臺灣博碩士論文加值系統

安非他命 (amphetamine) 是一種很強的中樞神經興奮劑，在過去的研究方向著重於中樞神經系統以及行為改變；後來研究發現，安非他命對於循環心血管系統、消化系統及生殖系統皆有影響。不論是以口服或是靜脈注射的方式使用安非他命，皆需透過肝臟的分解後運送至全身細胞引起藥物反應，目前已知安非他命誘發多種組織的氧化壓力傷害、發炎反應以及細胞凋亡，但在肝臟組織的相關機制仍不清楚。因此，本研究的主要目的是探討甲基安非他命是否透過MAPK訊息傳遞路徑及抑制抗氧化系統造成肝臟發炎及凋亡。本研究於SD雄鼠背部皮下注射甲基安非他命 (methamphetamine，每天每公斤10毫克甲基安非他命)，並分別於注射15天, 30天, 60天及120天後犧牲，取出肝臟組織，進行西方點墨法 (Western Blot)，分析肝臟組織的mitogen-activated protein kinase (MAPK) 家族、NF-kB、iNOS、eNOS、BCL-2以及BCL-x等蛋白表現量。此外，利用抗氧化套組 (assay kits) 測量肝臟組織glutathione (GSH) 的產量及抗氧化酵素的活性，如superoxide dismutase (SOD)、catalase、glutathione peroxidase (GPx)、glutathione reductase (GR)。組織內脂質過氧化則使用TBARS kit偵測。每次實驗各組至少重複三次，數據以Mean±SEM表示。先以 one-way analysis of variance (one-way ANOVA) 進行資料分析，若呈顯著，則繼以Duncan’s post-hoc 測驗法分析組間的差異，以p＜0.05 視為顯著水準。本研究結果顯示，長期注射甲基安非他命30天，顯著增加 t-AKT表現；注射120天後，顯著抑制p-JNK/t-JNK。控制組隨著年齡的增加，減少肝臟組織的SOD、catalase及GPx的活性。注射15天或30天甲基安非他命 (10 mg/kg/day)，顯著抑制SOD、catalase、GPx活性及GSH產量；並於注射甲基安非他命後120天，與控制組相較，顯著增加catalase及GPx等抗氧化酵素活性。在蛋白表現方面，控制組仍然隨著年齡週數的延長而減少eNOS、Cu/Zn-SOD、Mn-SOD、catalase、BCL-2以及BCL-x的蛋白表現。注射甲基安非他命15天後，顯著抑制eNOS、Cu/Zn-SDO、Mn-SOD、catalase、BCL-2以及BCL-x，但增加iNOS及NF-kB蛋白表現。但是注射甲基安非他命30天、60天或120天後，上述各種蛋白的表現與控制組相較無差異。本研究的結論是，注射甲基安非他命透過抑制抗氧化壓力系統的活性，造成發炎反應，並促進細胞凋亡產生。

Amphetamine is a strong central neuronal psycho-stimulant, resulted psychopathology, neurotoxicity, hepatotoxicity, and multiple organ degeneration. The amphetamine involved mechanisms included raised oxidant stress, inflammation and apoptosis. To understand wherether amphetamine enhanced the oxidative stress and inflammatory response through mitogen-activated protein kinase (MAPK) pathway, the SD male rats were injected subcutaneously with methamphetamine (10 mg/ml/kg BW/day) or PBS (1 ml/kg BW/day). After daily continuous injection for 15, 30, 60 and 120 days, the rats were scarified, and the livers were collected. The expression of MAPK family, NF-kB, iNOS, eNOS, BCL-2 and BCL-x were performed by Western Blot. Besides, the activity of superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), and glutathione reductase (GR) were measured by using commercial kits. Glutathione (GSH) assay kit and TBARS kit were used to analyze production of GSH and lipid peroxide. All values were presented as the mean ± standard error of the mean (SEM). The different between the treatment were tested by one-way analysis of variance (one-way ANOVA), and the difference between specific means were tested for significance by Duncan’s multiple range test. A difference between two means will be considered statistically significant when p< 0.05. The expression of t-AKT significantly (p<0.01) increased after methamphetamine injction for 30 days, and ratio of p-JNK/t-JNK significantly (p<0.05) decreased after methamphetamine injection for 120 days. In PBS groups, activities of SOD, catalase and GPx decreased following age increased. The activities of SOD, catalase, GPx and the production of GSH significantly decreased significantly (p<0.05) by injection methamphetamine (10mg/kg/per day) for 15 or 30 days. After administration of methamphetamine for 120 days, the activities of catalase and GPx were significantly higher than respective PBS group. As following the age-increased, protein expression of eNOS, Cu/Zn-SOD, Mn-SOD, catalase, BCL-2 and BCL-x decreased in PBS groups. Moreover, after injection of methamphetamine for 15 days, significant inhibited protein expression of eNOS, Cu/Zn-SOD, Mn-SOD, catalase, BCL-2 and BCL-x, but increased expression of iNOS and NF-kB. However, after injection of methamphetamine for 30, 60 and 120 days, all the protein expression were not different from the respective PBS groups. The conclusion is methamphetamine induced inflammation, and apoptosis through inhibited the activities of anti-oxidant system.

