益气补肾法对重离子辐射大脑旁效应诱发机体神经-内分泌-免疫系统网络变化调控机制的研究

With the extensive development and application of heavy ion radiation therapy of tumor,injury and its side effects induced by radiation are noticed by the academic circle day by day.A lot of research show that the biological damage by some side effects may be serious than the parts of radiation.Based on the radiation side effects,the research is combined with nerve-endocrine- immune system(NEI)of modern medicine to explain the mechanism of brain network.Supported by the theory “kidney stores essence,and marrow is communicated to the brain”,we’ll choose the classical prescription of Astragalus decoction to interfere in rats.After 12C6+ ion beam radiation of rats’brain,we’ll detect the expression of neurotransmitter and biological behaviour of brain cells by cell morphology, immunohistochemistry, biochip,RT-PCR and WB.Also,we”ll study the morphology of kidney and bone marrow cells,the expression of cells’ proliferation and apoptosis-associated genes,the molecular expression of nerve-endocrine system,the activity of spleen,peritoneal cells and the expression of immune molecules. Finally,we”ll focus our attention on the content changes of brain,kidney,spleen,thymus,bone marrow cells in condiations of radiation side effects and DNA methylation changes of brain,spleen,thymus,kidney and bone marrow cells.In a word,we want to explore the mechanism of regulation of radiation side effects in NEI system and the intervention effects by Astragalus decoction,so as to provide scientific basis for the protection from heavy ion radiation sideeffects by traditional Chinese medicine.

随着重离子辐射治疗肿瘤的广泛开展和应用，辐射诱发的损伤及其旁效应也日趋受到学术界的关注，大量研究表明部分旁效应所受到的生物学损伤可能比受辐射部分更强；本课题基于辐射旁效应结合现代医学神经-内分泌-免疫系统（NEI）网络机制的研究，根据中医“肾藏精，生髓通于脑”理论，选用补肾益气的经典方药黄芪汤干预大鼠，通过12C6+离子束辐射大脑，用细胞形态学、免疫化学、生物芯片、RT-PCR、WB 等方法检测脑组织细胞生物学及神经递质的表达；肾及骨髓细胞增殖与凋亡相关基因表达；神经内分泌网络分子表达；脾脏、腹腔细胞活性及免疫分子的表达；脑、肾、脾、胸腺、骨髓细胞旁效应相关指标的变化；脑、脾、胸腺、肾脏及骨髓细胞DNA甲基化变化等指标，从而探明重离子辐射大脑旁效应诱发机体神经-内分泌-免疫系统变化机制及补肾益气法的干预效果，以期为中医药防护重离子旁效应损伤提供科学依据。

近年来，辐射诱发的旁效应损伤日趋受到学术界的关注，研究表明旁效应所受到的生物学损伤可能比受辐射部位更强；基于此，本课题结合现代医学神经-内分泌-免疫系统（NEI）网络机制的研究，利用中医“肾藏精 ，生髓通于脑”理论，选用补肾益气的经典方药黄芪汤干预大鼠，通过12C6+离子束辐射鼠大脑，检测各组大鼠外周血WBC等血细胞数量及脑、肾、垂体的病理形态学，并观察黄芪汤对其影响，筛选出了4.0Gy辐射后7d为黄芪汤干预的最佳碳离子辐射条件；基于网络药理学技术分析了黄芪汤治疗放射性脑损伤的核心靶点176个，KEGG富集发现黄芪汤作用于放射性脑损伤的通路主要有MAPK、p53、NF-κB信号通路；利用现代生物学技术检测脑及肾病理形态变化，神经细胞凋亡、脑中IL-1β等炎症因子和Bcl-2等凋亡分子表达变化及肾中PCNA等凋亡分子和NF-κB等炎性分子表达变化，结果提示：黄芪汤对12C6+离子束辐射脑损伤模型鼠脑及旁效应器官肾损伤的防护机制可能与抑制炎症反应和抗凋亡相关；检测垂体病理变化及下丘脑中IL-1β等炎症因子、miRNA、TLR4/MyD88/NF-κB通路中分子的表达变化，结果提示：黄芪汤对12C6+离子束辐射脑损伤模型鼠旁效应下丘脑-垂体轴的保护机制可能是通过miRNA介导的TLR4/MyD88/ NF-κB的炎症信号通路来实现的；同时检测了外周血细胞数量，脾脏及血清IL-1β等炎症因子，血清CORT等激素和NO等氧化应激分子的含量变化，结果证实，黄芪汤对12C6+离子束辐射脑损伤模型鼠旁效应造血系统损伤、内分泌系统损伤及氧化应激损伤也具有防护作用。本实验探明了重离子辐射大脑旁效应诱发机体神经-内分泌-免疫系统变化机制及补肾益气法的干预效果，为中医药防护重离子旁效应损伤提供了科学依据。