Natural Products Research Institute, College of Pharmacy, Seoul National University, Seoul 151-742, Republic of Korea.
Division of Wood Chemistry & Microbiology, Department of Forest Products, Korea Forest Research Institute, Seoul 130-712, Republic of Korea.
Life Sci. 2015 Jun 1;130:25-30. doi: 10.1016/j.lfs.2015.02.026. Epub 2015 Mar 26.
The aim of this study was to prove the neuroprotective effect of 5,7-Dihydroxychromone (DHC) through the Nrf2/ARE signaling pathway. To elucidate the mechanism, we investigated whether 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in SH-SY5Y cells could be attenuated by DHC via activating the Nrf2/ARE signal and whether DHC could down-regulate 6-OHDA-induced excessive ROS generation
To evaluate the neuroprotective effect of DHC against 6-OHDA-induced apoptosis, FACS analysis was performed using PI staining. The inhibitory effect of DHC against 6-OHDA-induced ROS generation was evaluated by DCFH-DA staining assay. Additionally, translocation of Nrf2 to the nucleus and increased Nrf2/ARE binding activity, which subsequently resulted in the up-regulation of the Nrf2-dependent antioxidant gene expressions including HO-1, NQO1, and GCLc, were evaluated by Western blotting and EMSA.
Pre-treatment of DHC, one of the constituents of Cudrania tricuspidata, significantly protects 6-OHDA-induced neuronal cell death and ROS generation. Also, DHC inhibited the expression of activated caspase-3 and caspase-9 and cleaved PARP in 6-OHDA-induced SH-SY5Y cells. DHC induced the translocation of Nrf2 to the nucleus and increased Nrf2/ARE binding activity which results in the up-regulation of the expression of Nrf2-dependent antioxidant genes, including HO-1, NQO1, and GCLc. The addition of Nrf2 siRNA abolished the neuroprotective effect of DHC against 6-OHDA-induced neurotoxicity and the expression of Nrf2-mediated antioxidant genes.
Activation of Nrf2/ARE signal by DHC exerted neuroprotective effects against 6-OHDA-induced oxidative stress and apoptosis. This finding will give an insight that activating Nrf2/ARE signal could be a new potential therapeutic strategy for neurodegenerative disease.
本研究旨在通过 Nrf2/ARE 信号通路证明 5,7-二羟基色酮（DHC）的神经保护作用。为了阐明机制，我们研究了 DHC 是否可以通过激活 Nrf2/ARE 信号来减轻 6-羟多巴胺（6-OHDA）诱导的 SH-SY5Y 细胞毒性，以及 DHC 是否可以下调 6-OHDA 诱导的过量 ROS 生成。
使用 PI 染色通过 FACS 分析评估 DHC 对 6-OHDA 诱导凋亡的神经保护作用。通过 DCFH-DA 染色试验评估 DHC 对 6-OHDA 诱导的 ROS 生成的抑制作用。此外，通过 Western blot 和 EMSA 评估 Nrf2 向核易位以及增加 Nrf2/ARE 结合活性，进而上调 Nrf2 依赖性抗氧化基因表达，包括 HO-1、NQO1 和 GCLc。
DHC（一种枳椇子的成分）预处理可显著保护 6-OHDA 诱导的神经元细胞死亡和 ROS 生成。此外，DHC 抑制了 6-OHDA 诱导的 SH-SY5Y 细胞中激活的 caspase-3 和 caspase-9 的表达以及 PARP 的裂解。DHC 诱导 Nrf2 向核易位并增加 Nrf2/ARE 结合活性，导致 Nrf2 依赖性抗氧化基因表达上调，包括 HO-1、NQO1 和 GCLc。加入 Nrf2 siRNA 可消除 DHC 对 6-OHDA 诱导神经毒性和 Nrf2 介导的抗氧化基因表达的神经保护作用。
DHC 通过激活 Nrf2/ARE 信号发挥对 6-OHDA 诱导的氧化应激和凋亡的神经保护作用。这一发现将为激活 Nrf2/ARE 信号可能成为神经退行性疾病的新的潜在治疗策略提供启示。
