基于抗补体活性研究青蒿治疗狼疮肾炎的物质基础与作用机制

Lupus nephritis is the most serious complication of systemic lupus erythematosus, and there are no effective drugs to cure it up to date. The unwanted activation of the complement system plays an important role in the pathogenesis of Lupus nephritis. The Artemisia annua has a good effect on the treatment of the Lupus nephritis. Our preliminary experiment data showed that the Artemisia annua had the anticomplement activity through the classic pathway，but the mechanism remains unkown. So, we hyphothesize that the Artemisia annua treats the lupus nephritis through its inhibition of the complement system. Based on the above hypothesis, chemical investigations will be performed guided by the anticomplement activity in vitro and compounds will be elucidated by comprehensive NMR spectroscopy and MS. On this basis, the anticomplement compounds will be analyzed by HPLC/MS and enriched by pre-HPLC, and combined into a new effective composition. Then, we study the effect of the effective compositon on the differentiation of B lymphocyte activated by C3b and study the pharmacological effects and mechanism of the effective composition on MRL/lpr lupus nephritis mice. This project will elucidate the material basis and the mechanism of the Artemisia annua on the treatment of Lupus nephritis, and provide a new point of sight on the treatment of the Lupus nephritis.

狼疮肾炎（lupus nephritis, LN）是系统性红斑狼疮最严重的并发症，迄今尚无有效的治疗药物。补体系统的过度激活在LN的发病机制中起着重要作用。青蒿具有很好地防治LN的作用。我们的预实验结果显示青蒿具有抗补体活性，但二者是否存在相关性有待进一步探讨。为此，我们提出青蒿可能通过抗补体活性这一机制来治疗LN。基于以上假说，本项目以体外抗补体活性导向分离活性成分，采用现代波谱技术鉴定单体化合物的结构。在此基础上，利用制备HPLC富集抗补体活性成分组成活性组分，采用补体激活产物C3b刺激B淋巴细胞活化，研究活性组分对B淋巴细胞分化的影响，并在整体动物水平上探讨活性组分对MRL/lpr狼疮肾炎小鼠的药理作用与机制。目标是阐明青蒿治疗LN的物质基础与作用机制，为青蒿治疗LN提供现代科学依据，为LN的治疗提供新的思路。

本项目创新性提出“青蒿通过抑制补体过度激活可能是治疗狼疮肾炎的机制”之假说。为验证这一假说，本项目采用活性导向分离法，从青蒿的抗补体活性部位分离鉴定了12个化合物，并对这些化合物的抗补体活性和靶点进行了研究，从中发现9个化合物具有不同程度的抑制补体经典途径和旁路途径的活性，活性成分涉及黄酮类、倍半萜类化合物，在这些化合物中尤以青蒿素的抗补体活性最强，靶点研究发现青蒿素作用于补体C1q、C3、C4、C5和C9。动物实验研究发现，与阳性药强的松比，青蒿素联合强的松可以显著降低MRL/lpr狼疮小鼠尿蛋白和血清肌酐、ANA抗体、抗ds-DNA抗体水平，升高血清C1q、C3、C5的水平，改善脾脏肿大症状，提高MRL/lpr狼疮小鼠的生存率。肾脏病理学研究显示，青蒿素联合强的松组肾间质、肾小球炎性细胞浸润较模型组明显改善。进一步的机制研究表明联合用药组显著降低肾脏中C1q mRNA、C3 mRNA表达。免疫组化也显示，青蒿素联合强的松显著减轻C5b在肾脏的沉积。本项目的研究明确了青蒿的抗补体活性成分，阐明了青蒿治疗狼疮肾炎的物质基础与作用机制，同时为补体抑制剂治疗狼疮肾炎提供了理论基础。