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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 沈立言(Lee-Yan Sheen) | |
dc.contributor.author | Jung Pao | en |
dc.contributor.author | 包容 | zh_TW |
dc.date.accessioned | 2021-06-08T04:18:25Z | - |
dc.date.copyright | 2011-08-22 | |
dc.date.issued | 2011 | |
dc.date.submitted | 2011-08-18 | |
dc.identifier.citation | Adams JM, Cory S (1998) The Bcl-2 protein family: arbiters of cell survival. Science 281: 1322-1326
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/22465 | - |
dc.description.abstract | 皮膚癌是全球罹患率最高的癌症之一,可分為非黑色素細胞癌 (non-melanoma skin cancer, NMSC)與黑色素細胞癌(melanoma),而黑色素細胞癌(melanoma)是皮膚癌中最惡性的一種,容易轉移至肝、肺及腦,亦會抵抗化學療法及放射線療法,致死率較高。大蒜精油活性成分-二丙烯基硫化物(allyl sulfides)以二丙烯基單硫化物(diallyl sulfide, DAS)、二丙烯基二硫化物(diallyl disulfide, DADS)與二丙烯基三硫化物(diallyl trisulfide, DATS)對多種癌細胞株具有抑制活性,但目前大蒜精油在人類皮膚癌細胞的研究卻很少見。因此,本研究以水蒸氣蒸餾法萃取大蒜精油,並利用氣相層析儀分析其DAS、DADS及DATS分別約佔總精油量的5、61及22%,並進一步以體外及體內試驗探討大蒜精油對人類黑色素細胞癌之抑制效果及分子機制。於探討大蒜精油抑癌機轉方面,大蒜精油可抑制人類黑色素癌A375細胞株之細胞存活率,並具有劑量與時間效應;流式細胞儀及西方墨點法之結果發現,大蒜精油會導致細胞週期停滯於G2/M期,並改變粒線體膜通透性(mitochondrial membrane permeability transition, MPT)及促進細胞內活性氧物質(reactive oxygen species, ROS)的生成,進一步啟動粒線體活化路徑,包括增加A375細胞Bax/Bcl-2的比值,促使caspase-9、caspase-3及poly(ADP-ribose)polymerase (PARP)明顯有裂解產物的表現,誘導A375細胞凋亡的發生。在細胞轉移實驗方面,大蒜精油具有降低基質金屬蛋白酶(matrix metalloproteinase, MMP)的活性,如:MMP-2與MMP-9,有效抑制A375細胞移動(migration)與侵襲(invasion)的能力。在細胞增生實驗部分,利用固著依賴性細胞菌落分析(anchorage-dependent colony formation assay),大蒜精油可降低A375細胞株形成群落的能力,並經由MEK/MAPK [MAPK (mitogen-activated protein kinase)/ERK (extracellular signal regulated kinase) kinase]路徑抑制A375細胞增生。由以上結果證實,大蒜精油可藉由誘導A375細胞週期停滯G2/M期與細胞凋亡,並可抑制A375細胞株的轉移及增生。
本研究進一步以A375細胞株異種移植的模式,探討大蒜精油及DATS對黑色素細胞癌之抑制效果。將nu/nu裸鼠分為五組,分別為控制組 (葵花油0.1 mL)、大蒜精油低劑量組(58.8 mg/kg bw)與高劑量組(117.6 mg/kg bw)、DATS組(66.7 mg/kg bw)、正對照組(camptothecin i.v. 2 mg/kg bw )。裸鼠分別管餵大蒜精油或DATS一週後,將A375細胞株以皮下注射方式接種於裸鼠中,實驗期間每兩天管餵一次,記錄其體重、攝食量與腫瘤體積的變化,連續九週後犧牲,並進行血液生化值的分析。結果顯示,大蒜精油高劑量組(117.6 mg/kg bw) 及DATS可有效抑制腫瘤體積與重量,對腫瘤體積之抑制率分別為62及84% (p<0.05),而對腫瘤重量之抑制率分別為45及83% (p<0.05);控制組與處理組之間的體重、肝功能指數(AST、ALT)及腎功能指數(BUN、Creatine) 皆沒有顯著差異,顯示大蒜精油及DATS並不會造成不良的影響。由細胞與動物實驗的結果推論,大蒜精油具有抑制黑色素細胞癌生長與轉移的作用之潛力。 | zh_TW |
dc.description.abstract | The incidence of melanoma increased more rapidly than any other cancer in recent years. Malignancies arising in the skin are broadly divided into two categories: melanoma and non-melanoma skin cancers. Melanoma is the most serious, highly invasive and metastatic skin tumor, and even resistant to chemotherapy. Numerous studies have shown that garlic (Allium sativum L.) oil and their major active compounds - diallyl sulfide (DAS), diallyl disulfide (DADS) and diallyl trisulfide (DATS) exhibit the anticancer activity against several cancer types, but their effects and related mechanisms of garlic oil on human melanoma cells were not clear yet. The objective of our study was to investigate the anticancer effect of garlic essential oil on human melanoma A375 cells in vitro and in vivo assays. Garlic essential oil is obtained by steam distillation which yield accounts for 0.2–0.5%. We have evaluated the profiles of allyl sulfides in garlic oil, including DAS (5%), DADS (61%), DATS (22%) using gas chromatography.
