Central role of c-Src in NOX5- mediated redox signaling in vascular smooth muscle cells in human hypertension

Camargo, L. L. et al. (2021) Central role of c-Src in NOX5- mediated redox signaling in vascular smooth muscle cells in human hypertension. Cardiovascular Research, (doi: 10.1093/cvr/cvab171) (Early Online Publication)

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Abstract

Aims: NOX-derived reactive oxygen species (ROS) are mediators of signalling pathways implicated in vascular smooth muscle cell (VSMC) dysfunction in hypertension. Among the numerous redox-sensitive kinases important in VSMC regulation is c-Src. However, mechanisms linking NOX/ROS to c-Src are unclear, especially in the context of oxidative stress in hypertension. Here, we investigated the role of NOX-induced oxidative stress in VSMCs in human hypertension focusing on NOX5, and explored c-Src, as a putative intermediate connecting NOX5-ROS to downstream effector targets underlying VSMC dysfunction. Methods and results: VSMC from arteries from normotensive (NT) and hypertensive (HT) subjects were studied. NOX1,2,4,5 expression, ROS generation, oxidation/phosphorylation of signalling molecules, and actin polymerization and migration were assessed in the absence and presence of NOX5 (melittin) and Src (PP2) inhibitors. NOX5 and p22phox-dependent NOXs (NOX1–4) were down-regulated using NOX5 siRNA and p22phox-siRNA approaches. As proof of concept in intact vessels, vascular function was assessed by myography in transgenic mice expressing human NOX5 in a VSMC-specific manner. In HT VSMCs, NOX5 was up-regulated, with associated oxidative stress, hyperoxidation (c-Src, peroxiredoxin, DJ-1), and hyperphosphorylation (c-Src, PKC, ERK1/2, MLC20) of signalling molecules. NOX5 siRNA reduced ROS generation in NT and HT subjects. NOX5 siRNA, but not p22phox-siRNA, blunted c-Src phosphorylation in HT VSMCs. NOX5 siRNA reduced phosphorylation of MLC20 and FAK in NT and HT. In p22phox- silenced HT VSMCs, Ang II-induced phosphorylation of MLC20 was increased, effects blocked by melittin and PP2. NOX5 and c-Src inhibition attenuated actin polymerization and migration in HT VSMCs. In NOX5 transgenic mice, vascular hypercontractilty was decreased by melittin and PP2. Conclusion: We define NOX5/ROS/c-Src as a novel feedforward signalling network in human VSMCs. Amplification of this system in hypertension contributes to VSMC dysfunction. Dampening the NOX5/ROS/c-Src pathway may ameliorate hypertension-associated vascular injury.

Item Type:Articles
Additional Information:This work was funded by grants from the British Heart Foundation (BHF) (RG/13/7/30099, RE/18/6/34217) and the MRC (MC-PC-15076). RMT is supported through a BHF Chair award (CH/4/29762). ACM is supported by a University of Glasgow Walton Fellowship. MH is supported by the Higher Education Commission (HEC), Pakistan.
Status:Early Online Publication
Refereed:Yes
Glasgow Author(s) Enlighten ID:Wang, Yu and Jensen, Mr Thomas and Guzik, Professor Tomasz and Hartley, Professor Richard and Touyz, Professor Rhian and Rios, Dr Francisco and Montezano, Dr Augusto and Neves, Dr Karla and Zou, ZhiGuo and Hussain, Miss Misbah
Authors: Camargo, L. L., Montezano, A. C., Hussain, M., Wang, Y., Zou, Z., Rios, F. J., Neves, K. B., Alves-Lopes, R., Awan, F. R., Guzik, T. J., Jensen, T., Hartley, R. C., and Touyz, R. M.
College/School:College of Medical Veterinary and Life Sciences
College of Medical Veterinary and Life Sciences > Institute of Cardiovascular and Medical Sciences
Journal Name:Cardiovascular Research
Publisher:Oxford University Press
ISSN:0008-6363
ISSN (Online):1755-3245
Published Online:28 July 2021

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
167932Vascular Noxs as therapeutic targets and biomarkers in hypertensionRhian TouyzBritish Heart Foundation (BHF)RG/13/7/30099Institute of Cardiovascular & Medical Sciences
303944BHF Centre of ExcellenceRhian TouyzBritish Heart Foundation (BHF)RE/18/6/34217CAMS - Cardiovascular Science