中药五味子木脂素类成分抗肝损伤、抗肝纤维化作用评价与机理研究

Schisandrae Chinensis has the role of anti-liver injury and anti-liver fibrosis, and the Lignans are the main active ingredients in Schisandra Chinensis. The successful case of new drugs (Bifendate, and bicyclic alcohols) whose structures were modified from Schizandrae C reveals that it is feasibile to look for the drugs to treat liver disease from the Schisandrae Chinensis, but its specific mechanism of action is not clear. Cannabinoid G protein-coupled receptors, CB1 and CB2, are the the new action targets with anti-liver fibrosis action. And it is indicated that lignans in Schisandra can excite anti-liver fibrosis actions by inhibiting the CB1 receptor and activate CB2 receptors through our previous research. The project intends to study the lignans in Schisandrae Chinensis, build the classic liver injury and liver fibrosis animal model, and take a comprehensive pharmacological evaluation of the lignans in Schisandrae Chinensis in vitro and in vivo. Cell biology assays and radioligand binding assay were used to find the great potent ingredients preliminarily, combined with 2D/3D NMR and 3D virtual structure screening technology. According to the regulation of CB1 and CB2 receptors, the lignans of anti-liver injury and liver fibrosis actions can be focused, and the mechanism of anti-liver fibrosis can be clarified. This is very significant to improve the clinical applications of Schisandrae Chinensis, find new drug lead compounds, which has the effect of anti-liver injury and anti-liver fibrosis, and modify the structures of lead compounds.

五味子具有抗肝损伤和抗肝纤维化作用，木脂素类成分是其主要有效成分，经五味子丙素结构改造的新药发现成功案例（联苯双酯和双环醇）启示我们，从五味子木脂素类成分中寻找治疗肝病的药物可行性强，但目前其作用机理不清楚。大麻素G蛋白偶联受体CB1和CB2是最新发现的抗肝纤维化作用的靶点，前期的研究结果表明，五味子中的五味子醇甲可以通过拮抗CB1受体和激动CB2受体而起抗肝纤维化的作用。本项目拟以五味子木脂素类成分为研究对象，以CB1/CB2受体为抗肝纤维化的作用靶点，通过构建经典的肝损伤和肝纤维化动物模型，采用2D/3D NMR和3D虚拟结构筛选技术，结合体内外药效综合评价、细胞生物学测定和放射结合测定方法，定位作用较强的木脂素类成分,进而阐述木脂素类成分抗肝损伤、抗肝纤维化的作用机理，这对于提高五味子的临床应用和发现抗肝损伤、抗肝纤维化作用的新药先导化合物并进行结构改造具有重要意义。

肝损伤和肝纤维化都属于慢性肝脏疾病，是发展为肝硬化甚至肝癌的必经过程。对于肝纤维化，目前还没有有效的治疗药物，这是目前面临的主要问题。五味子木脂素类成分是中药五味子中的主要活性成分，具有显著的保肝降酶作用。本研究通过构建小鼠肝损伤和大鼠肝纤维化模型，对五味子木脂素类成分提取物（SLE）和五味子乙素进行抗肝损伤和抗肝纤维化作用评价，进而探讨其体内信号通路和分子机制,寻找可能的治疗靶点。结果表明SLE具有较好的抗肝损伤作用，其具体的作用机制为：SLE通过降低MDA含量和提高SOD，GSH和GSH-Px活力来发挥抗氧化应激作用；通过抑制NF-κB活力，降低炎症因子TNF-a, IL-1β和 IL-6水平，起到抗炎作用；通过抑制JNK活化及调节Bcl-2/Bax信号通路起到抗肝细胞凋亡的作用。综合以上研究，五味子木脂素类成分提取物是通过综合发挥抗氧化，抗炎和抗凋亡共同发挥保肝作用的。五味子乙素给药肝纤维化模型大鼠后，明显减轻了其肝纤维化症状，其抗肝纤维化的内在机制为：五味子乙素虽然不能抑制肝星状细胞的增殖，但可以通过抑制 TGF-β刺激α-SMA和Collagen-I的增加来发挥抗肝星状细胞活化的作用；此外，五味子乙素还能够抑制Smad非依赖的MAPK通路。不仅抑制了TGF-b刺激的Smad通路，还抑制了Smad非依赖的MAPK通路，提示我们五味子乙素有可能抑制了Smad和MAPK通路而抑制了肝星状细胞活化进而发挥抗肝纤维化的作用。通过构建肝纤维化靶点数据库，通过导入智能分析平台，进行在线成分-靶点对接，结合体外成分-蛋白竞争性结合数据，表明五味子乙素和丙素与CB2受体具有较好的结合，很有可能是其发挥抗肝纤维化作用的靶点，这为寻找木脂素类成分抗肝纤维化作用靶点奠定了基础。本项目已申请国家发明专利2项，发表SCI论文1篇和核心期刊论文1篇，投稿SCI论文1篇。培养1名硕士研究生。