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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 彭福佐(Fu-Chuo Peng) | |
dc.contributor.author | Wan-Ching Tsai | en |
dc.contributor.author | 蔡宛靜 | zh_TW |
dc.date.accessioned | 2021-06-16T16:05:26Z | - |
dc.date.available | 2018-09-24 | |
dc.date.copyright | 2013-09-24 | |
dc.date.issued | 2013 | |
dc.date.submitted | 2013-06-21 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/62604 | - |
dc.description.abstract | 氧化鋅(ZnO)是一種白色粉末,不溶於水,它被廣泛用作許多材料和產品中的添加劑,包括塑料、塗料、食品等等。在材料科學中的應用,氧化鋅具有高折射率,高導熱性,抗菌,防紫外線的性能及功效,因為ZnO具有可反射阻隔紫外線的效果,現今為人廣泛使用在防曬用產品。隨著奈米科技的逐步發展,奈米粒子之材料已應用於日常生活中,如防曬用產品也將ZnO奈米化為更能達到防曬效果。然而,有研究指出奈米級顆粒可以進入體內影響人類細胞,如人類肝細胞 (human liver cells, HepG2)。由於防曬產品首先接觸是皮膚,而文獻尚未詳細報導氧化鋅奈米粒子 (Zinc oxide nanoparticles, ZnO NPs) 對皮膚角質細胞的影響,因此本研究選用人類角質細胞(human keratinocyte cell, HaCaT cell) 來探討皮膚暴露於ZnO NPs後產生細胞毒性、氧化壓力以及細胞凋亡的傷害。細胞存活率顯示HaCaT cell暴露24小時於半抑制濃度(Concentration of 50 % inhibition, IC50) 50μg/ml的ZnO NPs,藉由流式細胞儀和螢光顯微鏡發現ZnO NPs對HaCaT cells 會產生活性氧化物(reactive oxygen species, ROS)。另外,以Annexin V 染劑分析細胞凋亡及 PI 染劑分析細胞壞死,結果顯示HaCaT細胞在18小時產生凋亡反應、24小時產生壞死現象,除此之外,以偵測細胞凋亡的 Hoechst 33342 染劑也顯現處理50 μg/ml ZnO NPs 18小時後產生凋亡反應。原子力顯微鏡(Atomic force microscopy, AFM ) 是一個高解析度的探針掃描顯微鏡,對於觀察細胞上奈米級的影象微結構是相當強大的工具,並且相當適合以物理特性測量細胞上的改變,所以AFM提供了用來研究細胞凋亡形態學特徵的技術優勢。HaCaT 細胞經由處理了ZnO NPs,細胞表面的粗糙度增加、硬度減少、黏滯力無明顯改變。綜合以上結果,本研究以生化與生物物理特徵之實驗數據證實ZnO NPs引發HaCaT 細胞死亡是藉由誘導ROS和細胞凋亡。 | zh_TW |
dc.description.abstract | Zinc oxide (ZnO) is a white powder that is insoluble in water, which is widely used as an additive in numerous materials and products including plastics, paints, foods. For material science applications, zinc oxide has high refractive index, antibacterial and UV-protection properties. ZnO are used in various products, including cosmetics and sunscreens due to UV-filtering properties. The production of engineered nanoparticles is growing rapidly as the field of nanotechnology continues to expand. Modern sunscreen contain insoluble ZnO nanoparticle (ZnO NPs), which is reflect ultraviolet (UV) more efficiently than larger particles. However, cosmetic research suggest that vesicle materials may enter human cell, like human liver cells (HepG2). Sunscreen products is the first contact with the skin, and the underlying mechanisms of zinc oxide adverse effects have not been fully characterized. This study was designed to investigate the cytotoxicity, oxidative stress and