Statins protect human endothelial cells from TNF-induced inflammation via ERK5 activation
Introduction
Inflammation plays a critical role in cardiovascular disease, and the inflammatory cascade is particularly important in the atherosclerotic process. The inflammatory mediator tumor necrosis factor (TNF, also known as TNF-α) has been implicated in the pathogenesis of a number of cardiovascular diseases, including atherosclerosis, myocardial infarction, heart failure, myocarditis and cardiac allograft rejection [1]. In response to TNF, vascular endothelial cells promote inflammation changes, which increase leukocyte adhesion, transendothelial migration and vascular leak and promote thrombosis, by displaying, in a distinct temporal, spatial and anatomical pattern [2], [3], [4], different combinations of adhesion molecules for leukocytes, including vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1) and E-selectin [5], [6]. In addition, TNF is vital for the ICAM-1-dependent recruitment of mononuclear cells and microvascular damage [7]. The central role of TNF in inflammation has also been demonstrated by the ability of TNF blocker to treat a range of cardiovascular disorders and inflammatory conditions, including acute myocardial infarction (AMI), heart failure, rheumatoid arthritis, diabetes and hyperlipidaemia [1], [8]. Despite the important pathological role of TNF in cardiovascular diseases, the exact mechanisms underlying TNF-induced vascular inflammation and dysfunction remain unresolved and there is no current scientific consensus.
Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors (statins) are potent inhibitors of cholesterol biosynthesis widely used to reduce serum cholesterol levels in hyperlipidemic patients [9]. In addition to lowering lipids, these drugs exhibit potent anti-inflammatory effects mediated by inhibition of macrophage function. Statins are the most effective agents available today for the reduction of vascular inflammation. However, the mechanisms by which statins may exert beneficial anti-inflammatory effects independent of lipid-lowering have not been completely identified.
Extracellular-signal-regulated kinase 5 (ERK5) is the newest member of the mitogen-activated protein kinase (MAPK) family. Similar to other MAPK family members, ERK5 plays a significant role in cell growth and differentiation. Nevertheless, emerging evidence suggests ERK5's unique functional characteristics [10]. Recent studies have revealed distinctive features of the ERK5 pathway: ERK5 is a key factor to inhibit endothelial inflammation [10] and has a key role in cardiovascular development [11]. Intriguingly, ERK5 is strongly activated by steady laminar flow (s-flow) that generates a frictional dragging force on the endothelium surface (called fluid shear stress), which is known to possess anti-inflammatory and antiatherosclerotic effects and to protect endothelial cells (ECs) from becoming dysfunctional [12], [13]. However, whether pharmaceutical stimulated ERK5 activation has a similar anti-inflammatory vasoprotective effect is not clear.
The accumulation of data implicating ERK5 as a key factor to inhibit endothelial inflammation prompted us to analyze the effect of statins on ERK5 activation and cross-talk between ERK5 and TNF inflammatory cascade in human endothelial cells. We found that TNF significantly increases NF-κB activity and VCAM-1 and ICAM-1 expression via Rac-1 activation and ROS generation. We provide evidence that statins can directly activate ERK5 and potently block TNF-induced Rac-1/ROS/NF-κB/VCAM-1 inflammation pathway. Therefore, the ERK5 activation and subsequent inhibition of TNF inflammation pathway in endothelial cells mediate statins-elicited anti-inflammatory vasoprotective effect.
Section snippets
Materials
Antibodies used in the present study and their commercial sources were as follows: anti-Rac-1, anti-IκB and anti-ERK5 (EMD Millipore Corporation, Billerica, MA, USA); anti-PMCA4 (plasma membrane calcium ATPase 4) (Sigma–Aldrich, St. Louis, MO, USA); anti-VCAM-1, ICAM-1 (OriGene Technologies, Rockville, MD, USA). Rac-1 inhibitor was purchased from Calbiochem (San Diego, CA, USA); PEG-catalase were purchased from Sigma; BAY 11-7085 and XMD 8-92 were from Santa Cruz Biotechnology (Santa Cruz, CA,
Statins inhibits TNF-induced Rac-1 activation and ROS formation in cultured ECs
To investigate whether TNF-α induces the liberation of ROS in HAECs, intracellular ROS levels were determined by DCF fluorescence. Stimulation of the HAECs with TNF-α (100 U/ml) led to a time-dependent increase of intracellular ROS, which was maximally 3.2-fold over control at 1 h (Fig. 1A). Recent research focused on the molecular and biochemical characterization of TNF-α-induced ROS production in endothelial cells. Several authors proved the role of the small GTPase Rac-1 in this context. Rac-1
Discussion
In the present study, we reveal the newest member of the MAPK family ERK5 as a critical mediator of the endothelial vasoprotective phenotype conferred by statins. Our data demonstrate that statins activate ERK5 in cultured human ECs, and importantly, that this activation is necessary for the statins-mediated decrease in TNF-activated expression of adhesion molecules such as VCAM-1 and ICAM-1, important inflammatory factors in ECs. Additionally, we have shown that the inhibition of Rac-1
Disclosures
None.
Acknowledgment
This work was supported by American Heart Association award – AHA Award #10POST3530033 to Y. Wu.
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2018, Advanced Drug Delivery ReviewsCitation Excerpt :GTPase family members Rac and RhoA, initiate downstream signalling that involves numerous intermediates (Fig. 3), most notably via the MAPK/NF-κB signalling axis to initiate synthesis of pro-inflammatory cytokines TNF-α, IL-6, IL-8 and pro-inflammatory intermediates intracellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 to name a few [29, 32, 53, 61, 63–66]. Importantly, cells treated with a variety of hydrophilic and lipophilic statins treatment show decreased Rac/RhoA-activity and therefore decreased downstream cytokine synthesis and secretion [32, 61, 65]. Additional studies have then further confirmed the role of prenylated GTPases play in regulating cytokine synthesis as chemical inhibition of prenylation reported similar results [18, 29, 30, 62, 67].