F. Jansen

Endothelial Microparticle–Mediated Transfer of MicroRNA-126 Promotes Vascular Endothelial Cell Repair via SPRED1 and Is Abrogated in Glucose-Damaged Endothelial Microparticles

Background— Repair of the endothelium after vascular injury is crucial for preserving endothelial integrity and preventing the development of vascular disease. The underlying mechanisms of endothelial cell repair are largely unknown. We sought to investigate whether endothelial microparticles (EMPs), released from apoptotic endothelial cells (ECs), influence EC repair. Methods and Results— Systemic treatment of mice with EMPs after electric denudation of the endothelium accelerated reendothelialization in vivo. In vitro experiments revealed that EMP uptake in ECs promotes EC migration and proliferation, both critical steps in endothelial repair. To dissect the underlying mechanisms, Taqman microRNA array was performed, and microRNA (miR)-126 was identified as the predominantly expressed miR in EMPs. The following experiments demonstrated that miR-126 was transported into recipient human coronary artery endothelial cells by EMPs and functionally regulated the target protein sprouty-related, EVH1 domain-containing protein 1 (SPRED1). Knockdown of miR-126 in EMPs abrogated EMP-mediated effects on human coronary artery endothelial cell migration and proliferation in vitro and reendothelialization in vivo. Interestingly, after simulating diabetic conditions, EMPs derived from glucose-treated ECs contained significantly lower amounts of miR-126 and showed reduced endothelial repair capacity in vitro and in vivo. Finally, expression analysis of miR-126 in circulating microparticles from 176 patients with stable coronary artery disease with and without diabetes mellitus revealed a significantly reduced miR-126 expression in circulating microparticles from diabetic patients. Conclusions— Endothelial microparticles promote vascular endothelial repair by delivering functional miR-126 into recipient cells. In pathological hyperglycemic conditions, EMP-mediated miR-126–induced EC repair is altered.

High glucose condition increases NADPH oxidase activity in endothelial microparticles that promote vascular inflammation

AIMSnDiabetes is a major risk factor for cardiovascular diseases. Circulating endothelial microparticles (EMP) are increased in diabetic patients, but their potential contribution in atherogenesis is unclear. We sought to determine the role of EMP derived under high glucose conditions in the development of atherosclerosis.nnnMETHODS AND RESULTSnEMP were generated from human coronary endothelial cells (HCAEC) exposed to high glucose concentrations in order to mimic diabetic conditions. These EMP were defined as injured EMP (iEMP) and their effects were compared with EMP generated from healthy untreated HCAEC. iEMP injection significantly impaired endothelial function in ApoE(-/-) mice compared with EMP and vehicle treatment. Immunofluorescent experiments showed increased macrophage infiltration and adhesion protein expression in atherosclerotic lesions of iEMP-treated ApoE(-/-) mice compared with controls. To further investigate the underlying mechanism of iEMP-induced vascular inflammation, additional in vitro experiments were performed. iEMP, but not EMP, induced activation of HCAEC in a time- and dose-dependent manner and increased monocyte adhesion. Further experiments demonstrated that iEMP induced activation of HCAEC by phosphorylation of p38 into its biologically active form phospho-p38. Inhibition of p38 activation abrogated iEMP-dependent induction of adhesion proteins and monocyte adhesion on HCAEC. Moreover, we could demonstrate that iEMP show increased NADPH oxidase activity and contain significantly higher level of reactive oxygen species (ROS) than EMP. iEMP triggered ROS production in HCAEC and thereby activate p38 in an ROS-dependent manner.nnnCONCLUSIONnHigh glucose condition increases NADPH oxidase activity in endothelial microparticles that amplify endothelial inflammation and impair endothelial function by promoting activation of the endothelium. These findings provide new insights into the pathogenesis of diabetes-associated atherosclerosis.

Endothelial microparticles reduce ICAM-1 expression in a microRNA-222-dependent mechanism

Endothelial microparticles (EMP) are released from activated or apoptotic endothelial cells (ECs) and can be taken up by adjacent ECs, but their effect on vascular inflammation after engulfment is largely unknown. We sought to determine the role of EMP in EC inflammation. In vitro, EMP treatment significantly reduced tumour necrosis factor‐α‐induced endothelial intercellular adhesion molecule (ICAM)‐1 expression on mRNA and protein level, whereas there was no effect on vascular cell adhesion molecule‐1 expression. Reduced ICAM‐1 expression after EMP treatment resulted in diminished monocyte adhesion in vitro. In vivo, systemic treatment of ApoE−/− mice with EMP significantly reduced murine endothelial ICAM‐1 expression. To explore the underlying mechanisms, Taqman microRNA array was performed and microRNA (miR)‐222 was identified as the strongest regulated miR between EMP and ECs. Following experiments demonstrated that miR‐222 was transported into recipient ECs by EMP and functionally regulated expression of its target protein ICAM‐1 in vitro and in vivo. After simulating diabetic conditions, EMP derived from glucose‐treated ECs contained significantly lower amounts of miR‐222 and showed reduced anti‐inflammatory capacity in vitro and in vivo. Finally, circulating miR‐222 level was diminished in patients with coronary artery disease (CAD) compared to patients without CAD. EMPs promote anti‐inflammatory effects in vitro and in vivo by reducing endothelial ICAM‐1 expression via the transfer of functional miR‐222 into recipient cells. In pathological hyperglycaemic conditions, EMP‐mediated miR‐222‐dependent anti‐inflammatory effects are reduced.

Circulating Microparticles Decrease After Cardiac Stress in Patients With Significant Coronary Artery Stenosis.

Cardiac stress leads to a dynamic increase of circulating microparticles (MPs) in healthy individuals that is diminished in individuals with vascular disease. The impact of coronary ischemia on circulating MP level is unknown. This study investigates the kinetics of circulating MPs during cardiac stress in patients with coronary artery stenosis.

MicroRNA-126-containing endothelial microparticles reduce neointimaformation and vascular smooth muscle cell proliferation

Background: Vascular Smooth Muscle Cell (VSMC) proliferation is of importance in the pathogenesis of vascular diseases such as restenosis or atherosclerosis. Endothelial Microparticles (EMP) have been shown to promote vascular regenration by transferring functional microRNAs, but whether they influence VSMC biology is largely unknown. We explored the influence of EMP on neointima formation in a model of acute vascular injury in vivo and on VSMC proliferation and migration in vitro.nnMethods and results: After wire injury of the carotid artery, mice were treated systemically with EMP generated from Human Coronary Artery Endothelial Cells (HCAEC). Mice treated with EMP showed a significantly reduced neointimal formation (0.63±0.03 vs. 0.47±0.04,p<0.05, n=8). Furthermore, VSMCs treated with EMPs showed significantly reduced proliferation and migration capacities, both critical steps in neointimaformation (p<0.05 after 6h, p<0.01 after 8 h, n=7-8). Following experiments revealed a time dependent uptake of EMP into VSMCs in vitro. In order to dissect the underlying mechanisms, Taqman microRNA-array was performed and microRNA (miR)-126 was identified as the predominantly expressed miR in EMP. Furthermore, miR-126 was transported into recipient VSMC by EMP (1±0.05 vs. 1.34±0.05, p<0.05). Interestingly, expression of the target protein LRP6, regulating VSMC proliferation, was altered in VSMCs after EMP treatment.nnConclusions: Endothelial microparticles reduce neointimal formation in a model of acute arterial injury in vivo and decrease proliferation and migration of vascular smooth muscle cells in vitro. The transfer of mir126 by EMP and subsequent regulation of LRP6 expression in VSMCs might be a possible pathway.