指導教授：李嗣涔臺灣大學：生醫電子與資訊學研究所蔡尚儒Tsai, Shang-RuShang-RuTsai2014-11-262018-07-052014-11-262018-07-052014http://ntur.lib.ntu.edu.tw//handle/246246/261849近年研究發現，紅外光可調控生物體的生理機能，活絡粒線體內的電子傳遞鏈，影響蛋白質含量和代謝，進而影響生物體成長形態。本論文主要是設計結合紅外光發射器之細胞培養箱，並探討癌症治療的過程中，癌細胞受不同波段紅外光照射和調控，其抑癌的效果與機制。 本文的第一個主題，是探討 3~5 um窄頻中紅外光影響肺癌細胞 A549的療效。利用三層結構 (金/二氧化矽/金)的波導熱輻射發射器(Waveguide thermal emitter)，其上下兩層金之間的二氧化矽激發共振腔膜態產生窄頻紅外光源，可抑制肺癌細胞 A549生長，造成癌細胞粒線體膜電位的異常。 本文的第二個主題，是探討 3~5 um窄頻中紅外光影響 Paclitaxel對子宮頸癌細胞HeLa的療效。利用波導熱輻射發射器產生窄頻紅外光源照射子宮頸癌細胞 HeLa，可提升 Paclitaxel的治療效果，破壞粒線體膜電位，刺激下游的凋亡訊息路徑，造成更多子宮頸癌細胞 HeLa死亡。 本文的第三個主題，是探討 810 nm近紅外光雷射結合光動力療法對骨癌細胞 MG-63的療效。研究發現 810 nm近紅外光雷射可影響細胞的電子傳遞鏈和產生大量ATP，造成癌細胞吸收更多標靶於溶酶體的 Mono-L-aspartyl chlorin(e6) (NPe6)，接著由紅光激發光敏劑引起光動力效應，產生大量活性氧毒殺癌細胞。 本文的研究成果可讓我們了解，利用紅外光適當調控生物體的生理機能，結合癌症治療，可以加強抑制惡性腫瘤細胞。未來也可應用於臨床和家庭醫療，透過非侵入性、低副作用的特性，可減少傷害正常人體組織，達成更好的功效。Recent studies found that infrared radiation (IR) can regulate physiology of organisms, stimulate mitochondrial electron transport chain, and affect expression of the genes, thereby regulating the morphology of cell growth. The goal of this thesis is to design the cell incubator equipped with IR emitter, and investigate the effectiveness and mechanisms of different wavelengths of IR on the cancer therapy. In the first topic of this thesis, it was clarified that the effect of mid-infrared radiation (MIR) with the peak wavelengths of 3~5 um range on the growth of lung cancer cell A549. The tri-layers Au/SiO2/Au waveguide thermal emitter (WTE) was used as the light source of narrow band IR. IR was originated from the resonance of cavity mode generated in the SiO2 layer between two Au films. The results showed that IR can inhibit the growth of cancer cell A549, leading to the collapse of mitochondrial membrane potential. In the second topic of this thesis, it was investigated that the effects of HeLa cells treated with Paclitaxel combined with 3~5 um MIR. Narrow band IR from WTE can promote the effectiveness of Paclitaxel through the collapse of mitochondrial membrane potential. In the third topic of this thesis, it was investigated that the effects of combination treatment of 810 nm near infrared radiation (NIR) laser and photodynamic therapy (PDT) in osteosarcoma cancer cell MG-63. The data showed that 810 nm NIR laser can regulate the electron transport chain and generate a large amount of ATP to enhance the uptake of lysosome-localized photosensitizer mono-L-aspartyl chlorin (e6) (NPe6). Therefore, overproduction of reactive oxygen species was activated through red light-induced PDT effects. The results of this thesis imply that the combination of cancer therapy and IR-regulated physiological activity can enhance the therapeutic effects on cancer cells. In future, IR can be applied to clinical and home health care to achieve therapeutic purposes without damaging normal tissue based on the characteristics of non-invasive and low side effects.致謝...................................................I 中文摘要..............................................III Abstract..............................................IV Contents..............................................VI List of abbreviations.................................XI Figure legends.......................................XII Chapter 1 Introduction.................................1 1.1 The mechanism of infrared in photobiology..........1 1.1.1 IR and mitochondria..............................5 1.1.2 IR and ROS.......................................7 1.1.3 IR and water molecule............................11 1.2 The objectives of this thesis......................13 1.3 The framework of this thesis.......................15 Chapter 2 The infrared light source....................18 2.1 The fundamentals of waveguide thermal emitter......18 2.1.1 Surface plasmons on smooth surfaces..............18 2.1.2 Surface plasmons on the metal surface perforated with hole