臺灣博碩士論文加值系統

酒精性脂肪肝主要是由長期飲酒所造成的肝臟脂肪堆積病症，當脂肪進入人體會刺激脂肪生合成相關路徑並造成氧化壓力並加速脂肪堆積。甚且，若脂肪肝不及時給予治療會更加惡化造成後續肝硬化、肝癌等病症。因此，尋找天然以及具有潛力的中草藥物來治療酒精性脂肪已是重要課題。
綬草 (Spiranthes sinensis)是個尚未被開發及了解的中草藥物。本研究透過綬草萃取物用於酒精及油酸誘導肝細胞組織FL83B，並透過紅油染色及及時聚合酶鏈式反應分析，發現綬草萃取物具有改變油滴堆積之功效並且抑制脂肪生合成之相關基因Sterol regulatory element-binding proteins 1C (SREBP-1C)及Acetyl-CoA carboxylases 1 (ACC1)表現；其次利用酒精誘導之酒精性脂肪肝小鼠給予綬草萃取物後，在血液生化值及肝臟脂肪含量有具有減緩功效，並抑制脂肪生合成之相關蛋白Fatty acid synthesis (FASN)及ACC1，且增加脂肪分解相關蛋白Carnitine palmitoyltransferase 1B (CPT1B)、Peroxisome proliferator-activated receptor alpha (PPARα)以及5’ AMP-activated protein kinase (AMPK)。在肝臟切片中也可看出綬草萃取物可減緩脂肪堆積。總結，綬草不僅在酒精及油酸誘導細胞平台以及酒精誘導動物平台皆具有減脂功效，因此，綬草對在酒精性脂肪肝具有保護肝臟作用的藥物。

Alcoholic fatty liver disease (AFLD), mainly caused by long-term alcohol consumption and led to the accumulation of fat in liver tissues. Fat can cause oxidative stress and triggers the signal pathway and subsequently accelerates lipid droplet accumulation via the activation of adipogenesis in liver. Furthermore, it will get worse to cirrhosis or hepatoma when AFLD is not impeded shortly. Therefore, the exploration of potential candidate from natural sources for alleviation of AFLD renders interesting issue. Spiranthes sinensis, a folk herb, is still unexplored in phytochemical properties for medicinal value. This study was conducted to investigate the biological activity of S. sinensis extract for applying in FL83B cells in vitro by oil red o staining to measure lipid droplet and the results showed that S. sinesis extract could reduce lipid droplets in alcohol plus oleic acid-induced FL83B cells. S. sinensis extract also significantly reduced lipogensis SREBP-1C and ACC1 mRNA levels by qPCR. Moreover, the alcohol-induced fatty liver C75BL/6 mice was treated with S. sinensis extract to determine the efficacy of alcoholic-fatty liver via lipotropic factors using qPCR and micrographic analysis by H & E staining of liver tissues. The results revealed that the reduction of ACC1 and FASN with the increase of CPT1B, PPARα, and AMPKα with S. sinensis extract treatment resulted in alleviation of fatty liver via Western blot and qPCR. Histopathological review, SS extract slightly reduced lipid droplets in liver of alcohol-induced mice. Taken together, S. sinesis extract possesses lipotropic effect on the alcohol plus oleic-induced FL83B cells and the alcohol-induced mice as well, suggesting S. sinensis extract is evident as a candidate of liver-protection source particularly alcoholic fatty liver.

摘要 I
Abstract V
Contents VII
Table contents XI
Figure contents XIII
Abbreviations XV
Introduction 1
1. Epidemiology of FLD 1
2. Definition and classification of FLD 1
3. Causes of FLD 2
4. Steatosis 3
5. Reducing equivalents (NADH) 3
6. Role of sterol regulatory element-binding proteins 4
7. Role of AMPK 5
8. Role of mammalian sirtuin-1 5
9. Role of peroxisome proliferator-activated receptor-alpha (PPAR-α) 6
10. Treatment of ALDH 7
11. Spiranthes sinensis 7
12. Motivation and aim 8
Material and methods 11
1. Plant material and crude extract 11
2. Cell culture 11
3. MTT assay 11
4. Oil red O staining 12
5. Animal experiments 12
6. Fatty liver induced and S. sinensis treatment 13
7. Blood for biochemical analysis 13
8. Fat content of liver measurement 14
9. Western Blot 14
10. RNA isolation and RT-PCR 15
11. Real-time PCR 15
12. Tissue histopathology 16
13. Statistical analysis 16
Results 17
1. Cytotoxicity of S. sinensis crude extract and taurine in FL83B cells. 17
2. Lipid accumulation on FL83B cells after treatments with S. sinensis extract and taurine. 17
3. The altered genes expression levels of S. sinensis crude extract and taurine on FL83B cells. 18
4. Blood biochemical levels (ALT, AST, cholesterol, triglyceride, HDL, and LDL) in fatty liver mice. 18
5. The appearance of fatty liver in mice. 19
6. Body weight and fat content of liver in mice. 19
7. The altered genes expression levels of S. sinensis extract and taurine in alcoholic fatty liver mice. 20
8. The altered AMPKα protein levels of S. sinensis crude extract and taurine in alcoholic fatty liver mice. 21
9. The altered FASN protein levels of S. sinensis crude extract and taurine in alcoholic fatty liver mice. 22
10. The altered ACC1 and p-ACC1 protein levels of Spiranthes sinensis crude extract and taurine in alcoholic fatty liver mice. 22
11. The altered PPARα protein levels of Spiranthes sinensis crude extract and taurine in alcoholic fatty liver mice. 23
12. Histopathological changes in tissue sections. 23
Discussion 25
Conclusions 28
Future work 29
References 31

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