Department of Bioscience Technology, Chung Yuan Christian University, Chungli, Taiwan.
Department of Neurosurgery, Chi Mei Medical Center, Tainan, Taiwan.
Am J Physiol Cell Physiol. 2021 Apr 1;320(4):C635-C651. doi: 10.1152/ajpcell.00515.2019. Epub 2020 Dec 23.
Disruption of copper homeostasis is closely involved in neurodegenerative disorders. This study examined whether a hybrid copper-binding compound, ()-2-(4-(dimethylamino)phenylimino)methyl)quinolin-8-ol (DPMQ), is able to protect NG108-15 cells against oxidative stress. We found that treatment of cells with rotenone or hydrogen peroxide increased cellular oxidative stress and resulted in mitochondrial dysfunction and apoptosis. The cellular levels of Nrf2 and the Cu chaperone DJ-1 were also decreased. These oxidative detrimental effects were all inhibited when cells were cotreated with DPMQ. DPMQ increased cellular Cu content, DJ-1 protein level, superoxide dismutase (SOD) activity, and Nrf2 nuclear translocation under basal state. The activity of SOD decreased under redox imbalance and this decrease was blocked by DPMQ treatment, while the protein level of SOD1 remained unaltered regardless of the oxidative stress and DPMQ treatment. Using endogenous proteins, coimmunoprecipitation showed that DJ-1 bound with SOD1 and Nrf2 individually. The amount of Nrf2, bound to DJ-1, consistently reflected its cellular level, while the amount of SOD1, bound to DJ-1, was potentiated by DPMQ, being greater in the basal state than under redox imbalance. Simultaneous inclusion of nonpermeable Cu chelator tetrathiomolybdate or triethylenetetramine during DPMQ treatment blocked all aforementioned effects of DPMQ, showing that the dependency of the effect of DPMQ on extracellular Cu. In addition, silencing of DJ-1 blocked the protection of DPMQ against oxidative stress. Taken all together, our results suggest that DPMQ stabilizes DJ-1 in a Cu-dependent manner, which then brings about SOD1 activation and Nrf2 nuclear translocation; these together alleviate cellular oxidative stress.
铜稳态的破坏与神经退行性疾病密切相关。本研究探讨了一种混合铜结合化合物（）-2-（4-（二甲基氨基）苯基亚氨基）甲基）喹啉-8-醇（DPMQ）是否能够保护 NG108-15 细胞免受氧化应激。我们发现，用鱼藤酮或过氧化氢处理细胞会增加细胞氧化应激，导致线粒体功能障碍和细胞凋亡。Nrf2 和铜伴侣 DJ-1 的细胞水平也降低了。当细胞用 DPMQ 共处理时，这些氧化损伤作用都被抑制了。DPMQ 在基础状态下增加了细胞内的铜含量、DJ-1 蛋白水平、超氧化物歧化酶（SOD）活性和 Nrf2 核易位。在氧化还原失衡下，SOD 的活性降低，而 DPMQ 处理阻止了这种降低，而 SOD1 的蛋白水平无论是否存在氧化应激和 DPMQ 处理都保持不变。使用内源性蛋白，共免疫沉淀表明 DJ-1 分别与 SOD1 和 Nrf2 结合。与 DJ-1 结合的 Nrf2 的量始终反映其细胞水平，而与 DJ-1 结合的 SOD1 的量则被 DPMQ 增强，在基础状态下比在氧化还原失衡时更大。在 DPMQ 处理期间同时包含不可渗透的铜螯合剂四硫钼酸盐或三乙烯四胺时，阻断了 DPMQ 的所有上述作用，表明 DPMQ 的作用对细胞外铜的依赖性。此外，DJ-1 的沉默阻断了 DPMQ 对氧化应激的保护作用。总的来说，我们的结果表明，DPMQ 以铜依赖的方式稳定 DJ-1，从而导致 SOD1 的激活和 Nrf2 的核易位；这些共同减轻细胞氧化应激。
