我国特色食药用真菌抗阿尔茨海默病新结构活性次级代谢产物的研究

With the great increase of Alzheimer's patients in the aging population of China, the need for new anti-Alzheimer's drug and healthy food with intellectual property is becoming urgent. Edible and medicinal fungi are the key group of the fungal reservoir of China, and the important components of Traditional Chinese Medicine. Edible and medicinal fungi are well known for their production secondary metabolites with neurotrophic activity, and their effective in the prevention and treatment of Alzheimer's. To investigate the bioactive secondary metabolites with anti-Alzheimer's bioactivity from edible and medicinal fungi, the fermentation techniques with high efficiency and the high throughput screening method was combined to screen the culture extract of two hundred strains. The culture extracts of three strains belonging to Cyathus sp., Coprinus sp., and Pleurotus sp. were found to have the potential of stimulating the transformation of neuroglial cells into neuronal cells and further the development of neural synapses bodies. The HPLC fingerprint profile analysis of the culture extracts from these three strains indicated higher chemical diversity of the secondary metabolites. We applied interdisciplinary theory and methods to identify the bioactive secondary metabolites with the anti-Alzheimer's bioactivity from the edible and medicinal fungi in this project, which will greatly enhance the research and development of the edible and medicinal fungi in China, and further promote the protection for the resource of edible and medicinal fungi.

我国人口老龄化加剧，阿尔茨海默病患者大幅增加，急需研发具有自主知识产权的药物和健康产品。食药用真菌是我国真菌资源宝库和中医药体系的重要组成部分。食药用真菌能够产生神经营养和保护的活性次级代谢产物，可以有效地预防阿尔茨海默病的发生和延缓阿尔茨海默病的进程。本课题将食药用真菌高效发酵技术与高通量活性筛选相结合，对200余株食药用真菌的发酵提取物进行抗阿尔茨海默活性次级代谢产物的筛选。发现一株黑蛋巢属、一株鬼伞属和一株侧耳属真菌的发酵物可以促进神经胶质细胞向神经元细胞转化和神经突触小体的形成。高效液相指纹图谱分析显示这三株真菌含有丰富的次级代谢产物。本项目以食药用真菌为研究对象，应用多种发酵技术结合次级代谢产物分离及体内外活性评价，获得食药用真菌抗阿尔茨海默病活性次级代谢产物。本项目应用多学科的理论和方法，开展食药用真菌资源研究，对保护我国特色食药用真菌资源及深入开发利用具有重要意义。

以前期筛选得到的活性菌株侧耳属真菌Pleurotus sp.（泡囊侧耳P. cystidiosush和金顶侧耳P. citrinopileatus）、金针菇（Flammulina velutipes）、猴头菇Hericium erinaceush和Aspergillus oryzae的发酵产物为研究对象，通过乙酸乙酯萃取得到粗提物，采用各种色谱分离手段和波谱技术，构建了包含130个化合物（63个新结构化合物）的次级代谢产物库。利用H2O2诱导PC12细胞氧化损伤模型评价了化合物的神经保护作用，结果显示分离自泡囊侧耳、金针菇等食用菌的15个化合物对H202诱导PC12细胞损伤具有中等强度的保护作用（200μM浓度下，其生存率在70-90%）；18个化合物具有NGF样作用，促进PC12细胞的轴突生长，这一结果为食用菌在老年痴呆方面的应用提供了理论基础。另外，我们建立GFP-LC3稳定表达的HeLa细胞筛选平台，从小分子化合物库筛选到一个能够诱导自噬的二倍半萜类化合物X15-2，该化合物能通过诱导自噬降解α-synuclein，大量研究证明α-synuclein的异常聚集与帕金森病的发病密切相关，因此该研究为神经退行性疾病的治疗带来新的思路。