Endothelial Microparticles from Acute Coronary Syndrome Patients Induce Premature Coronary Artery Endothelial Cells Ageing and Thrombogenicity: Role of the Ang II/AT1 Receptor/NADPH Oxidase-mediated Activation of MAPKs and PI3-kinase Pathways
Background—Microparticles (MPs) have emerged as a surrogate marker of endothelial dysfunction and cardiovascular risk. This study examined the potential of MPs from senescent endothelial cells (ECs) or from patients with acute coronary syndrome (ACS) to promote premature ECs ageing and thrombogenicity.
Methods—Primary porcine coronary ECs were isolated from the left circumflex coronary artery. MPs were prepared from ECs and venous blood from patients with ACS (n=30) and from healthy volunteers (n=6) by sequential centrifugation. The level of endothelial senescence was assessed as senescence-associated beta-galactosidase (SA-β-gal) activity using flow cytometry, oxidative stress using the redox-sensitive probe dihydroethidium, Tissue factor activity using an enzymatic Tenase assay, the level of target protein expression by Western blot analysis, platelet aggregation using an aggregometer, and shear stress using a cone-and-plate viscometer.
Results—Senescence, as assessed by SA-β-gal activity, was induced by passaging of porcine coronary artery ECs from passage P1 to P4, and associated with a progressive shedding of procoagulant MPs. Exposure of P1 ECs to MPs shed from senescent P3 cells or circulating MPs from ACS patients induced increased SA-β-gal activity, oxidative stress, early phosphorylation of MAPKs and Akt, and up-regulation of p53, p21 and p16. Ex vivo, the pro-senescent effect of ACS-MPs was evidenced only under low shear stress condition. Depletion of endothelial-derived MPs (EMPs) from ACS-MPs reduced the induction of senescence. Pro-senescent MPs promoted ECs thrombogenicity through tissue factor up-regulation, shedding of procoagulant MPs, eNOS down-regulation and reduced NO-mediated inhibition of platelet aggregation. These MPs exhibited angiotensin-converting enzyme (ACE) activity, and up-regulated AT1 receptors and ACE in P1 ECs. Losartan, an AT1 receptor antagonist, and inhibitors of either MAPKs or PI3-kinase prevented the MPs-induced endothelial senescence.
Conclusions—These findings indicate that EMPs from ACS patients induce premature endothelial senescence under atheroprone low shear stress and thrombogenicity through angiotensin II-induced redox-sensitive activation of MAPKs and PI3-kinase/Akt. They further suggest that targeting EMPs shedding and/or their bioactivity may be a promising therapeutic strategy to limit the development of an endothelial dysfunction post ACS.
- Received April 29, 2016.
- Revision received September 16, 2016.
- Accepted October 19, 2016.