人類自然殺手細胞毒殺率快速檢測微型化仿生平台-以血癌細胞為例

Abstract

自然殺手細胞療法是近代癌症治療方法之一，相較於傳統化療、放射線治療等方法，其具備有治療副作用小，且可使患者維持較好的生活品質等優勢。自然殺手細胞為人體內先天免疫系統的細胞，會主動辨識癌細胞並進行攻擊，藉由分泌細胞因子或炎性物質，啟動癌細胞凋亡程序，進而達成消滅癌細胞之目的。 本研究使用微流體技術，開發一人類自然殺手細胞毒殺率快速檢測微型化仿生平台，將自然殺手細胞(NK-92MI)與標靶癌細胞(K562)接觸後，利用微流道出入口水頭壓力差的原理，將懸浮的細胞混合液導入晶片當中，並以微結構捕捉固定後，利用螢光染色法來判定 K562細胞是否存在早期細胞凋亡的現象，用以檢測NK-92MI對 K562細胞毒殺率。利用計算流體力學（computational fluid dynamics, CFD）來預估微流體中應力作用對細胞的影響，確認在此微系統環境下，流場剪應力對於細胞存活率並無顯著影響，以增加微型化仿生平台檢測可靠度。實驗方面針對傳統流式細胞儀與本平台，比較不同接觸時間與不同效應細胞對目標細胞比例(effector to target cell ratio, E:T ratio)混合之樣本，進行自然殺手細胞毒殺能力的檢測與分析。結果顯示使用微型化仿生平台所獲得的結果與流式細胞儀一致性極高，此外本平台能捕捉少量(約〖10〗^2)懸浮細胞樣本，提供自然殺手細胞療法一嶄新的毒殺率檢測方式。並實際抽取人體血樣，驗證人類自然殺手細胞毒殺率變化，證明此裝置於自然殺手細胞療法中對醫師有臨床檢測功效。未來可規劃循環式流動系統，以達模擬真實血管中自然殺手細胞毒殺癌細胞的作用進行觀察。 本微型化仿生平台成功且有效檢測出自然殺手細胞之細胞毒殺率，可應用於為患者尋找合適的自然殺手細胞捐贈來源，提高自然殺手細胞療法成功率，實現個人化醫療(personalized medicine)的目標。此外本平台尚擁有製造低成本和低樣品消耗量等特性，充分具備未來於醫療實務應用上的發展潛力。

Natural killer cell therapy is one of the most recent forms of cancer treatment. Natural killer cell therapy has fewer side effects and better quality of life than chemotherapy and radiotherapy. Natural killer cells are cells of the innate immune system in the human body. They actively attack cancer cells by secreting cytokines or inflammatory cell apoptosis programs. Cytotoxicity assays were performed on natural killer cells and K562 cells. By manipulating microfluidics, the cell mixture was injected into the chip to detect cytotoxicity. In this study, computational fluid dynamics (CFD) software was used to study the effect of shear stress on microfluidic cells. It was confirmed that the shear stress had no significant effect on cell in the flow. Increase the reliability of the miniaturized biomimetic platform. The cells are mixed according to the effector to target cell ratio (E: T ratio). The cytotoxicity of natural killer cells was compared with traditional flow cytometry. The results show that the analytical results using a miniaturized biomimetic platform are highly consistent with flow cytometry. In addition, the miniaturized biomimetic platform can capture a small amount of suspended cell samples (about 10^2 cells) and improve the shortcomings that flow cytometry requires a large number of cells (about 5 × 10^5 ~10^6). Blood was drawn off from donors to verify changes in cytotoxicity. This device proved to be clinically effective for physicians in natural killer cell therapy. The miniaturized biomimetic platform successfully and effectively detects the cytotoxicity of natural killer cells. It can be used to find suitable natural killer cell donation sources for patients. Increase the success rate of natural killer cell therapy. In addition, this platform still has manufacturing features such as low cost and low sample consumption. This research has the potential for development in practical medical treatment.