丹参经表观遗传调控Nrf2/ARE通路及降低核苷酸类似物肾毒性的作用机制研究

项目名称： 丹参经表观遗传调控Nrf2/ARE通路及降低核苷酸类似物肾毒性的作用机制研究

项目编号： No.81473489

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 蒋学华

作者单位： 四川大学

项目金额： 75万元

中文摘要： 中西药联用的临床目的在于增效减毒，揭示联用时增效减毒的分子机制则有助于为临床用药提供科学依据。本课题组的前期研究结果初步提示，丹参具有潜在的降低核苷酸类似物(PNAs)肾毒性的作用。为揭示该现象产生的机理，本项目在已有的工作基础上，围绕丹参是否通过影响Nrf2/ARE信号通路，继而诱导MRP4的表达，减少肾小管上皮细胞的药物蓄积，实现对PNAs肾毒性的减毒作用开展研究。研究工作选择阿德福韦为PNAs的代表，采用报告基因系统、染色质免疫共沉淀等方法，揭示Nrf2调控MRP4的机理；应用DNA甲基化免疫沉淀和亚硫酸氢盐测序技术探索丹参对于肾上皮细胞中Nrf2的表观遗传调控机制；结合细胞摄取实验和基因敲除的动物模型，宏观呈现联用状态下，药物外排率及对小鼠肾功能的影响。研究工作将揭示丹参降低PNAs肾毒性的作用机制，为丹参与PNAs联用提供科学依据，也为中西药联合应用的分子机制研究开阔思路。

中文关键词： 丹参；表观遗传；Nrf2/ARE信号通路；核苷酸类似物；肾毒性

英文摘要： The purpose of herb drug combination is to exert the synergistic effect as well as attenuate side effect. Revealing the underlying will provide a scientific basis for clinical use. Our previous studies showed that Salvia miltiorrhiza (danshen) extract could decrease the nephrotoxicity of adefovir, a nucleotide analog antiviral medicine. The induction of MRP4 expression via Nrf2/ARE pathway leading to enhanced efflux of adefovir in renal proximal epithelium could be a potential mechanism. In this proposal, the following experiments will be carried out to identify the exact mechanisms. The luciferase reporter gene system and chromatin immunoprecipate techniques will be utilized to verify the regulatory effect of Nrf2 on MRP4 through binding the ARE in MRP4 promoter. The methylation DNA immunoprecipitation and bisulfite genomic sequencing method will then be applied to investigate the epigenetic regulation of Nrf2 by Salvia miltiorrhiza extract as well as the active components in renal proximal cells. Based on the results of the above in vitro studies, the cellular efflux as well as in vivo efficacy study using both normal mice and Nrf2(-/-) mice will be performed to validate our findings of reducing nephrotoxicity of adefovir by co-administration of Salvia miltiorrhiza. The outcome from the current project will not only provide the scientific evidence of Salvia miltiorrhiza effect on reducing toxicity, but broaden the area of future application of herb-drug combination.

英文关键词： Salvia miltiorrhiza;Epigenetics;Nrf2/ARE pathway;nucleotide analogs;Nephrotoxicity