Vascular Biology Center, Medical College of Georgia at Augusta University, Augusta, Georgia.
Departments of Pharmacology and Toxicology, Medical College of Georgia at Augusta University, Augusta, Georgia.
Am J Physiol Cell Physiol. 2019 Dec 1;317(6):C1161-C1171. doi: 10.1152/ajpcell.00264.2019. Epub 2019 Sep 25.
NADPH oxidase (NOX)-derived reactive oxygen species (ROS) and copper (Cu), an essential micronutrient, have been implicated in vascular inflammatory diseases. We reported that in proinflammatory cytokine TNF-α-stimulated endothelial cells (ECs), cytosolic Cu chaperone antioxidant-1 (Atox1) functions as a Cu-dependent transcription factor for the NOX organizer p47phox, thereby increasing ROS-dependent inflammatory gene expression. However, the role and mechanism of Atox1 nuclear translocation in inflamed ECs remain unclear. Using enface staining and nuclear fractionation, here we show that Atox1 was localized in the nucleus in inflamed aortas from ApoE mice with angiotensin II infusion on a high-fat diet, while it was found in cytosol in those from control mice. In cultured human ECs, TNF-α stimulation promoted Atox1 nuclear translocation within 15 min, which was associated with Atox1 binding to TNF-α receptor-associated factor 4 (TRAF4) in a Cu-dependent manner. TRAF4 depletion by siRNA significantly inhibited Atox1 nuclear translocation, p47phox expression, and ROS production as well as its downstream VCAM1/ICAM1 expression and monocyte adhesion to inflamed ECs, which were rescued by overexpression of nuclear targeted Atox1. Furthermore, Atox1 colocalized with TRAF4 at the nucleus in TNF-α-stimulated inflamed ECs and vessels. In summary, Cu-dependent Atox1 binding to TRAF4 plays an important role in Atox1 nuclear translocation and ROS-dependent inflammatory responses in TNF-α-stimulated ECs. Thus the Atox1-TRAF4 axis is a novel therapeutic target for vascular inflammatory disease such as atherosclerosis.
NADPH 氧化酶（NOX）衍生的活性氧（ROS）和铜（Cu），作为一种必需的微量营养素，与血管炎症性疾病有关。我们报道，在促炎细胞因子 TNF-α刺激的内皮细胞（ECs）中，细胞质铜伴侣抗氧化剂-1（Atox1）作为 NOX 组织因子 p47phox 的 Cu 依赖性转录因子发挥作用，从而增加 ROS 依赖性炎症基因表达。然而，Atox1 核转位在炎症内皮细胞中的作用和机制仍不清楚。使用 Enface 染色和核分离，我们在这里显示，在高脂肪饮食加血管紧张素 II 输注的 ApoE 小鼠的炎症主动脉中，Atox1 定位于细胞核中，而在对照小鼠的主动脉中则位于细胞质中。在培养的人 ECs 中，TNF-α刺激在 15 分钟内促进 Atox1 核转位，这与 Atox1 以 Cu 依赖性方式与 TNF-α受体相关因子 4（TRAF4）结合有关。用 siRNA 耗尽 TRAF4 可显著抑制 Atox1 核转位、p47phox 表达和 ROS 产生，以及其下游 VCAM1/ICAM1 表达和单核细胞对炎症内皮细胞的黏附，而过表达核靶向 Atox1 则可挽救这些变化。此外，在 TNF-α刺激的炎症内皮细胞和血管中，Atox1 与 TRAF4 在核内共定位。总之，Cu 依赖性 Atox1 与 TRAF4 的结合在 TNF-α 刺激的 ECs 中 Atox1 核转位和 ROS 依赖性炎症反应中发挥重要作用。因此，Atox1-TRAF4 轴是动脉粥样硬化等血管炎症性疾病的新的治疗靶点。
