碘化N-正丁基氟哌啶醇通过调控Nrf2信号通路减轻心肌缺血再灌注损伤机制的研究

Nuclear factor E2-related factor (Nrf2) is a redox-sensitive nuclear transcription factor that up-regulates the gene expression of a battery of antioxidants and cytoprotective enzymes by binding to antioxidant responsive element. Recently, emerging evidence demonstrate that Nrf2 is an important endogenous cardio-protective signal that protects against oxidant-mediated I/R injury. A novel haloperidol derivative, N-n-butyl haloperidol iodide (F2), has been designed and synthesized by us. Our previous studies have shown F2 can inhibit the oxidative stress and confers protection against myocardial ischemia/reperfusion (I/R). Furthermore，we found that F2 could effectively activate Nrf2 and enhance the translocation of PKCε from cytosol to mitochondria. Thus, a hypothesis that F2 may activate Nrf2 and subsequently decrease the oxidant level via regulating the PKCε/AMPK signaling axis was proposed. In the present project, both in vivo mouse heart models and in vitro cardiac myocyte models were exploited to examine the potential for F2 to limit I/R insult. Then, Nrf2 deficient mice will be employed to verify whether Nrf2 is the pivotal target in the cardio-protective effects of F2 against I/R lesion. The regulative mechanism of F2 on Nrf2 in the process of myocardial I/R will be deciphered by utilizing specific inhibitors and siRNA approach. More importantly, the current project will provide novel avenues to the prophylaxis and therapy of myocardial I/R injury.

核因子2相关因子（Nrf2）是一种氧化还原敏感核转录因子，通过与抗氧化反应元件结合调控许多抗氧化蛋白和解毒酶的表达。此外，它是一种重要的内源性心脏保护因子，可通过减轻缺血/再灌注（I/R）诱导的氧化应激抗心肌损伤。碘化N-正丁基氟哌啶醇（F2）是本课题组合成的一种与抑制活性氧自由基密切相关的化合物，具有显著拮抗心肌I/R损伤，改善心功能的作用，且发现F2能有效激活Nrf2和影响PKCε的转位。据此，提出F2通过激活Nrf2降低氧化应激水平可能与其调控PKCε/AMPK信号通路有关。本课题拟采用整体心脏I/R模型和离体心肌细胞H/R模型，明确F2在体内外对心肌的保护作用；然后利用Nrf2基因敲除小鼠，明确Nrf2是否是F2抗心肌I/R损伤的作用靶点；通过采用特异性抑制剂和siRNA等方法，在分子水平上揭示F2对心肌I/R过程中Nrf2的调控机理，确证假说，为防治心肌I/R损伤提供新思路。

本课题分别从在体小鼠I/R模型以及离体小鼠心肌细胞H/R模型上系统观察了F2对心肌缺血损伤的保护作用并进一步深入探究其作用机制。主要结果：1.F2对心肌I/R损伤具有良好的保护作用；2.F2对心肌细胞H/R损伤亦具有良好的保护作用；3.I/R或H/R激活Nrf2，而F2可进一步增加Nrf2的活性；4.心脏特异性敲除Nrf2导致F2对I/R或H/R损伤的保护作用消失；5.I/R或H/R激活PKCε-AMPK信号通路，而F2可进一步提高此信号通路的活化；6.PKCε-AMPK信号通路参与F2对Nrf2的调控。以上工作明确了Nrf2是F2抗心肌缺血损伤的关键靶点并阐明了F2对Nrf2调控的分子机制，为临床防治心肌I/R损伤提供了新思路。