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系統識別號 U0026-1801201816334200
論文名稱(中文) 光驅動奈米平台在生物醫學的應用
論文名稱(英文) Light-triggered Nanoplatform in Biomedical Applications
校院名稱 成功大學
系所名稱(中) 化學系
系所名稱(英) Department of Chemistry
學年度 106
學期 1
出版年 106
研究生(中文) 黃昀凱
研究生(英文) Yun-Kai Huang
電子信箱 st51031520@hotmail.com
學號 L38021064
學位類別 博士
語文別 英文
論文頁數 125頁
口試委員 指導教授-葉晨聖
召集委員-廖奕翰
口試委員-蘇家豪
口試委員-何佳安
口試委員-黃志嘉
中文關鍵字 核殼結構  光熱治療  磁振造影  超音波影像  滅殺除癌錠  光驅動藥物釋放  傑納斯奈米粒子  單態氧  光動力治療 
英文關鍵字 yolk-shell  photothermal therapy  MRI  ultrasound image  methotrexate  phototriggered drug release  Janus nanoparticle  ROS  photodynamic therapy 
學科別分類
中文摘要 多年來,各式各樣的多功能奈米粒子被開發應用於奈米生醫領域,如何有效率地將奈米粒子運送至治療部位,驅動治療效果並減少對其他組織的傷害成為一個重要的課題。在各種驅動方式中,光驅動是一種較方便的驅動方式,只需要一個光源如雷射光或紅外光治療燈就可以進行藥物釋放或治療,不需要昂貴的大型儀器。另一方面,光驅動為一種非侵入式的治療方式,與傳統手術相比可以大幅減少病人的痛苦,也可以將治療範圍集中於光照射的範圍,減少其他組織的傷害及副作用。本研究開發三種不同的奈米粒子,分別設計作為光熱治療、光驅動藥物釋放、光動力治療,以下就三個主題進行介紹。
第二章介紹了碳-氧化鐵奈米複合材料的合成。外面的碳殼具有吸光產熱的功能,在雷射的照射下可以進行熱治療,另一方面,包覆在內的氧化鐵具有磁性,可以利用於磁標靶及磁振造影顯影劑。特殊的蛋黃-蛋殼結構也讓我們可以利用中間的空腔包覆低沸點的液體PFH,在照光產熱的情況下會汽化產生氣泡,作為超音波的顯影劑。在我們實驗設計上,我們先利用磁鐵吸引氧化鐵,將奈米粒子引導至腫瘤部位,使用磁振造影找到奈米粒子累積最多的時間點,再使用雷射驅動光熱治療,同時觀察超音波影像的增強現象。此奈米粒子的設計不僅擁有光驅動治療的優點,同時也有標靶及影像的多重功能,具有生物醫學臨床的潛力。
第三章我們欲利用光驅動藥物釋放的優勢,控制藥物的釋放時機,進而研究抗癌藥物”滅殺除癌錠”對癌症細胞的最佳治療方式。我們在上轉換粒子表面修飾上光斷鍵分子,並在其上接上抗癌藥物滅殺除癌錠,當上轉換奈米粒子接受到980 nm的雷射光照射,此奈米粒子會放出一紫外波段的光,此光斷鍵分子收到此紫外光,即進行斷鍵而放出抗癌藥物。在先前的研究中指出,海拉細胞上具有還原葉酸載體及葉酸受體兩個路徑可以進行滅殺除癌錠的藥物攝取,如果使用單純藥物,還原葉酸載體唯一較佳路徑,然而使用奈米粒子為載體的研究卻都使用葉酸受體這個路徑,所以我們想利用我們的光驅動藥物釋放系統來研究此兩路徑。我們將此系統與海拉細胞一同培養,在細胞外及細胞內釋放抗癌藥物,比較此兩方法對癌症細胞的毒殺能力。結果顯示,細胞內釋放似乎是一個比較好的藥物運送路徑。
第四章中我們利用了傑納斯奈米粒子兩面不同的表面特性,修飾上單態氧的產生器以及偵測器,使這個奈米粒子具有自我偵測單態氧的功能。在奈米醫學中,單態氧被使用於光動力治療。此傑納斯奈米粒子表面的TBO,在633 nm 雷射的照射下具有產生單態氧的能力,我們的研究證實此單態氧對癌症細胞有毒殺的效果。另一方面,TBO產生的單態氧可以被另一面修飾的單態氧偵測器APF偵測到,而放出螢光物質,此一結果也在螢光光譜儀上得到證實。我們開發的這個系統不僅具有光動力治療的應用價值,也可以開發進行生物影像的應用。
英文摘要 In recent years, various nanoparticles have been developed for application in biomedicine. How to deliver nanoparticles to the affected area then trigger the therapy and reduce the side effect have become a big issue. Among the triggering methods, light-triggering is a more convenient way. What you need is a light source like laser or infrared treatment instrument, then you can perform treatment or drug release. You don’t need expensive and huge facilities. On the other hand, light-triggering is a non-invasive therapy method. Compared to traditional surgery, the pain that patients suffered from can be drastically reduced. We can also focus the therapy area at wherever we want to reduce side effect. In this research, we developed three different kinds of nanoparticles for photothermal therapy, light-triggered drug release, and photodynamic therapy. The followings are the introductions.