The results indicated that garlic oil inhibited cell viability in a dose dependent manner. Flow cytometry analysis and Western blot results showed garlic oil significantly induced G2/M phase arrest of A375 cells. We further demonstrated that garlic oil elicited apoptotic events such as mitochondrial membrane depolarization, increasing intracellular reactive oxygen species (ROS) generation, activation of caspase-9, caspase-3 and poly(ADP-ribose)polymerase (PARP). Additionally, garlic oil inhibited cell migration and invasion of A375 cells by in vitro anti-invasive assay. The activities and protein expressions of matrix metalloproteinase (MMP)-2 and -9 in A375 cells were inhibited by treatment with garlic oil. Moreover, we also showed that garlic oil suppressed anchorage-independent growth of A375 cells. The anti-proliferative potency might relate to inhibit phosphorylation of MEK/MAPK [MAPK (mitogen-activated protein kinase)/ERK (extracellular signal regulated kinase) kinase] pathway. We also studied the anti-tumor activity of garlic oil and DATS in A375 tumor-bearing nude mice model. Garlic oil and DATS was administered for 1 week before inoculation of A375 cells and continued for 9 weeks. This results indicated that oral administration of garlic oil (117.6 mg/kg bw) and DATS (66.7 mg/kg bw) has potential to reduce tumor volume and weight as compared with negative control in xenografted nude mice; tumor volume was inhibited by 62 and 84% (p<0.05); tumor weight was inhibited by 45 and 83% (p<0.05), respectively. Furthermore, there was no significant difference in body weights among the various groups at the end of the study. The AST, ALT, BUN and creatine values showed no significant difference between treatment groups and control group. The results implied that garlic oil and DATS feeding to mice did not appear to induce any adverse effects. In conclusion, garlic oil inhibited cell growth of human melanoma cells via induction of cell cycle arrest and apoptosis, and might provide the anti-metastatic and anti-proliferative potency. Therefore, these finding demonstrated that garlic oil and DATS exhibit anti-tumor effects in vivo model. These results suggest that garlic oil provide the potential anticancer effect on anti-melanoma. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T04:18:25Z (GMT). No. of bitstreams: 1 ntu-100-R98641037-1.pdf: 4643322 bytes, checksum: 4ec3a8b38fb101d5008b5d980a6e8d9d (MD5) Previous issue date: 2011 | en |
dc.description.tableofcontents | 中文摘要 I
