Atherosclerotic conditions promote the packaging of functional microRNA-92 into endothelial microvesicles

Abstract

Objectives: Microvesicle (MV)-incorporated microRNAs (miRs) are biomarkers and effectors of cardiovascular diseases. Whether MV-miRs expression is regulated in coronary artery disease (CAD) is unknown. We aimed to explore the expression of circulating MV-miRs in patients with CAD.<br /> Methods and results: Circulating MVs were isolated from patients’ plasma using ultracentrifugation. The electron microscope was used for MVs size characterization. Taqman miR array revealed that MV-miRs are significantly regulated in patients with stable CAD compared to ACS patients. To validate miR array results, 180 patients with angiographically excluded CAD (n=41), stable CAD (n=77) and acute coronary syndrome (ACS, n=62) were prospectively studied. Nine miRs involved in the regulation of vascular performance - miR-126, miR-222, miR-let7, miR-21, miR-26, miR-92, miR-133, miR-30 and miR-199 – were quantified in circulating MVs by real-time PCR. Among those, miR-92 was significantly increased in patients with CAD compared to non-CAD patients.<br /> MV-sorting experiments showed that endothelial cells (ECs) were the major cell source of MVs (EMVs) containing miR-92. In vitro, oxLDL stimulation dose-dependently increased miR-92 expression both in EMVs and ECs in a STAT3-dependent way. MiR-92 and EMV labeling demonstrated that functional miR-92 was transported into recipient ECs promoting EC migration and proliferation. Knockdown of miR-92 in EMVs abrogated EMV-mediated effects on EC migration and proliferation and blocked vascular network formation in matrigel plug. PCR-based gene profiling showed that the expression of THBS1, a target of miR-92 and an inhibitor of angiogenesis, was significantly reduced in ECs by EMVs. Knockdown of miR-92 in EMVs abrogated EMV-mediated inhibition of THBS1 gene and protein expression.<br /> Conclusion: Atherosclerotic conditions promote the packaging of endothelial miR-92 from ECs into EMVs. EMV-mediated transfer of functional miR-92 regulates angiogenesis in recipient ECs in a THBS1-dependent mechanism.