藏药七味铁屑方对大鼠非酒精性脂肪肝的干预作用及机制研究

Non-alcoholic fatter liver disease(NAFLD) is a clinical syndrome with lesion of steatosis and fat storage in hepatic lobules but without alcohol abuse.The pathological features are fatty degeneration of hepatocytes, injury and inflammatory cells infiltration.And its path-physiological hallmark is insulin resistance and oxidative stress. The prevalence of NAFLD increases year by year, NAFLD has been one of the most common causes of chronic 1iver diseas and abnormal liver function globally. Howerver，for a long time，studies have failed to clearly define the molecular theory and the optimal treatment has not been established domestic. Qiwei Tiexie Fang ， the classic complex prescription of Tibetan drug，is based on the three-cause theory.The major functions of Qiwei Tiexie Fang is promoting flow of qi and blood circulation，calming the liver，clearing heat， relieving pain. Application of various acute or chronic liver disease for a long time, it has a good effect on fatty liver disease prevention and control. The mechanism of Qiwei Tiexie Fang in the teatment NAFLD is unc1ear and needs furthur study. In this study， the effect of Qiwei Tiexie Fang on glucose uptake and fatty acid oxidation and inflammatory reaction was measured in the 3T3-L1 adipocyte and the hepatic tissue of NAFLD rats. Furthermore，important factors involved in glucose and lipid metabolism and inflammatory reaction such as PPARs、LXRα、CPT-l、IκBα、 iNOS、 Akt、 P-Akt 、MCP-1 etc. were detected to explore the mechanism of the therapy effect .

非酒精性脂肪肝（NAFLD）是以肝细胞脂肪变性、损伤及炎症细胞浸润为病理特征，但无过量饮酒史的临床综合征，主要病理生理机制为胰岛素抵抗和氧化应激，发病率呈逐年上升趋势，成为全球引起慢性肝病、肝功能异常的首要原因之一。但其发病机制尚未完全阐明，缺乏有效的药物治疗。七味铁屑方是根据"三因"理论形成的藏药经典复方，主治行气活血，平肝清热止痛，长期应用各种急慢性肝病，对脂肪肝防治疗效显著，但其发挥作用的具体机制尚不明确。本研究以3T3-L1脂肪细胞、NAFLD大鼠模型肝脏为研究对象，通过七味铁屑方的干预，检测对糖脂代谢、氧化代谢及炎症反应相关指标的影响以及发挥重要作用的PPARs、LXRα、CPT-l、 IκBα、iNOS、Akt、P-Akt 、MCP-1等表达变化，以探讨其作用机制。

七味铁屑胶囊(QWTX)是藏药的代表方剂，广泛用于慢性肝病和非酒精性脂肪肝(NAFLD)的长期治疗。本研究探讨QWTX对3T3-L1脂肪细胞和NAFLD的作用及机制。本研究采用3T3-L1前脂肪细胞和NAFLD大鼠模型。在3T3-L1细胞中，用CKK-8检测QWTX的细胞毒性，2-deoxy-D-[3H]葡萄糖和[1-14C]棕榈酸分别测定葡萄糖摄取和脂肪酸氧化。抗肉毒碱棕榈酰基转移酶-1抗体(CPT-1),肝X受体α(LXRα)、过氧化物酶体增殖物激活受体-γ(PPAR)γ,肝X受体α(LXRα)和蛋白激酶B的表达水平是由PCR和免疫印迹决定。采用脂肪乳剂加蔗糖共9周建立NAFLD。采用HE染色和透射电镜观察QWTX对NAFLD大鼠脂质代谢、肝功能和肝脏形态的影响。血清一氧化氮(NO)和游离脂肪酸(FFA) 水平、超氧化物歧化酶(SOD)和丙二醛(MDA) 在肝脏中的含量,以及单核细胞趋化蛋白-1 (MCP-1) CPT-1, LXRα,PPARα,PPARβ/δ,PPARγ, iNOS的表达水平都检测到。QWTX对3T3-L1前脂肪细胞无细胞毒性，14CO2的生成速率增加到4.15，表明脂肪积累减少。在NAFLD中，通过HE染色和电镜检查QWTX可减轻肝脏脂肪变性、脂肪空泡和炎症。QWTX治疗后氧化应激生物标志物血清FFA水平降低，血清NO水平升高。在肝脏组织中，NAFLD中SOD降低，MDA明显升高，均通过QWTX恢复。在非酒精性脂肪肝NF-κB和CYP2E1也调节,而由QWTX使其表达下调。在QWTX作用于3T3-L1脂肪细胞和非酒精性脂肪肝模型中检测到LXRα,PPARγ和iNOS向下调节。