薑精油透過抗氧化、抗發炎及調節脂質作用等機制而具改善小鼠非酒精性脂肪肝炎之功效

Abstract

根據台灣肝病防治學術基金會調查，脂肪肝（fatty liver）是台灣國人普遍的肝臟問題，故脂肪肝相關疾病已成為一個日益嚴重的公共衛生問題。非酒精性脂肪肝病（nonalcoholic fatty liver disease, NAFLD）涵蓋了不同程度的脂肪肝病變，包含肝細胞脂肪變性、非酒精性脂肪性肝炎（nonalcoholic steatohepatitis, NASH）、肝纖維化、肝硬化等，甚至可能會惡化成肝癌，且NAFLD與肥胖、血脂異常、糖尿病等代謝疾病具有高度相關性。NAFLD的致病機轉相當複雜，最著名的假說之一為Day與James於1998年所提出之two-hit theory，其first hit是指脂質代謝異常而導致脂肪在肝臟中過度累積；second hit則為氧化壓力及發炎反應導致肝臟進一步發生脂肪肝炎以及纖維化的病變。薑（Zingiber officinale Roscoe）是一種含有刺激性香味的根莖植物，已被廣泛的運用，作為調味品及中藥藥材等，且有研究證實，薑具有調節血脂、抗氧化和抗發炎等活性。本研究所使用的樣品為經水蒸氣蒸餾法所萃取而得的薑精油（ginger essential oil, GEO），根據先前研究中所獲得的結果，本研究的假說為薑精油可藉由其抗發炎、抗氧化和調節血脂作用來改善NASH。因此，本研究目的為應用高脂飼料誘導非酒精性脂肪肝炎之實驗動物模式探討薑精油之護肝功效。根據實驗結果發現，給予12週的高脂飼料可成功誘導小鼠產生NASH，而同時給予GEO及GEO中含量最高之純物質-檸檬醛（citral）後，可使小鼠血清中肝功能生化指標-麩胺酸轉胺酶（aspartate aminotransferase, AST）、丙酮酸轉胺酶（alanine aminotransferase, ALT）之酵素活性、血脂及胰島素降低，亦可使小鼠體重和脂肪組織重量下降及減少肝中三酸甘油酯和膽固醇含量；由肝臟組織病理切片結果得知，樣品之給予可減少小鼠肝臟細胞脂質堆積的情況，此外，還可提升小鼠肝臟中的抗氧化能力和降低發炎因子的表現量；進一步以西方墨點法評估小鼠肝臟中與脂質代謝相關之蛋白質表現量，GEO和citral可顯著抑制小鼠肝臟中固醇調節因子結合蛋白1（sterol regulatory element-binding protein-1c, SREBP-1c)、乙醯輔酶A羧化酶（acetyl-CoA carboxylase, ACC）、脂肪酸合成酶（fatty acid synthase, FAS）及抑制3-Hydroxy-3-methylglutaryl-coenzyme A reductase（HMGCR）之蛋白質表現量，達到減少脂肪酸及膽固醇之生合成作用，進而減少高脂所造成的脂質代謝異常情形，並可藉由抑制被誘導代謝過多游離脂肪酸之細胞色素 P450 2E1（cytochrome P450 2E1, CYP2E1）酵素活性下降，而減少ROS生成所造成的氧化壓力。綜合以上結果可推測，GEO和citral可能具有改善高脂飼料誘導小鼠非酒精性脂肪肝炎之功效，其作用機制可能是藉由降低肝臟中脂質的生合成作用與堆積，並提升肝中抗氧化能力及降低發炎反應而達成。

According to the survey of Taiwan liver research foundation, fatty liver is a common liver disease in Taiwan. Consequently, fatty liver disease has become an increasingly serious public health problem. Non-alcoholic fatty liver disease (NAFLD) represents a wide spectrum of liver pathology ranging from hepatic steatosis to nonalcoholic steatohepatitis (NASH), fibrosis, cirrhosis and liver cancer. In addition, NAFLD exhibit an important relationship with obesity, hyperlipidemia and type II diabetes mellitus. Previously, researches proposed a two-hit theory to explain the progression of NAFLD. The first hit constitutes the accumulation of triglycerides in the hepatocyte due to abnormal lipid metabolism. The second hit promote oxidative stress, inflammation, cell death, and fibrosis. In decades, some specific nutrients or phytochemicals have been demonstrated that provide potential hepatoprotective effects. Ginger (Zingiber officinale Roscoe) is a rhizome plant containing pungent flavor and has been used widely in various foods, beverages, and traditional Chinese medicine. Ginger essential oil (GEO) was obtained by steam distillation of the rhizome of Z. officinale. GEO has been exhibited several biological benefits including anti-inflammatory, anti-oxidative and lipid reducing effects. The hypothesis of this study was improved NASH through anti-inflammation, anti-oxidation and lipid regulation of GEO. Therefore, this study was investigated the hepatoprotective effects of GEO and its active compound in high fat diet-induced NASH mice. In our results, it showed GEO and GEO pure substance – citral can decrease enzyme activity of aspartate aminotransferase (AST), alanine aminotransferase (ALT), lipids and insulin in serum. In addition, GEO and citral reduce body weight, adipose tissue weight, triglycerides and cholesterol level in liver from mice which are induced NASH for 12 weeks. Moreover, the liver biopsy results show the samples can be given to reduce the phenomenon of lipid accumulation in mouse liver cell. Furthermore, GEO and citral can enhance the antioxidant capacities and reduce inflammation response in mouse liver; Further assessment by western blotting associated with lipid metabolism in mouse liver, GEO and citral can inhibit the protein expression of sterol regulatory element-binding protein-1c (SREBP-1c), acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS) and 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) significantly, which make to reduce the biosynthesis of fatty acid and cholesterol. Additionally, they may also decrease the enzyme activities of cytochrome P450 2E1 (CYP2E1) induced by metabolism of excessive free fatty acid, and reducing the oxidative stress by ROS. Based on the above results can be speculated, GEO and citral may exhibit the hepatoprotective effects on nonalcoholic steatohepatitis induced by high-fat diet in mice, and the mechanism are including the reduction of lipid biosynthesis in liver, the elevation of hepatic anti-oxidative capacities and decrease of inflammation response.