基于1-磷酸鞘氨醇受体的冬虫夏草免疫抑制药效物质基础及作用机制研究

冬虫夏草因具有多种功效被历代医家推崇，在临床上虽被广泛用于免疫调节，但其免疫抑制的药效物质尚不明晰。冬虫夏草稀有贵重，难以获得足够量的样品是制约其药效物质研究的瓶颈。申请人前期首次从冬虫夏草中分离获得一类结构特殊且主含的脑苷脂化合物，该类化合物在体内代谢生成1-磷酸鞘氨醇后作用于1-磷酸鞘氨醇受体，从而发挥作用。虚拟靶点对接和体外活性结果提示该类化合物是冬虫夏草免疫抑制的药效物质。本项目拟在脑苷脂基因簇被阐明和冬虫夏草菌液体发酵菌丝体中含有相同脑苷脂成分的基础上，采用异源表达和液体发酵策略大量获得该类成分，运用免疫抑制经典动物模型开展药效和作用机制研究，采用成熟的[γ-35S] GTP结合实验进行靶点验证，由此阐明冬虫夏草免疫抑制的药效物质和作用机制，为产品开发和质量标准提供科学依据，从而推动中医药文化传承和产业发展。此外，异源表达和液体发酵策略，也为稀有贵重中药的研究提供方法借鉴。

Cordyceps sinensis has been admired by doctors in all Dynasties for its extensive pharmacological effects. And it has been widely used in clinical for immunoregulation. However, its immunosuppressive substance has not been clarified. Cordyceps sinensis is rare and precious. So, it is difficult to obtain a sufficient amount of sample from Cordyceps sinensis, which are the bottleneck of the study on the effective substances. In our previous study, the cerebrosides with distinctive structures from Cordyceps sinensis were obtained as the main ingredients for the first time. These compounds will become to sphingosine 1-phosphate in body and act on the sphingosine 1-phosphate receptor. The virtual target docking and activity results in vitro showed that these cerebrosides were the main effective substances of Cordyceps sinensis for immunosuppressive activity. Meanwhile, the gene cluster of cerebroside has been elucidated and we found that the liquid fermented mycelium of Cordyceps sinensis also contained the same cerebrosides. In our research, the strategies of heterologous expression and liquid fermentation will be used to obtain such components in large quantities. The immunosuppressive efficacy and mechanism of cerebrosides will be investigated by using the classic animal model of immunosuppression. The target of cerebrosides will be investigated in a [γ-35S] GTP functional assay. The above results will clarify the effective substances and mechanism of Cordyceps sinensis for immunosuppressive effect, which will provide a scientific basis for the development and quality control of Cordyceps sinensis. And it will promote the cultural heritage of Traditional Chinese Medicine and the development of Cordyceps sinensis industry. In addition, the strategies of heterologous expression and liquid fermentation will also provide a method for the study of rare and precious Traditional Chinese Medicine.