and connect them with external flow channels, a layer of fibronectin and endothelial cells are lined on the microchannels. It is demonstrated that blood vessels can form connections to the external channels. Using the developed tumor-on-a-chip device, the total length of 17-day angiogenic grew vessels can be 87.2% longer than that of vessels grew in 14 days, and the length can be as long as 1304.98 μm. Furthremore, branches of sprouted vessels of 17 days also has more than twice of that in 14-day culture. In summary, we successfully develop a microfluidic device that can induce both vasculogenic and angiogenic processes on a chip and induce angiogenic sprouting toward the microtumor. The maturation of this technology can provide a vascularized tumor model that can mimic the in vivo tumor for drug delivery studies.