arrays............................................23 2.2 The extraordinary light transmission and waveguide mode...................................................26 2.3 Fabrication and measurement of infrared emitter....28 2.4 Incubator for cell illuminated by IR...............36 2.5 The light source used in combination treatment of NIR and PDT................................................37 Chapter 3 The effect of narrow band 3~5 um MIR on the growth of lung cancer cell A549 for 48 hours...........38 3.1 Motivation.........................................38 3.2 Material and methods...............................42 3.2.1 Measurement of cell number and diameter..........42 3.2.2 Cell mitochondrial membrane potential analysis...44 3.3 Results and discussion.............................45 3.3.1 MIR inhibits the growth of A549 cells............45 3.3.2 MIR increases collapse of mitochondrial membrane potential..............................................48 Chapter 4 The effect of narrow band 3~5 um MIR in enhancing the antitumor activity of paclitaxel...................51 4.1 Motivation.........................................51 4.2 Material and methods...............................55 4.2.1 Cell culture.....................................55 4.2.2 Measurement of cell viability....................55 4.2.3 Measurements by Annexin V-FITC/PI (Propidium iodide) staining...............................................56 4.2.4 Cell mitochondrial membrane potential analysis...56 4.2.5 Oxidative stress analysis........................57 4.3 Results............................................58 4.3.1 Dosages of paclitaxel for HeLa cells.............58 4.3.2 MIR enhanced antitumor activity of paclitaxel....60 4.3.3 MIR increases apoptosis in HeLa cells treated with paclitaxel.............................................62 4.3.4 MIR-induced collapse of mitochondrial membrane potential of HeLa cells treated with paclitaxel........64 4.3.5 MIR increases ROS generation in HeLa cells treated with paclitaxel........................................66 4.4 Discussion.........................................68 Chapter 5 The effect of 810 nm NIR laser potentiates NPe6-mediated PDT...........................................71 5.1 Motivation.........................................71 5.2 Material and methods...............................74 5.2.1 Cell culture.....................................74 5.2.2 Cell viability and PDT treatment.................74 5.2.3 Cellular uptake of NPe6..........................77 5.2.4 Measurement of ROS generation induced by NIR combined with PDT...............................................77 5.2.5 Cell Localization of NPe6 via Confocal Microscopy.78 5.2.6 ATP activity assay................................79 5.2.7 Pretreatment with mitochondrial respiratory chain inhibitor antimycin A...................................79 5.2.8 Statistical Analysis..............................80 5.3 Results.............................................80 5.3.1 NIR-LLLT enhanced PDT effectiveness...............80 5.3.2 Cellular uptake of NPe6 after NIR-LLLT............82 5.3.3 NIR-LLLT increases PDT-induced ROS generation.....82 5.3.4 Intracellular localization of NPe6 in MG-63 cells.85 5.3.5 Activation of ATP production by NIR-LLLT..........85 5.3.6 NIR-LLLT potentiation of NPe6 PDT cytotoxicity is abrogated by antimycin A, an inhibitor of ATP synthesis.88 5.4 Discussion..........................................90 Chapter 6 Conclusion and perspectives...................93 6.1 Conclusion..........................................93 6.2 Perspectives........................................98 References.............................................1003404434 bytesapplication/pdf論文公開時間：2017/08/17論文使用權限：同意有償授權(權利金給回饋學校)光動力療法低能量光療法波導熱輻射紅外光發射器HeLa 子宮頸癌細胞A549 肺癌細胞MG-63骨肉癌細胞[SDGs]SDG3thesishttp://ntur.lib.ntu.edu.tw/bitstream/246246/261849/1/ntu-103-F95921105-1.pdf