英文摘要 III 縮寫對照表 V 表 次 XII 圖 次 XIII 第一章 前言 1 第二章 文獻回顧 2 第一節 皮膚癌(Skin cancer) 2 一、皮膚癌種類與發生率 2 二、皮膚癌之危險因子 5 第二節 飲食與皮膚癌 12 第三節 黑色素細胞癌的轉移 16 一、黑色素細胞癌的分期 16 二、基質金屬蛋白酶(Matrix metalloproteinases, MMPs) 21 三、MAPK (Mitogen-activated protein kinase) 24 第四節 細胞生長週期與細胞凋亡(Cell cycle and apoptosis) 28 一、細胞週期的調控 29 二、細胞凋亡的主要路徑 30 三、氧化壓力與細胞凋亡 38 第五節 大蒜之生理功能 39 一、大蒜的組成分 40 二、大蒜含硫化合物及其生理功效 41 三、大蒜之生理功能 47 四、大蒜含硫化合物及其生物可利用性 55 五、大蒜的安全性研究 56 第三章 研究目的與實驗架構 57 第一節 研究目的 57 第二節 實驗架構 58 第四章 實驗材料與方法 61 第一節 實驗材料與儀器設備 61 一、實驗樣品-大蒜精油 61 二、實驗細胞株 62 三、化學藥品與試劑 63 四、儀器設備 64 五、動物來源與管理 67 第二節 實驗方法 67 一、細胞株之解凍、繼代培養及保存 67 二、細胞存活率分析(Cell viability analysis) 69 三、利用倒立式顯微鏡觀察細胞形態 70 四、利用DAPI染色之觀察(DAPI staining for nuclear morphology detection) 70 五、細胞週期分析(Cell cycle analysis) 71 六、細胞粒線體膜電位檢測(ΔΨm determination) 73 七、測定細胞內活性氧物質含量(Intracellular ROS measurement via carboxy-H2DCFDA) 73 八、免疫螢光染色(Immunofluorescence staining) 75 九、西方墨點法(Western blot analysis) 75 十、細胞移行實驗(Cell migration assay) 77 十一、細胞侵犯能力試驗(Matrigel invasion assay) 78 十二、明膠蛋白酵素電泳法(Gelatin zymography) 78 十三、固著依賴性細胞菌落分析(Anchorage-dependent colony formation assay) 79 十四、動物試驗 80 十五、統計分析 82 第五章 實驗結果 83 第一節 氣相層析儀(Gas chromatograms)分析大蒜精油其活性成分之含量 83 第二節 大蒜精油對人類黑色素細胞癌A375細胞生長之影響 83 一、大蒜精油對A375細胞的存活率 83 二、大蒜精油對人類正常角質細胞HaCaT細胞株的存活率 84 三、大蒜精油對A375細胞型態的影響 84 四、大蒜精油對A375細胞週期的影響 84 五、利用西方點墨法檢測A375細胞週期蛋白質表現 85 第三節 大蒜精油誘導人類黑色素細胞癌A375細胞凋亡 86 一、以免疫螢光染色觀察大蒜精油對A375細胞骨架之影響 86 二、利用DAPI螢光染色法檢測A375細胞凋亡之型態變化 86 三、大蒜精油對A375細胞內ROS生成的影響 87 四、大蒜精油對A375細胞粒線體膜電位(ΔΨm)的影響 87 五、大蒜精油對A375細胞凋亡相關蛋白質的表現 88 第四節 大蒜精油對人類黑色素細胞癌A375細胞轉移之影響 88 一、大蒜精油對A375細胞移動(Migration)影響 89 二、大蒜精油對A375細胞侵犯力(Invasion)的影響 89 三、大蒜精油對A375細胞分泌MMP-2與MMP-9酵素活性之影響 90 四、大蒜精油對A375細胞分泌MMP-2與MMP-9蛋白質表現之影響 90 第五節 大蒜精油對人類黑色素細胞癌A375細胞增生之影響 90 一、大蒜精油抑制A375細胞固著依賴性細胞群落分析 (Anchorage-dependent colony formation assay) 91 二、大蒜精油對A375細胞增生相關蛋白質的表現 91 第六節 以動物腫瘤模式探討大蒜精油及其活性成分DATS之抑癌活性 92 一、腫瘤動物模式之建立 92 二、大蒜精油及DATS對人類黑色素癌細胞異種移植裸鼠腫瘤生長之影響 92 三、肝功能指數與其他血液生化值 94 第六章 討論 95 第一節 探討大蒜精油抑制人類黑色素細胞癌A375細胞抑癌活性 95 一、大蒜精油對A375細胞生長之抑制效果 95 二、大蒜精油造成人類黑色素細胞癌A375細胞週期停滯 97 第二節 探討大蒜精油誘導人類黑色素癌A375細胞的凋亡 99 一、細胞骨架對人類黑色素癌A375細胞生長之探討 99 二、大蒜精油促進人類黑色素癌A375細胞內ROS的生成而抑制細胞生長 100 三、藉由粒線體途徑誘導人類黑色素癌A375細胞capase dependent apoptosis 101 第三節 探討大蒜精油抑制人類黑色素細胞癌A375細胞的轉移及侵襲 102 第四節 探討大蒜精油抑制人類黑色素細胞癌A375細胞的增生 104 第五節 以體內實驗探討大蒜精油及其有效成分DATS對人類黑色素癌A375細胞株之影響 105 一、大蒜精油及其有效成分DATS在動物實驗的抑癌活性 105 二、大蒜精油及DATS對人類黑色素癌A375細胞異種移植腫瘤生長之影響 106 三、大蒜精油及DATS對異種移植腫瘤血液生化值之影響 108 第七章 結論 109 第八章 實驗圖表 111 第九章 參考文獻 140 | |
dc.language.iso | zh-TW | |
dc.title | 大蒜精油對人類皮膚黑色素細胞癌之抑制活性與機制探討 | zh_TW |
dc.title | Investigation of anticancer activity and mechanism of garlic essential oil against human melanoma cells | en |
dc.type | Thesis | |
dc.date.schoolyear | 99-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 何其儻,羅翊禎,謝淑貞,鍾景光,李宗貴 | |
dc.subject.keyword | 大蒜,黑色素細胞癌,細胞凋亡,細胞轉移,異種移植, | zh_TW |
dc.subject.keyword | garlic,melanoma,apoptosis,melanoma,xenografted nude mice, | en |
dc.relation.page | 173 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2011-08-18 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 食品科技研究所 | zh_TW |
顯示於系所單位: | 食品科技研究所 |
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