目錄
論文考試審定書 ii
授權書 iii
名詞縮寫 iv
中文摘要 v
英文摘要 vii
謝誌 ix
目錄 x
圖目錄及表目錄 xii
第一章、緒論 1
一、研究背景 1
二、研究目的 1
三、研究假設 1
第二章、文獻探討 2
一、安非他命的歷史發展 2
二、安非他命對生理功能的影響 4
三、安非他命與氧化壓力及一氧化氮的關係 6
四、安非他命與MAPK訊息傳遞路徑 10
第三章、研究方法與研究步驟 12
一、動物的選取與照料 12
二、實驗設計 12
三、實驗流程 13
四、實驗材料與分析 14
五、資料處理及統計分析 25
第四章、結果 26
一、甲基安非他命對大鼠體重、肝臟組織重量及肝臟體重比值的影
響 26
二、甲基安非他命對肝臟組織抗氧化系統及脂質過氧化的影響 26
三、甲基安非他命對肝臟組織特定蛋白表現量的影響 27
第五章、討論 32
參考文獻 37
圖及表 44

圖目錄及表目錄
圖1-1.安非他命與甲基安非他命結構圖 44
圖1-2.連續注射甲基安非他命120天對雄性SD大鼠體重的影響 45
圖1-3.注射甲基安非他命15, 30, 60, 120天對雄性SD大鼠體重的影響
46
圖1-4.連續注射甲基安非他命30或120天對雄性SD大鼠肝臟重量的
影響 47
圖1-5.注射甲基安非他命30及120天對雄性SD大鼠肝臟體重比值的
影響 48
圖1-6.連續注射甲基安非他命對大鼠肝臟SOD活性的影響 49
圖1-7.連續注射甲基安非他命對大鼠肝臟catalase活性的影響 50
圖1-8.連續注射甲基安非他命對大鼠肝臟GPx活性的影響 51
圖1-9.連續注射甲基安非他命對大鼠肝臟GR活性的影響 52
圖1-10.連續注射甲基安非他命對大鼠肝臟GSH產量的影響 53
圖1-11.連續注射甲基安非他命對大鼠肝臟TBARS濃度的影響 54
圖1-12.連續注射甲基安非他命對大鼠肝臟eNOS蛋白的影響 55
圖1-13.連續注射甲基安非他命對大鼠肝臟iNOS蛋白的影響 56
圖1-14.連續注射甲基安非他命對大鼠肝臟Cu/Zn-SOD蛋白的影響
57
圖1-15.連續注射甲基安非他命對大鼠肝臟Mn-SOD蛋白的影響
58
圖1-16.連續注射甲基安非他命對大鼠肝臟catalase蛋白的影響 59
圖1-17.連續注射甲基安非他命對大鼠肝臟AKT蛋白的影響 60
圖1-18.連續注射甲基安非他命對大鼠肝臟JNK蛋白的影響 61
圖1-19.連續注射甲基安非他命對大鼠肝臟ERK蛋白的影響 62
圖1-20.連續注射甲基安非他命對大鼠肝臟NF-kB蛋白的影響 63
圖1-21.連續注射甲基安非他命對大鼠肝臟BCL-2蛋白的影響 64
圖1-22.連續注射甲基安非他命對大鼠肝臟BCL-x蛋白的影響 65
表2-1、連續注射甲基安非他命對大鼠肝臟特定蛋白表現的影響
66

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郭家驊、劉昉青、祁業榮、劉針芳、張振崗、邱麗玲、郭婕 合著(民90)。運動營養學 (初版)。台中市：華格那。
Tortora簡明人體解剖學與生理學 (陳金山、徐淑媛譯) (民96)。臺北市：合記。(原著出版年：2007)。
運動生理學 (林正常 等譯) (民91)。台北市：麥格羅希爾。(原著出版年：2002)。
行政院管制藥品管理局青少年藥物濫用調查 (九十二年臺灣地區藥物濫用趨勢)，2004，http://www.pcc.vghtpe.gov.tw/k-n-08.asp