Large Extracellular Vesicle-Associated Rap1 Accumulates in Atherosclerotic Plaques, Correlates With Vascular Risks and Is Involved in Atherosclerosis

Abstract

Rationale: Metabolic syndrome (MetS) is a cluster of interrelated risk factors for cardiovascular diseases and atherosclerosis. Circulating levels of large extracellular vesicles (lEVs), submicrometer-sized vesicles released from plasma membrane, from MetS patients were shown to induce endothelial dysfunction but their role in early stage of atherosclerosis and on vascular smooth muscle cells (SMC) remain to be fully elucidated. Objective: To determine the mechanisms by which lEVs lead to the progression of atherosclerosis in the setting of MetS. Methods and Results: Proteomic analysis revealed that the small GTPase, Rap1 was overexpressed in lEVs from MetS patients compared to those from non-MetS (nMetS) subjects. Rap1 was in GTP-associated active state in both types of lEVs and Rap1-lEVs levels correlated with increased cardiovascular risks including stenosis. MetS-lEVs, but not nMetS-lEVs, increased Rap1-dependent endothelial cell permeability. MetS-lEVs significantly promoted migration and proliferation of human aortic SMC and increased expression of pro-inflammatory molecules and activation of ERK5/p38 pathways. Neutralization of Rap1 by specific antibody or pharmacological inhibition of Rap1 with GGTi-298, completely prevented the effects of lEVs from MetS patients. HFD-fed ApoE -/- mice displayed an increased expression of Rap1 both in aortas and circulating lEVs. lEVs accumulated in plaque atherosclerotic lesions depending on the progression of atherosclerosis. lEVs from HFD-fed ApoE -/- mice, but not those from mice fed with a standard diet, enhanced SMC proliferation. Human atherosclerotic lesions were enriched in lEVs expressing Rap1. Conclusions: These data demonstrate that Rap1 carried by MetS-lEVs participates in the enhanced SMC proliferation, migration, pro-inflammatory profile and activation of ERK5/p38 pathways leading to vascular inflammation and remodeling, and atherosclerosis. These results highlight that Rap1 carried by MetS-lEVs may be a novel determinant of diagnostic value for cardiometabolic risk factors and suggest Rap1 as a promising therapeutic target against the development of atherosclerosis.

Extracellular vesicle signalling in atherosclerosis

“…Platelet-derived EVs can induce SMC proliferation and migration by inducing a proinflammatory phenotype in SMCs [ 162 , 163 ]. Proteomic analysis of EVs, revealed that Ras-related protein 1 (Rap1) was overexpressed in patients with metabolic syndrome compared to healthy donors [ 164 ]. A larger proportion of Rap1 + EVs originating from platelets, promoted SMC migration and proliferation probably via ERK5 activation in vitro and these effects were abolished after Rap1 inhibition [ 164 ].…”

“…Proteomic analysis of EVs, revealed that Ras-related protein 1 (Rap1) was overexpressed in patients with metabolic syndrome compared to healthy donors [ 164 ]. A larger proportion of Rap1 + EVs originating from platelets, promoted SMC migration and proliferation probably via ERK5 activation in vitro and these effects were abolished after Rap1 inhibition [ 164 ]. Elevated levels of Rap1 were detected on circulating EVs from high fat fed apoE −/− mice too [ 164 ].…”

“…A larger proportion of Rap1 + EVs originating from platelets, promoted SMC migration and proliferation probably via ERK5 activation in vitro and these effects were abolished after Rap1 inhibition [ 164 ]. Elevated levels of Rap1 were detected on circulating EVs from high fat fed apoE −/− mice too [ 164 ]. Evidence also suggests that they are responsible for monocyte recruitment to the atheroprone endothelium [ 165 ].…”

“…Elevated levels of EV-derived CD14 showed a strong correlation with vascular disease [ 208 ]. Lastly, circulating EVs from patients with cardiometabolic syndrome are enriched with Rap1 and could be a surrogate biomarker for early atherosclerosis detection [ 164 ]. Taken together these data suggest that EVs could be used as liquid biomarkers to monitor CVD progression.…”

Effects of REDOX in Regulating and Treatment of Metabolic and Inflammatory Cardiovascular Diseases

“…Atherosclerosis is one of the main causes of cardiovascular death in developed countries [17]. More and more evidence shows that oxidative stress plays a key role in the formation of atherosclerosis [44,45]. The activation of proinflammatory signal pathway, the expression of cytokines/chemokines, and the increase of oxidative stress are some of the mechanisms underlying atherosclerosis [20].…”