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| 題名: | 油脂對非酒精性脂肪肝疾病大鼠及其粒線體功能之影響
Effects of oils on nonalcoholic fatty liver disease in rats and its mitochondrial function |
| 作者: | 吳宜倩
Wu Yi-Chien |
| 貢獻者: | 保健營養學研究所
謝榮鴻 |
| 關鍵詞: | 碩士
指導教授-謝榮鴻
委員-葉添順
委員-施純明
非酒精性脂肪肝疾病;粒線體;魚油;橄欖油;茶油
nonalcoholic fatty liver disease;mitochondria;fish oil;olive oil;tea seed oil |
| 日期: | 2010 |
| 上傳時間: | 2010-10-20 12:46:54 (UTC+8) |
| 摘要: | 非酒精性脂肪肝疾病(nonalcoholic fatty liver disease, NAFLD)是由非大量攝取酒精所引起的肝臟相關疾病，全世界罹患NAFLD的人口約有10%至24%，其病程包含單純性脂肪肝、非酒精性脂肪肝炎 (nonalcoholic steatohepatitis, NASH)、肝纖維化、肝硬化、肝衰竭及肝癌等，其中又以單純性脂肪肝及NASH盛行率最高。目前由單純性脂肪肝發展成NASH的機制尚不明確，"二次傷害假說"為目前最廣泛被接受的原因，此兩次傷害分別為胰島素抗性及氧化壓力上升。粒線體為脂肪酸進行β氧化及產生自由基的主要場所，近年研究發現，NAFLD之發生及進展與粒線體功能異常有密切關係。過去研究亦發現不同種類的脂肪酸可影響粒線體功能及其β氧化速率，進而調控肝臟中脂質代謝。
本次研究的目的為給予具有脂肪肝之大鼠不同種類油脂的飲食，並觀察魚油、橄欖油及茶油等是否會影響粒線體功能，並達到改善NAFLD病程之進展。本次實驗使用八週齡大雄性Sprague Dawley (SD)大鼠，分為控制組及高脂飲食組。高脂飲食組藉由給予脂質占總熱量71%，且富含豬油之飲食14週以建立單純性脂肪肝之動物模式。確認高脂飲食組大鼠發生脂肪肝後，再隨機分為魚油組、橄欖油組、茶油組、低脂豬油組及高脂豬油組，介入6週後予以犧牲。
結果顯示低脂豬油組、茶油組、橄欖油組及魚油組其肝臟三酸甘油酯含量及胰島素抗性皆較高脂豬油組低 (P＜0.05)，而又以魚油組改善情況最佳。高脂豬油組之粒線體complex I及 II的呼吸控制比值 (Respiratory control ratio, RCR)顯著低於控制組 (P＜0.05)，而低脂豬油組、茶油組、橄欖油組及魚油組較高脂豬油組高 (P＜0.05)，其中又以茶油組回升效果最佳，橄欖油次之。粒線體電子傳遞鏈複合體酵素活性方面，高脂豬油組之cytochrome c oxidase (CCO)顯著低於控制組 (P＜0.05)，魚油組、橄欖油組、茶油組之CCO則較高脂豬油組顯著上升。高脂豬油組之肝臟粒線體數量較控制組少，且膜有破裂的現象，而低脂豬油組、茶油組、橄欖油組及魚油組其粒線體膜較完整且數量較高脂豬油組多。由結果顯示經由高脂飲食引發脂肪肝之大鼠其粒線體酵素、呼吸速率及其控制比值及粒線體結構皆有異常的現象，而攝取較低比例的油脂即可改善脂肪肝大鼠之肝臟TG堆積情形及其粒線體功能，但隨著不同油脂之介入影響亦不相同，魚油對胰島素抗性及脂質堆積改善效果最佳，茶油及橄欖油則對改善粒線體功能較好。
Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease with worldwide prevalence that occurs in subjects who do not abuse alcohol. It is defined as a spectrum of hepatic steatosis, nonalcoholic steatohepatitis (NASH), fibrosis, necrosis and cirrhosis. The prevalence is about 10 to 24% in general population, and the mortality is up to 13%. Simple fatty liver and NASH are the most common characteristics in NAFLD, but the pathogenesis of NASH is not well defined. ”Two-hit hypothesis” have been proposed for the pathophysiology of NAFLD and NASH. Insulin resistance and oxidative stress are the first and second hits, respectively. Mitochondrion is an important organelle confers beta-oxidation of fatty acids and also the major site to produce by-product of oxygen reactive species. Mitochondrial dysfunction might play an important role in both hits. It is reported previously, there are distinct effects on mitochondrial function by treating different kinds of fatty acids.
The objects of this study were to investigate whether oils, such as fish oil, olive oil, tea seed oil and lard, could influence mitochondrial function and improve fatty liver disease. Male Sprague Dawley (SD) rats were fed with 71% high fat diet to induce simple fatty liver, afterward, randomly divided into 5 groups: high-lard (HL), low-lard (LL), tea seed oil (TO), olive oil (OO) and fish oil (FO) diets.