apoptosis by ZnO nanoparticles in human keratinocyte cell (HaCaT cell). Cell viability assays indicated an IC50 approximately 50 μg/ml of ZnO nanoparticles after 24 h of exposure. ZnO nanoparticles were found to induce reactive oxygen species (ROS) generation in HaCaT cells by flowcytometry and fluoresce microscopy. ZnO nanoparticles were also investigated using Annexin V staining for apoptosis analysis and P2 staining for necrosis analysis. ZnO nanoparticles displaced apoptosis at 18 hr and then necrosis at 24 hr. Additional, Hoechst 33342 staining revealed apoptosis in HaCaT cells after 50 μg/ml ZnO nanoparticles 18hr treatment. Atomic force microscopy (AFM) is a very high resolution type of scanning probe microscopy. It has been shown to be a powerful tool for imaging materials at the nanometer level and for observing the ultrastructure of a cell. This method is appropriate for measuring the change in the biophysical properties of the cell. AFM offers an advantages over morphological characterization technique used to study apoptosis. After ZnO nanoparticles treatment the HaCaT cells, the surface roughness was increased, the stiffness was decreased and the adhesion was no significant changed. In conclusion, this study demonstrated that ZnO nanoparticles induced cell death of HaCaT cells was via induction of ROS and apoptosis by biochemical feature and AFM. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T16:05:26Z (GMT). No. of bitstreams: 1 ntu-102-R00447004-1.pdf: 4521925 bytes, checksum: 5f88ede9d6da8c0b74d60f2e10beee17 (MD5) Previous issue date: 2013 | en |
dc.description.tableofcontents | 目錄
第一章 緒言(introduction) 1 1-1奈米科技 1 1-1-1氧化鋅(Zinc Oxide, ZnO) 2 1-1-2奈米氧化鋅 (Zinc Oxide nanoparticles, ZnO NPs) 3 1-2人類角質細胞 5 1-3 原子力顯微技術(Atomic Force Microscope, AFM) 7 1-3-1源起 7 1-3-2 AFM優缺點 10 1-3-3 AFM的裝置 11 1-3-4 AFM原理 13 1-3-5 AFM的模式 16 1-3-6三種模式的比較 19 1-3-7樣品的要求 20 1-3-8 AFM在生物醫學上的應用 21 1-4 研究動機與實驗目的 (Motivations and aims) 22 第二章 材料與方法 23 2-1實驗材料 23 2-1-1實驗藥品 23 2-1-2實驗儀器 24 2-2實驗方法 25 2-2-1細胞培養(cell culture) 25 2-2-2細胞冷凍保存 25 2-2-3細胞存活率測試(MTT assay) 25 2-2-4細胞氧化自由基測試(ROS assay) 26 2-2-5細胞凋亡測試(cell apoptosis assay) 27 2-2-6原子力顯微術掃描 28 第三章 論文結果 32 3-1 HaCaT 人類角質細胞的型態 32 3-2 奈米氧化鋅對於HaCaT 人類角質細胞存活率的影響 32 3-3 奈米氧化鋅處理HaCaT 人類角質細胞產生氧化自由基的影響 32 3-4 奈米氧化鋅處理HaCaT 人類角質細胞造成細胞凋亡的影響 33 3-5 原子力顯微鏡檢測奈米氧化鋅處理HaCaT人類角質細胞造成細胞凋亡的影響 34 3-5-1原子力顯微鏡檢測凋亡細胞型態結構的變化 34 3-5-2原子力顯微鏡檢測凋亡細胞粗糙度的變化 34 3-5-3原子力顯微鏡檢測凋亡細胞高度的變化 34 3-5-4原子力顯微鏡檢測凋亡細胞黏滯力和軟硬度的變化 35 第四章 討論 36 第五章 未來展望 40 第六章 結論 41 第七章 參考文獻 42 附圖目錄 附圖 1:奈米尺度規模大小 46 附圖 2:奈米材料在醫療臨床上的運用。 