线粒体ROS
基因敲除
线粒体
细胞生物学
生物
活性氧
内皮功能障碍
氧化磷酸化
神经颗粒素
伊诺斯
一氧化氮
分子生物学
信号转导
一氧化氮合酶
细胞凋亡
内分泌学
生物化学
蛋白激酶C
作者
Ashton N. Jorgensen,Nabil A. Rashdan,K.N. Shashanka Rao,Luisa F. Delgadillo,Gopi K. Kolluru,David M. Krzywanski,Christopher B. Pattillo,Christopher Kevil,Hyung Wook Nam
出处
期刊:Redox biology
[Elsevier]
日期:2024-04-01
卷期号:70: 103085-103085
标识
DOI:10.1016/j.redox.2024.103085
摘要
Endothelial dysfunction and endothelial activation are common early events in vascular diseases and can arise from mitochondrial dysfunction. Neurogranin (Ng) is a 17kD protein well known to regulate intracellular Ca2+-calmodulin (CaM) complex signaling, and its dysfunction is significantly implicated in brain aging and neurodegenerative diseases. We found that Ng is also expressed in human aortic endothelial cells (HAECs), and depleting Ng promotes Ca2+-CaM complex-dependent endothelial activation and redox imbalances. Endothelial-specific Ng knockout (Cre-CDH5-Ngf/f) mice demonstrate a significant delay in the flow-mediated dilation (FMD) response. Therefore, it is critical to characterize how endothelial Ng expression regulates reactive oxygen species (ROS) generation and affects cardiovascular disease. Label-free quantification proteomics identified that mitochondrial dysfunction and the oxidative phosphorylation pathway are significantly changed in the aorta of Cre-CDH5-Ngf/f mice. We found that a significant amount of Ng is expressed in the mitochondrial fraction of HAECs using western blotting and colocalized with the mitochondrial marker, COXIV, using immunofluorescence staining. Seahorse assay demonstrated that a lack of Ng decreases mitochondrial respiration. Treatment with MitoEbselen significantly restores the oxygen consumption rate in Ng knockdown cells. With the RoGFP-Orp1 approach, we identified that Ng knockdown increases mitochondrial-specific hydrogen peroxide (H2O2) production, and MitoEbselen treatment significantly reduced mitochondrial ROS (mtROS) levels in Ng knockdown cells. These results suggest that Ng plays a significant role in mtROS production. We discovered that MitoEbselen treatment also rescues decreased eNOS expression and nitric oxide (NO) levels in Ng knockdown cells, which implicates the critical role of Ng in mtROS-NO balance in the endothelial cells.
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