Atheroprone flow activates inflammation via endothelial ATP-dependent P2X7-p38 signalling

Abstract

ObjectiveAtherosclerosis is a focal disease occurring at arterial sites of disturbed blood flow that generates low oscillating shear stress. Endothelial inflammatory signalling is enhanced at sites of disturbed flow via mechanisms that are incompletely understood. The influence of disturbed flow on endothelial adenosine triphosphate (ATP) receptors and downstream signalling was assessed.Methods and resultsCultured human endothelial cells were exposed to atheroprotective (high uniform) or atheroprone (low oscillatory) shear stress for 72 h prior to assessment of ATP responses. Imaging of cells loaded with a calcium-sensitive fluorescent dye revealed that atheroprone flow enhanced extracellular calcium influx in response to 300 µM 2'(3')-O-(4-Benzoylbenzoyl) adenosine-5'-triphosphate. Pre-treatment with pharmacological inhibitors demonstrated that this process required purinergic P2X7 receptors. The mechanism involved altered expression of P2X7, which was induced by atheroprone flow conditions in cultured cells. Similarly, en face staining of the murine aorta revealed enriched P2X7 expression at an atheroprone site. Functional studies in cultured endothelial cells showed that atheroprone flow induced p38 phosphorylation and up-regulation of E-selectin and IL-8 secretion via a P2X7-dependent mechanism. Moreover, genetic deletion of P2X7 significantly reduced E-selectin at atheroprone regions of the murine aorta.ConclusionsThese findings reveal that P2X7 is regulated by shear forces leading to its accumulation at atheroprone sites that are exposed to disturbed patterns of blood flow. P2X7 promotes endothelial inflammation at atheroprone sites by transducing ATP signals into p38 activation. Thus P2X7 integrates vascular mechanical responses with purinergic signalling to promote endothelial dysfunction and may provide an attractive potential therapeutic target to prevent or reduce atherosclerosis.

Bacterial lipopolysaccharide and antimicrobial LL-37 enhance ICAM-1 expression and NF-κB p65 phosphorylation in senescent endothelial cells

“…These observations suggested that upregulated P2X7 may have a more important role in the enhanced response to LL-37 in senescent endothelial cells. In this regard, it is interesting to note that endothelial P2X7 is increased in atherosclerotic lesions of mouse aorta (39) and is required for inflammatory signaling in endothelial cells exposed to low shear stress mimicking atherogenic conditions (39). These observations suggest that LL-37 localized on endothelial cells of human atherosclerotic lesion (14) may contribute to the enhanced inflammatory response during atherogenesis possibly via the upregulated P2X7.…”

P2 Receptors as Therapeutic Targets in the Salivary Gland: From Physiology to Dysfunction

“…et al, 2017). The P2X7R is widely expressed in diverse tissues, including hematopoietic cells (Feng et al, 2016), neurons (Miras-Portugal et al, 2017), glia (Stokes et al, 2015;Kaczmarek-Hajek et al, 2018), bone (Agrawal and Gartland, 2015), muscle (Fabbrizio et al, 2019), endothelium (Green et al, 2018), epithelium (Woods et al, 2012), and immune cells (Ferrari et al, 1997). In the exocrine pancreas, P2X7Rs have been shown to be primarily expressed in pancreatic ductal cells where they may contribute to secretory regulation through induction of cation fluxes and interaction with cholinergic signaling (Novak et al, 2010;Burnstock and Novak, 2012).…”

Computational Characterization of the Dish-In-A-Dish, A High Yield Culture Platform for Endothelial Shear Stress Studies on the Orbital Shaker

“…Cells subjected to this type of shear stress align poorly and show an increased expression of proinflammatory markers such as monocyte chemoattractant protein 1 (MCP1), a chemokine involved in monocyte attraction, and cadherin-2 (CDH2), a mesenchymal marker [5][6][7][8][9]. High laminar flow profiles (shear stresses > 1 Pa) are considered atheroprotective [7,10]. Cells that are exposed to an atheroprotective flow align in the direction of the flow, form a tight vascular barrier and express anti-inflammatory genes [11].…”