知母盐炙转化多糖与入肾经药效相关性及分子通路研究

In the past, the research on Anemarrhena asphodeloides Bunge was focused on small molecules or original polysaccharides. Previous studies have shown that the main change of salt processed Anemarrhena asphodeloides Bunge was polysaccharides which form a new salt polysaccharide and make the polysaccharide chain shorter. The protein combined with polysaccharide is more easily removed and the activities of polysaccharide have been enhanced. It was revealed that the purpose of salt processed Anemarrhena asphodeloides Bunge was to get salt polysaccharide. It was found that the activity of salt polysaccharide was stronger. It was related to the traditional effect of salt processed into the kidney and releasing deficiency fire of kidney. We provide the hypothesis that the transformation of polysaccharide should be one of the important mechanisms of the traditional Chinese medicine processing. Salt polysaccharide, compared with the original polysaccharide, the content of protein combined with the polysaccharide is much lower than that of the original polysaccharide, which may be easily removed by heating, made protein denaturation and play better effectiveness. In this study, it has essential meaning to find out the new active polysaccharide component, clarify its immunosuppressive effect and mechanism, and clarify the connotation of traditional Chinese medicine. We will make a systematic chemical study on original polysaccharides and salt polysaccharides, explain the structure of polysaccharide, screen the active homologous polysaccharide, investigate the structure - activity relationship, explore the molecular mechanism of polysaccharide action, and clarify the relationship between foundation and efficacy from the point of view of the whole composition of the split.

以往对知母的研究大多关注的是小分子或原生多糖，我们前期研究证明：知母的盐炙炮制主要改变了大分子多糖成分，形成新的转化多糖,使糖链变短、与糖结合的蛋白成分更易于去除、糖的活性增强。从而揭示：获得转化多糖应为知母盐炙的炮制目的，并发现转化多糖活性更强，与盐炙入肾、泻肾经虚火的传统功效正相关。提出了“多糖的炮制转化应是中药炮制原理的重要机制之一。转化多糖相比原生多糖，所含的与糖天然结合的蛋白含量大为降低，可能由于加热炮制会使蛋白变性而易于除去，可以使多糖进入机体后更好发挥与机体受体蛋白的接合，而更好地发挥效能”的假说。本研究对于发现新的活性多糖组分，阐明其免疫抑制作用及机制，阐明中药炮制的内涵具有重要意义。拟对知母的原生多糖、转化多糖进行系统的化学研究，阐明多糖的精细结构,筛选活性均一多糖，考察构效关系，探讨多糖作用的分子机制通路；从全成分拆分的角度，阐明物质基础与药效的关联性。

我们前期研究发现，知母经盐炙炮制后产生了活性更高、分子量减少，含蛋白量较低的大量新的转化多糖。转化多糖的药理作用与其入肾经、治疗肾经疾患的传统药效正相关，所以提出“转化多糖应为其药效物质基础的重要组成”的假说，并围绕其开展项目研究。本项目筛选出知母转化多糖生成的最佳炮制工艺；完成了知母原生多糖、转化多糖的分离、纯化，制备出均一多糖，阐明了均一多糖的结构，建立多糖化学研究方法学平台；对于知母原生多糖、转化多糖进行药效学评价、代谢组学、转录组学研究，研究发现知母转化多糖的抗炎与免疫调节作用、利尿作用、解热作用以及治疗系膜增生性慢性肾小球肾炎效果更佳显著，知母转化多糖主要参与甘油磷脂代谢途径，α-亚麻酸代谢途径来发挥抗炎作用，通过调节体内免疫系统下调TNF信号通路来影响炎症代谢，来延缓、改善系膜增生性肾小球肾炎的症状；确定知母转化均一多糖具有显著抗炎与免疫调节作用，其作用机制为抑制炎症因子IL-1β、IL-6及TNF-α的释放且能下调 NF-κB 蛋白的表达；完成了知母化学物质组成的拆分，并证明拆分组分化学成分互不交叉；对拆分组分与知母原生多糖、转化多糖进行解热、抗炎与免疫调节等药效学评价，建立了物质基础与效应的关联分析。发表相关学术论文4篇，其中SCI收录1篇，中文核心期刊收录1篇，一般论文2篇。培养博士毕业生1名，硕士毕业生1名。