In chapter 2, we introduced the fabrication of carbon-iron oxide nanocomposite. The carbon shell has photothermal effect. With irradiation of laser, it can produce heat for photothermal therapy. Moreover, the iron oxide encapsulated inside can serve as magnetically targeting and magnetic resonance imaging. The unique yolk-shell structure enables us to utilize the cavity for encapsulating low boiling point liquid PFH. Under photothermal effect, it will vaporize and produce bubble for the use in ultrasound imaging. In our experimental design, we guided our particles by magnet to the tumor site, and find the most accumulation time point by MRI. Later, laser was irradiated on the tumor site for photothermal therapy. At the same time, the enhanced ultrasound image was also observed. This design not only has the merit of light-triggering therapy, but also have targeting ability and multifunctional imaging, which have potential use in clinical biomedical applications.
In chapter 3, we want to use the advantage of light-triggered drug release to study the optimized therapy for cancer cells of anti-cancer drug” methotrexate” (MTX) by control the time point of drug release. We modified a photocleavable molecule and then the drug on the surface of upconversion nanoparticles. Upon irradiation of 980 nm laser, this kind of nanoparticle will emit light in UV region, which will cleave photocleavable molecule and then the drug will be released. In previous research, people showed that there are two pathways for uptake of MTX in HeLa cells. One is reduced folate carrier and another is folate receptor. If we deliver pure drug, reduced folate carrier will be a better choice. However, the research using nanoparticles as carrier deliver drug always by folate receptor. It is interesting to use our photo-triggering drug delivery system to study these two pathways. We incubated the HeLa cell with our material, and released the drug extracellularly and intracellularly to compared the cytotoxicity of cancer cell. The results showed that intracellularly delivery seems to be a better choice.
In chapter 4, we used the different surface property of Janus nanoparticles to modify ROS generator and ROS sensor, making the Janus nanoparticle have self-sensing function. In nanomedicine, ROS is used for photodynamic therapy. With the irradiation of 633 nm laser, the TBO on surface of Janus nanoparticle can generate ROS. Our research demonstrate that this generated ROS has ability to kill cancer cell. Besides, the generated ROS from TBO can be detected by APF modified on the other side of Janus nanoparticles. Upon receiving ROS, APF will release strong fluorescence species. This result can be proved by fluorescence spectra. The system we developed not only can be applied in photodynamic therapy, but also have potential application in bio-imaging study.
論文目次 摘要……………………………………………………………………………I
Abstract……………………………………………………………………...III
致謝…………………………………………………………………………..V
Content…………………………………………………………………….. VII
Figure content………………………………………………………………..XI
Chapter 1. Introduction……………………………………………………..…1
1.1 Nanoparticles in cancer therapy……………………………………….…1
1.2 Photothermal therapy………………………………………………….…2
1.3 Phototriggered drug release………………………………………………7
1.4 Photothermal therapy…………………………………………………...12
Chapter 2. Fabrication of Silica-Coated Hollow Carbon Nanospheres
Encapsulating Fe3O4 Cluster for Magnetically and MR Imaging Guided NIR Light Triggering Hyperthermia and Ultrasound Imaging……………………17
2.1 Introduction……………………………………….……………………17
2.1.1 Carbon nanoparticles………………………………………………..18
2.1.2 IONP-C core-shell nanostructure……………………………………20
2.1.3 yolk-shell carbon nanomaterials…………………………………….22
2.2 Motivation...……………………………………………………………25
2.3 Materials and instruments………………………………………………26
2.4 Experiments section ……………………………………………………29
