Cigarette smoke extract profoundly suppresses TNF\u03b1-mediated proinflammatory gene expression through upregulation of ATF3 in human coronary artery endothelial cells

Jack E Teasdale,Alasdair M G Peachey,Georgina G J Hazell,Mark Bond,Graciela B Sala-Newby,Stephen J White,Andrew C Newby,Tristan R Mckay,Charles C T Hindmarch

doi:10.1038/srep39945

Abstract

Endothelial dysfunction caused by the combined action of disturbed flow, inflammatory mediators and oxidants derived from cigarette smoke is known to promote coronary atherosclerosis and increase the likelihood of myocardial infarctions and strokes. Conversely, laminar flow protects against endothelial dysfunction, at least in the initial phases of atherogenesis. We studied the effects of TNFα and cigarette smoke extract on human coronary artery endothelial cells under oscillatory, normal laminar and elevated laminar shear stress for a period of 72 hours. We found, firstly, that laminar flow fails to overcome the inflammatory effects of TNFα under these conditions but that cigarette smoke induces an anti-oxidant response that appears to reduce endothelial inflammation. Elevated laminar flow, TNFα and cigarette smoke extract synergise to induce expression of the transcriptional regulator activating transcription factor 3 (ATF3), which we show by adenovirus driven overexpression, decreases inflammatory gene expression independently of activation of nuclear factor-κB. Our results illustrate the importance of studying endothelial dysfunction in vitro over prolonged periods. They also identify ATF3 as an important protective factor against endothelial dysfunction. Modulation of ATF3 expression may represent a novel approach to modulate proinflammatory gene expression and open new therapeutic avenues to treat proinflammatory diseases.

Highlights

Endothelial dysfunction caused by the combined action of disturbed flow, inflammatory mediators and oxidants derived from cigarette smoke is known to promote coronary atherosclerosis and increase the likelihood of myocardial infarctions and strokes
The athero-protective phenotype that endothelial cells adopt in normal physiological laminar shear stress (12–16 dynes/cm[2] in the coronary circulation5–7) is predominantly mediated by increased expression of KLF2 and KLF4, and the activation of Nrf[2], all of which combine to activate a program of gene expression and epigenetic changes that reduce endothelial dysfunction[8,9]
We focus on the interaction of shear stress with the effects of TNFαand cigarette smoke extract (CSE) as a model of endothelial dysfunction, which contributes to the development of atherosclerosis, our intention being to identify new signalling interactions that might be useful for future therapy or as biomarkers

Summary

Introduction

Endothelial dysfunction caused by the combined action of disturbed flow, inflammatory mediators and oxidants derived from cigarette smoke is known to promote coronary atherosclerosis and increase the likelihood of myocardial infarctions and strokes. We studied the effects of TNFα and cigarette smoke extract on human coronary artery endothelial cells under oscillatory, normal laminar and elevated laminar shear stress for a period of 72 hours. The haemodynamic environment plays an important contributing role in the initiation and progression of atherosclerosis, with disease developing and progressing most rapidly in regions of oscillatory, low and disturbed wall shear stress[1,2,3,4] Underlying this response is the mechanosensitivity of endothelial cells, whose behaviour is profoundly modulated by the shear environment to which they are exposed. The athero-protective phenotype that endothelial cells adopt in normal physiological laminar shear stress (12–16 dynes/cm[2] in the coronary circulation5–7) is predominantly mediated by increased expression of KLF2 and KLF4, and the activation of Nrf[2], all of which combine to activate a program of gene expression and epigenetic changes that reduce endothelial dysfunction[8,9]. We have previously demonstrated that fresh aqueous cigarette smoke extract (CSE) induces a stress response in human coronary artery endothelial cells, activating transcription factor Nrf[2] and increasing the expression of cytochrome p450 subunits[23]

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