CLOCK disruption aggravates carotid artery stenosis through endoplasmic reticulum stress-induced endothelial-mesenchymal transition

Hanfei Tang; Song Xue; Gefei Zhao; Chao Fang; Liang Cai; Zhenyu Shi; Weiguo Fu; Ruizhe Qian; Pengfei Zhang; Xiao Tang; Daqiao Guo

CLOCK disruption aggravates carotid artery stenosis through endoplasmic reticulum stress-induced endothelial-mesenchymal transition

Am J Transl Res. 2020 Dec 15;12(12):7885-7898. eCollection 2020.

Authors

Hanfei Tang¹, Song Xue¹, Gefei Zhao², Chao Fang¹, Liang Cai¹, Zhenyu Shi¹, Weiguo Fu¹, Ruizhe Qian³, Pengfei Zhang², Xiao Tang¹, Daqiao Guo¹

Affiliations

¹ Department of Vascular Surgery, Institute of Vascular Surgery, Zhongshan Hospital, Fudan University Shanghai, China.
² Department of Thoracic and Cardiovascular Surgery, The Affiliated Drum Tower Hospital, Medical School of Nanjing University Nanjing, Jiangsu, China.
³ Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Shanghai Key Laboratory of Clinical Geriatric Medicine, Fudan University Shanghai, China.

PMID: 33437367
PMCID: PMC7791501

Abstract

Carotid artery stenosis is a leading cause of ischemic stroke, but the underlying mechanism remains unclear. We aimed to determine the molecular mechanisms of carotid plaque progression. We analyzed the molecular and morphometric characteristics of carotid plaque samples obtained from 30 patients who underwent carotid endarterectomy. Additionally, we established a mouse model of carotid atherosclerosis by partially ligating the left common carotid arteries of male Clock^Δ19/Δ19 (Clk) and wild-type (WT) C57BL/6J mice fed a high-fat diet. Clk and WT primary mouse aortic endothelial cells (pMAECs) were exposed to disturbed flow (DF) or undisturbed flow (UF) with or without treatment with the IRE-1α inhibitor STF-083010 or the PERK inhibitor GSK2606414. In human carotid artery plaques, CLOCK expression was lower in the lipid-rich necrotic core than in transitional regions, especially in the endothelium. Decreased CLOCK mRNA levels were associated with more extensive stenosis, intraplaque hemorrhage, and complex plaque in human carotid plaques. In mice, the Clock^Δ19/Δ19 mutation significantly increased neointima formation and neovascularization but decreased collagen content and lumen area in partially ligated carotid arteries. In addition, Clock^Δ19/Δ19 mutants exhibited significantly decreased Cdh5 expression and increased expression of endothelial-mesenchymal transition (EndMT) and endoplasmic reticulum (ER) stress markers in mice with partially ligated carotid arteries and pMAECs exposed to DF. Notably, inhibition of the IRE1α-XBP1 axis abrogated the increased EndMT caused by Clock^Δ19/Δ19 mutation and DF in pMAECs. In conclusion, the disruption of CLOCK function aggravates EndMT via the IRE1α-XBP1 axis, contributing to carotid artery stenosis.

Keywords: Carotid artery stenosis; circadian locomotor output cycles kaput; disturbed flow; endoplasmic reticulum stress; endothelial-mesenchymal transition.