2.4.1 Preparation of Fe3O4 NPs …………………………………………..29
2.4.2 Preparation of h-Fe3O4@P NPs……………………………………..30
2.4.3 SiO2 coated h-Fe3O4@P (SiO2/h-Fe3O4@P) NPs……………………30
2.4.4 Preparation of SiO2/h-Fe3O4@C NPs……………………………….30
2.4.5 PEG Surface modification of SiO2/h-Fe3O4@C…………………….31
2.4.6 Encapsulating of PFH in PEG coated SiO2/h-Fe3O4@C……………31
2.4.7 In vitro photothermal conversion experiments………………………31
2.4.8 Cell culture and cytotoxicity experiments…………………………..32
2.4.9 In vitro cellular uptake experiments…………………………………32
2.4.10 In vitro cell viability with laser irradiation…………………………33
2.4.11 In vivo evaluation of magnetic resonance image…………………..33
2.4.12 In vivo antitumor efficacy of PEG coated SiO2/h-Fe3O4@C containing PFH NPs…………………………...…………………………..35
2.4.13 In vitro and in vivo ultrasound imaging……………………………36
2.4.14 In vivo blood analysis, H&E stain, and biodistribution……………37
2.5 Results and discussion…………………………………………………..37
2.5.1 Synthesis and characterization of SiO2/h-Fe3O4@C…………………37
2.5.2 Photothermal effect in HeLa cells for SiO2/h-Fe3O4@C……………47
2.5.3 Loading PFH in SiO2/h-Fe3O4@C hollow structure…………………51
2.5.4 MR imaging behavior of SiO2/h-Fe3O4@C containing PFH………..53
2.5.5 Photothermal efficacy in vivo for SiO2/h-Fe3O4@C containing PFH..55
2.5.6 Echogenic behavior of SiO2/h-Fe3O4@C containing PFH…………..57
2.5.7 Toxicity of SiO2/h-Fe3O4@C containing PFH……………………….60
2.6 Conclusion……………………………………………………………...63
Chapter 3. Stimuli Triggering Antifolates Release: Extracellular or Intracellular Delivery? ……………………………………………………………………64
3.1 Introduction………………………………………………………….…64
3.1.1 Reduced folate carrier…………………………………………….…64
3.1.2 Folate receptor……………………...…………………………….…65
3.1.3 Methotrexate- a kind of antifolate…………...………………………67
3.2 Motivation……………………………………………………………...69
3.3 Materials………………………………………………………………..70
3.4 Experiment section……………………………………………….……..71
3.4.1 Synthesis of NaYF4:Yb,Tm@NaYF4 nanorods……………………..71
3.4.2 Synthesis of CS-UCNR@SiO2………………………………………72
3.4.3 Synthesis of CS-UCNR@SiO2/APTES……………………………..72
3.4.4 Synthesis of photocleavable CS-UCNR@SiO2/NPA/MTX…………72
3.4.5 Cytotoxicity of CS-UCNR@SiO2/APTES…………………………..73
3.4.6 In vitro cellular uptake………………………………………………74
3.4.7 Laser confocal images……………………………………………….74
3.4.8 Cells analysis by flow cytometry……………………………………74
3.5 Results and discussion…………………………………………………..75
3.5.1 Fabrication and characterization of CS-UCNR@SiO2/NPA/MTX…75
3.5.2 Light-triggering release of CS-UCNR@SiO2/NPA/MTX…………...81
3.5.3 In vitro cellular uptake of CS-UCNR@SiO2/NPA/MTX……………85
3.5.4 Extracellular and intracellular release………………………………..87
3.6 Conclusions …………………………………………………………….93
Chapter 4. Janus Nanoparticle as Self ROS Detection Platform and Photodynamic Therapy in Cells……………………………...………………90
4.1 Introduction……………………………………………………...……..95
4.1.1 Introduction of Janus nanoparticles…………………………………95
4.1.2 Janus nanoparticles in biomedical applications……………………..98
4.1.3 ROS detection……………………………………………………….99
4.2 Motivation………………………………………..…………………...101
4.3 Materials……………………………………………………………....102
4.4 Experiment section…………………………………………….……...102
4.4.1 Preparation of Au/PS Janus NPs……………………………………102
4.4.2 Surface modification of APF and TBO on Au/PS Janus NPs………103
4.4.3 Reactive oxygen species (ROS) transfer efficiency measurement.…104
4.4.4 In vitro cell cytotoxicity and PDT effect……………………………104
4.5 Results and discussion…………………………………………………105
4.5.1 Structure characterization of Au-PS Janus nanoparticles……..……105
4.5.2 Surface modification of Janus NPs by TBO and APF………………108
4.5.3 Self ROS detection of Janus NPs measured by fluorescence spectra.111
4.5.4 In vitro cell cytotoxicity and PDT effect of Janus-TBO-APF………112
4.6 Conclusion…………………………………………………………….116
Reference…………………………………………………………………...118
Appendix………………………………………………………………...…125
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