具有治療非酒精性脂肪肝之中草藥之篩選

Identification of Chinese herbal medicines for treatment of nonalcoholic fatty liver disease

賴昱昇(Yu-Sheng Lai)

指導教授 ： 呂紹俊

國立臺灣大學/醫學院/生物化學暨分子生物學研究所/碩士(2009年)

https://doi.org/10.6342/NTU.2009.10356

摘要

非酒精性脂肪肝，定義為非酗酒過量者之肝細胞脂肪重量達到肝總重5%以上。致病原因可歸納於脂肪酸合成增加及 (或) 脂肪酸分解減低。近年來國內成人人口脂肪肝盛行率高達26-34%。臨床研究顯示非酒精性脂肪肝與肥胖、糖尿病及代謝症候群相關疾病有高度正相關。此外，近年研究更顯示，非酒精性脂肪肝有相當比例發展成肝纖維化、肝硬化甚至是肝癌。然而目前臨床上並無令人滿意的藥物，因此使得非酒精性脂肪乾的治療藥物開發有相當的空間。
我們從中醫古籍上記載具有保肝功能之中藥篩選可降低細胞三酸甘油酯的中草藥，由18種複方即60種單方中草藥中，我們篩選出可降低細胞三酸甘油酯的5種複方及18種單方，並探討其用於治療非酒精性脂肪肝的可能性。
首先我們以LXR agonist—T0901317存在與否發現有2種複方可降低HepG2細胞中三酸甘油酯含量。之後以西方點墨法發現22種單方中有13種單方降低細胞中FASN蛋白含量，顯示這些藥材具有抑制脂肪酸合成的功效。接著以intralipid直接提供脂肪酸來源並抑制脂肪酸合成，發現有3種單方仍可降低肝細胞TG顯示可能具促進脂肪酸氧化分解的作用。經由中醫師之建議，將上述篩選出可抑制脂肪酸合成與促進分解之中草藥單方組成兩個配方：A配方及B配方。
之後我們以十一週大C57BL/6雌性小鼠為動物模式，將之分為四組，分別為控制組、Lipogenic diet組 (35% corn　starch + 35% sucrose)、Lipogenic diet + A配方組與Lipogenic diet + B配方組。飼養8週後，灌食B配方可顯著降低肝臟總膽固醇含量，也減低腹腔脂肪重量，肝組織切片亦發現脂肪空泡堆積明顯減少之情形。然而各組間血液AST與ALT活性並無顯著差異，但由RT-qPCR分析發炎反應基因，發現比起控制組餵食Lipogenic diet顯著增加MCP-1、F4/80及TNF-α表現，而灌食B配方可顯著抑制餵食Lipogenic diet引發MCP-1、F4/80及TNF-α之mRNA表現。另外， Lipogenic diet組織SREBP-1 mRNA表現顯著高於控制組，但灌食B配方可顯著降低其表現；而CPT-1a與ACO在餵食Lipogenic diet後表現顯著低於控制組，灌食A配方及B配方皆可提升其表現量。顯示此動物模式下，B配方可影響全身性脂肪之代謝同時抑制肝臟發炎現象，然而A配方雖可改善肝臟脂肪堆積但未抑制發炎反應。
動物實驗結果發現B配方具有改善肝臟脂肪堆積與降低發炎指標之功效，然而B配方是否在其他動物模式仍然有效還有待後續實驗分析，因此如何以不同動物模式印證B配方之效果，找出B配方之有效成分及其作用機轉為日後後續研究之重點。

關鍵字

非酒精性脂肪肝 ； 三酸甘油酯 ； 肝發炎反應 ； 中草藥

並列摘要

Non-alcoholic fatty liver disease (NAFLD) is a disease with over 5% fat (w/w) in the liver of people who consumed limited amount of alcohol. The pathology of NAFLD can be resulted from the increase of fatty acid synthesis and/or decrease of fatty acid β-oxidation. The prevalence of NAFLD in Taiwan is about 26-34%. Clinical and epidemiological studies revealed that NAFLD is associated with metabolic syndrome, such as insulin resistance and obesity. Furthermore, NAFLD is predicted as a risk factor of liver fibrosis, cirrhosis and cancer. Still, there is no satisfying medicines to treat NAFLD. Thus, it makes developing of medicines for treatment of NAFLD more important.
We screened a total of 60 Chinese Herbal Medicines (CHMs) in HepG2 for their effects on cellular triacylglycerol (TG). Five CHM prescriptions were identified, which could decrease cellular TG in the presence or absence of LXR agonist – T0901317. Twenty-two CHMs were identified could decrease cellular TG for at least 20%. Western blot and RT-PCR showed that 13 of these CHMs could inhibited fatty acid synthase (FASN) expression and three of them could increase fatty acid oxidation. Two mixtures, mixture A and B, that contained CHMs could inhibit FASN, and the CHMs which could induce FA oxidation were prepared according to the suggestion of doctors of traditional Chinese medicine.
An animal experiment was carried out using 11-week old C57BL/6 female mice. Mice were divided into four groups: control, lipogenic diet, lipogenic diet + mixture A and lipogenic diet + mixture B, mixtures A and B were given by gavage. After 8 weeks of treatment, mixture B significantly decreased hepatic content of total cholesterol and the weight of abdominal fat. Histological analysis showed that mixture B improved liver steatosis signifcantly. No difference was found in the plasma AST and ALT activities among four groups. However, RT-Q-PCR analysis revealed that feeding lipogenic diet induce increased mRNA levels of macrophage infiltration and hepatic inflammatory cytokines, such as MCP-1, F4/80 and TNF-

並列關鍵字

NAFLD ； triglyceride ； hepatic inflammation ； Chinese Herbal Medicine

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