风轮菜黄酮类成分调控Nrf2/ARE信号通路诱导Ⅱ相解毒酶抗心肌缺血再灌注损伤的分子机制及构效关系研究

The Nrf2-ARE signaling pathway has been proved to be the key endogenous antioxidant responses. The coordinated transcriptional activation of Nrf2-mediated phase II detoxification enzymes is a potential mechanism to maintain redox homeostasis and avoid the deleterious effects of myocardial ischemia reperfusion injury (MIRI), suggesting that Nrf2 is a new target for MIRI. Clinopodium chinense(Benth.)O.Kuntze, affiliated to the genus Clinopodium of family Labiate, has obvious effects of promoting blood circulation and stanching. Previous studies found that the total flavonoids of Clinopodium chinensis and its monomer component luteolin possessed considerable cardiovascular regulatory effects, and the mechanisms may be associated with enhancing cellular antioxidant defense capacity through the activation of Akt and ERK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction. These results indicate that the protective effects of Clinopodium chinensis flavonoid constituents against MIRI may be related to activate Nrf2 and induce phase II enzymes through the antioxidant response element. In our project, through the studies on the structure activity relationship of the protective effects of thirteen Flavonoid components in Clinopodium chinensis against MIRI, we hope to establish the theoretical foundation for developing novel leading compounds, and elucidate the association of its cardioprotective effects with the regulation of PI3K/Akt and other upstream signal pathways that activates Nrf2, induces phase II detoxification enzymes, and protects cardiomyocytes from oxidant-induced apoptosis in a Nrf2-dependent manner, exploring the material base and molecular mechanisms of the protective effects of Clinopodium chinensis flavonoids against MIRI.

Nrf2/ARE 通路是重要的内源性抗氧化应激通路，通过诱导Ⅱ相解毒酶的表达调控MIRI的氧化应激稳态失衡，是探寻抗MIRI药物的新靶标。风轮菜为唇形科风轮菜属植物，具有止血、活血的功效，前期我们研究发现其总黄酮、单体成分木犀草素等具有显著的心血管活性，其机制与增加抗氧化能力，调节上游信号通路PI3K/Akt、ERK进而激活Nrf2/ARE通路并诱导HO-1表达相关。提示风轮菜黄酮类成分抗MIRI作用可能通过干预Nrf2/ARE信号通路并调控信号传导上游环节，进而诱导Ⅱ相解毒酶的表达有关。本项目拟对风轮菜13种不同类型黄酮类成分进行抗MIRI的构效关系研究，为开发新颖结构的先导化合物奠定理论基础。并阐明其心肌保护作用可能与调控PI3K/Akt等信号传导上游环节，通过磷酸化修饰，影响Nrf2/ARE的结合，诱导Ⅱ相解毒酶表达，减少细胞凋亡有关。探索其抗MIRI的药效物质基础及分子机制。

Nrf2/ARE 通路是重要的内源性抗氧化信号通路，可通过诱导Ⅱ相解毒酶的表达调控MIRI的氧化应激稳态失衡。风轮菜为唇形科风轮菜属植物，具有止血、活血的功效。本项目对从风轮菜中分离提取出的风轮菜总黄酮（TFCC）及其单体成分进行了心肌保护活性筛选、构效关系及机制研究。表明TFCC对心肌缺血再灌注损伤保护作用显著，其中单体成分柚皮苷（Naringin）及紫丹酸（Tournefolic acid B,TAB）活性较强。进一步研究发现，风轮菜总黄酮抗缺血性心肌病主要与激活AKT/Nrf2信号通路，增加细胞内源性抗氧化活性有关，同时发现了风轮菜总黄酮对阿霉素诱导的心肌损伤亦有显著的保护作用。柚皮苷可通过诱导ERK、AKT、PKC的磷酸化，进而激活Nrf2的核转位及抗氧化蛋白及分子HO-1、GCLC的表达，对A/R诱导的H9C2氧化应激损伤具有很好的保护作用。TAB对缺血性心脏病有确切的保护作用，其机制与细胞凋亡及PI3K/AKT和MAPK信号通路相关。本研究主要取得了如下研究成果：创新性发现风轮菜总黄酮抗心肌缺血作用；首次发现TAB活性分子，并采用合成化学方法可成功合成TAB，证明其可能成为治疗缺血性心肌病的潜在候选药物；该研究为揭示风轮菜黄酮类化合物的抗心肌缺血作用的药效物质基础、发现先导化合物提供了科学依据，并为基于Nrf2/ARE信号通路抗缺血性心肌病的新药发现提供示范。本项目已发表相关学术论文8篇，其中包括SCI论文7篇，核心期刊论文1篇；另有1篇SCI论文待发表；申请国家发明专利2项；该项研究工作获得2015年北京药学会年会优秀论文二等奖，2015中国医学科学院药用植物研究所药植论坛优秀论文三等奖；获中华中医药学会中青年创新人才称号1名。培养毕业硕士研究生3名和博士研究生2名。