基于Keap1-Nrf2/ARE信号通路探讨橙皮素防治AMD的作用机制

Age-related macular degeneration (AMD) is an irreversible vision loss disease induced by multiple risk factors. Oxidative stress, which is closely related to aging, plays an important role in the development and progression of AMD. The nuclear factor erythroid 2-related factor 2 (Nrf2) is an important transcription factor for cells to mediate the resistance to oxidative stress. The Keap1-Nrf2/ARE signalling pathway is essential for cellular redox homeostasis and its activation is one of the main defense mechanisms against oxidative stress. Activation of the Keap1-Nrf2/ARE pathway has been identified as a major target in cellular defense against oxidative stress of AMD. In our previous study, we screened several bioactive compounds which have antioxidant potential. Hesperetin (Hesp), a common flavanone glycoside, has been suggested to exhibit the role of anti-oxidant in ARPE-19 cells and anti-inflammatory in RAW 264.7 cells via activating Nrf2/HO-1 pathway. In addition, Hesp dramatically inhibited H2O2-induced Aβ protein expression, which is a known constituent of drusen triggering the devolvement of AMD. Hesp remarkably blocked Aβ-induced downregulation of Nrf2 and production of ROS. Based on the effect of Hesp in pre-experiment, we speculate that Hesp may protect AMD via activating Keap1-Nrf2/ARE pathway. In the present study, we will use Aβ-induced AMD in vitro and in vivo to investigate the inhibitory effect of Hesp on inflammation, oxidative stress and apoptosis, and further demonstrate that Hesp targets the transcription factor Nrf2 to ameliorate oxidative stress, inflammation, apoptosis multiple signaling axes to suppress AMD. These findings will support a novel therapeutic value for Hesp in the treatment of AMD.

年龄相关性黄斑变性（AMD）是由多因素诱发的一种严重致盲性眼科疾病，与衰老密切相关的氧化应激在AMD的发生发展中发挥重要作用。Nrf2作为抑制氧化应激的重要转录因子，其小分子激活剂的应用成为防治AMD的新策略。课题组前期筛选了大量的Nrf2小分子激活剂，其中橙皮素可显著抑制氧化应激损伤及炎症应答，降低H2O2诱导的视网膜色素上皮细胞（ARPE-19）中AMD致病蛋白Aβ的表达、抑制Aβ诱导的Nrf2下调。这些发现提示橙皮素可能通过激活Keap1-Nrf2/ARE信号通路防治AMD。本研究拟采用Aβ诱导的视网膜色素上皮细胞模型和小鼠模型，进一步应用Nrf2敲除的上皮细胞和工具鼠，层次证明橙皮素通过激活Keap1-Nrf2/ARE抗氧化通路抑制炎症、凋亡和炎症小体信号通路，发挥防治AMD作用这一机制。此研究将为临床上AMD的预防和治疗提供新思路。

年龄相关性黄斑变性（AMD）是由多因素诱发的一种严重致盲性眼科疾病，与衰老密切相关的氧化应激在AMD的发生发展中发挥重要作用。Nrf2作为抑制氧化应激的重要转录因子，其小分子激活剂的应用成为防治AMD的新策略。课题组前期筛选了大量的Nrf2小分子激活剂，其中橙皮素可显著抑制氧化应激损伤及炎症应答，降低H2O2诱导的视网膜色素上皮细胞（ARPE-19）中AMD致病蛋白β-淀粉样蛋白（Aβ）的表达、抑制Aβ诱导的Nrf2下调。这些发现提示橙皮素可能通过激活Keap1-Nrf2/ARE信号通路防治AMD。本研究拟采用Aβ诱导的视网膜色素上皮细胞模型和小鼠模型，研究橙皮素对Nrf2/ARE抗氧化通路、炎症和凋亡信号轴、以及敲除Nrf2后其对氧化应激、炎症和凋亡的作用。我们的研究结果发现，橙皮素通过减少炎性细胞因子的分泌、调节氧化标记物、抑制凋亡蛋白的表达等几方面减缓了Aβ诱导的AMD的发生发展。此外，橙皮素通过抑制MAPK、NF-κB和Txnip/NLRP3信号通路有效缓解炎症反应。同时，橙皮素诱导Keap1-Nrf2/ARE信号通路的激活，从而发挥保护视网膜的活性。综上所述我们的研究表明，橙皮素对年龄相关性黄斑变性具有保护作用，可能与Keap1-Nrf2/ARE的激活密切相关。