Liver triglyceride content and HOMA-IR index in LL, TO, OO, FO groups were lower than HL group (P＜0.05), moreover, FO group was significantly reduced in body weight and visceral fat. Respiratory control rate (RCR) in HL group was lower than control group (P＜0.05), and increased in other groups, especially in TO group. The activity of cytochrome c oxidase (CCO) in HL group was lower than control group (P＜0.05), besides, CCO was increased significantly in TO, OO and FO group compared with HL groups (P＜0.05). In conclusion, all of fish oil, olive oil and tea seed oil might provide benefits for fatty liver disease, but they should via diverse mechanisms to affect the progression of NAFLD. |
| 關聯: | 77頁 |
| 描述: | 目錄
中文摘要 I
Abstract III
致謝 V
縮寫表 VII
藥品試劑列表 IX
目錄 XI
表目錄 XV
圖目錄 XVII
第一章 序論 1
第二章 文獻回顧 3
第一節 非酒精性脂肪肝疾病 (nonalcoholic fatty liver disease, NAFLD) 3
第二節 粒線體與自由基 5
第三節 粒線體與脂肪酸β氧化 6
第四節 粒線體與非酒精性脂肪肝炎 7
第五節 脂肪酸與非酒精性脂肪肝疾病 8
第六節 茶油 (茶籽油) 10
第三章 研究目的 11
第四章 材料與方法 12
第一節 實驗動物分組 12
第二節 飼料組成 13
第三節 實驗流程 14
第四節 油脂之脂肪酸成分分析 15
(一) 脂肪酸萃取 15
(二) 脂肪酸成分分析 15
第五節 血液樣品分析 16
(一) 血液樣品收集 16
(二) 分析項目與方法 16
第六節 肝臟樣品分析 19
(一) 肝臟樣品收集 19
(二) 分析項目與方法 20
第七節 粒線體功能分析 21
(一) 肝臟粒線體電子顯微鏡切片 21
(二) 萃取肝臟粒線體 21
(三) 萃取肝臟total DNA 22
(四) 萃取肝臟總蛋白質 22
(四) 分析項目與方法 23
第八節 統計分析 27
第五章 結果 28
第一節 大豆油、豬油、茶油、橄欖油及魚油之脂肪酸成份百分比組成 28
第二節 脂肪肝動物模式之建立 28
第三節 攝食量及體重 29
第四節 血糖、血漿胰島素及HOMA-IR 30
第五節 肝臟三酸甘油酯含量、組織病理切片及腹部脂肪 31
第六節 血漿脂質濃度及肝功能指數 32
第七節 肝臟脂質過氧化產物 33
第八節 肝臟電子顯微鏡切片 34
第九節 粒線體呼吸傳遞鏈複合體酵素活性 34
第十節 粒線體呼吸速率及呼吸控制比值 (RCR) 35
第十一節 肝臟粒線體生合成 36
第六章 討論 37
第一節 富含豬油之71%高脂飲食對於誘發脂肪肝動物模式之影響 37
第二節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠其血糖、血漿胰島素及HOMA-IR之影響 37
第三節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟脂質代謝之影響 39
第四節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠血漿脂質代謝之影響 40
第五節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟氧化壓力之影響 40
第六節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟粒線體結構之影響 41
第七節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟粒線體呼吸傳遞鏈複合體酵素活性之影響 42
第八節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟粒線體呼吸速率及呼吸控制比值 (RCR)之影響 43
第九節 豬油、茶油、橄欖油及魚油對於脂肪肝大鼠肝臟粒線體生合成之影響 44
第七章 總結 45
第八章 參考文獻 46
表目錄
表一. 控制組及高脂飲食組之飼料組成分 51
表二. 魚油組、茶油組、橄欖油組及低脂豬油組飲食之飼料組成 52
表三. 本次實驗所使用之油脂脂肪酸成分百分比 (%) 53
表四. 即時聚合酶連鎖反應分析使用之引子對序列 54
表五 西方點墨法分析使用之一級抗體 55
表六：於實驗期第0、14 及20週各組之體重。 56
表七 於實驗期第0至14 週及14至20週之平均攝食量 57
表八. 實驗期第0、12、14、20週之禁食血漿葡萄糖濃度 58
表九. 實驗期第14、20週之血漿胰島素濃度 59
表十. 實驗期第14、20週之HOMA-IR值 60
表十一. 實驗期第14、20週之肝臟三酸甘油酯含量 61
表十三. 實驗期第14、20週之腹部脂肪重量 62
表十三. 實驗期第0、14、20週之血漿三酸甘油酯含量 63
表十四. 實驗期第0、14、20週之血漿總膽固醇含量 64
表十五. 實驗期第0、14、20週之血漿HDL-c含量 65
表十六. 實驗期第0、14、20週之血漿LDL-c含量 66
表十七. 實驗期第0、14、20週之血漿AST含量 67
表十八. 實驗期第0、14、20週之血漿ALT含量 68
表十九. 實驗期第20週粒線體呼吸傳遞鏈複合體酵素活性 69
表二十. 實驗期第20週肝臟粒線體呼吸速率及控制比值 70
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