47 附圖 3:European Commission(EU)統計的奈米成長趨勢圖。 48 附圖 4:防曬的運用原理及原料分為化學濾光劑和物理反光劑 49 附圖 5:奈米氧化鋅大於20 nm 能穿透可見光 50 附圖 6:奈米氧化鋅及二氧化鈦奈米顆粒大小的穿透度 51 附圖 7:比較氧化鋅的顆粒差異 52 附圖 8:皮膚構造為表皮(epidermis)、真皮(dermis)及皮下組織(subcutis) 53 附圖 9:常見顯微術比較 54 附圖 10:技術演進與功能 55 附圖 11:原子力顯微鏡的硬體架構示意圖 56 附圖 12:微小懸桿(cantilever)藉由折返移動來感測原子之間力的變化量 57 附圖 13:原子力顯微鏡懸臂樑及針尖示意圖 58 附圖 14:原子力顯微鏡結合系統將樣本的表面特性呈現示意圖 59 附圖 15:電子雲斥力的作用圖 60 附圖 16:光二極體量測示意圖(光槓桿偵測原理) 61 附圖 17:原子力顯微鏡掃描探針的操作模式 62 附圖 18:酵素免疫分析測讀儀 63 附圖 19:流式細胞儀 63 附圖 20:螢光顯微鏡 63 附圖 21: SPA 300HV 原子力顯微鏡 64 附圖 22:80×80μm2壓電掃描頭 64 附圖 23:細胞存活率測試示意圖 65 附圖 24:螢光顯微鏡偵測DHE染劑示意圖 66 附圖 25:Annexin V及PI染劑偵測細胞凋亡的原理示意圖 67 附圖 26:螢光顯微鏡偵測Hoechst 33342染劑示意圖 68 附圖 27:原子力顯微鏡系統所量測到探針的共振頻率圖 69 附圖 28:原子力顯微鏡氣相專用探針載台 70 附圖 29:原子力顯微鏡掃描細胞影像求得粗糙值參數Ra、Rt及Rms 71 附圖 30:原子力顯微鏡在力-距離曲線下的軟硬度與黏滯力分析 72 附圖 31:奈米顆粒聚集形成聚集體 73 附圖 32:人體內存在大量的抗氧化劑 74 附圖 33:服用或接觸的藥物可能使細胞造成損傷 75 附圖 34:未來展望的細部流程圖 76 結果圖目錄 圖 1:人類角質細胞(HaCaT)的細胞型態 77 圖 2:奈米氧化鋅對人類角質細胞HaCaT之細胞存活率 78 圖 3:奈米氧化鋅對人類角質細胞HaCaT產生之氧化自由基反應 79 圖 4:奈米氧化鋅對人類角質細胞HaCaT產生最大量氧化自由基 80 圖 5:螢光顯微鏡觀測氧化自由基 81 圖 6:奈米氧化鋅對人類角質細胞HaCaT產生之細胞凋亡反應 82 圖 7:奈米氧化鋅對人類角質細胞HaCaT產生細胞凋亡量化圖 83 圖 8:螢光顯微鏡觀測細胞凋亡 84 圖 9:原子力顯微鏡檢測奈米氧化鋅處理下凋亡細胞型態結構的變化 85 圖 10:原子力顯微鏡檢測過氧化氫處理下凋亡細胞型態結構的變化 86 圖 11:原子力顯微鏡檢測奈米氧化鋅處理下凋亡細胞粗糙度的變化 87 圖 12: 以數值比較奈米氧化鋅與過氧化氫凋亡正控制組高度的變化 88 圖 13:原子力顯微鏡檢測奈米氧化鋅處理下凋亡細胞黏滯力的變化 89 圖 14:原子力顯微鏡檢測奈米氧化鋅處理下凋亡細胞軟硬度的變化 90 表格目錄 表格 1: 以數值比較奈米氧化鋅與過氧化氫凋亡正控制組粗糙度的變化 91 表格 2: 奈米氧化鋅與過氧化氫凋亡正控制組比較凋亡細胞高度的變化 92 表格 3:奈米氧化鋅與過氧化氫凋亡正控制組比較凋亡細胞黏滯力與硬度的變化 93 | |
dc.language.iso | zh-TW | |
dc.title | 以原子力顯微鏡探討奈米氧化鋅粒子經活性氧自由基造成人類角質細胞凋亡之現象 | zh_TW |
dc.title | Zinc oxide nanoparticles induce ROS mediated apoptosis in human keratinocyte cells research by atomic force microscopy | en |
dc.type | Thesis | |
dc.date.schoolyear | 101-2 | |
dc.description.degree | 碩士 | |
dc.contributor.coadvisor | 林世明(Shi-Ming Lin) | |
dc.contributor.oralexamcommittee | 洪錦堂(Jim-Tong Horng) | |
dc.subject.keyword | 奈米氧化鋅,人類角質細胞,活性氧化物,細胞凋亡,原子力顯微鏡, | zh_TW |
dc.subject.keyword | Zinc oxide nanoparticles (ZnO NPs),Human keratinocyte,reative oxygen species (ROS),apoptosis,atomic force microscopy (AFM ), | en |
dc.relation.page | 93 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2013-06-21 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 毒理學研究所 | zh_TW |
顯示於系所單位: | 毒理學研究所 |
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