Extracellular Vesicles Secreted by Atherogenic Macrophages Transfer MicroRNA to Inhibit Cell Migration.

During inflammation, macrophages secrete vesicles carrying RNA, protein, and lipids as a form of extracellular communication. In the vessel wall, extracellular vesicles (EVs) have been shown to be transferred between vascular cells during atherosclerosis; however, the role of macrophage-derived EVs in atherogenesis is not known. Here, we hypothesize that atherogenic macrophages secrete microRNAs (miRNAs) in EVs to mediate cell-cell communication and promote proinflammatory and proatherogenic phenotypes in recipient cells. We isolated EVs from mouse and human macrophages treated with an atherogenic stimulus (oxidized low-density lipoprotein) and characterized the EV miRNA expression profile. We confirmed the enrichment of miR-146a, miR-128, miR-185, miR-365, and miR-503 in atherogenic EVs compared with controls and demonstrate that these EVs are taken up and transfer exogenous miRNA to naive recipient macrophages. Bioinformatic pathway analysis suggests that atherogenic EV miRNAs are predicted to target genes involved in cell migration and adhesion pathways, and indeed delivery of EVs to naive macrophages reduced macrophage migration both in vitro and in vivo. Inhibition of miR-146a, the most enriched miRNA in atherogenic EVs, reduced the inhibitory effect of EVs on macrophage migratory capacity. EV-mediated delivery of miR-146a repressed the expression of target genes IGF2BP1 (insulin-like growth factor 2 mRNA-binding protein 1) and HuR (human antigen R or ELAV-like RNA-binding protein 1) in recipient cells, and knockdown of IGF2BP1 and HuR using short interfering RNA greatly reduced macrophage migration, highlighting the importance of these EV-miRNA targets in regulating macrophage motility. EV-derived miRNAs from atherogenic macrophages, in particular miR-146a, may accelerate the development of atherosclerosis by decreasing cell migration and promoting macrophage entrapment in the vessel wall.

Nguyen MA ，Karunakaran D ，Geoffrion M ，Cheng HS ，Tandoc K ，Perisic Matic L ，Hedin U ，Maegdefessel L ，Fish JE ，Rayner KJ ... -  《-》

Hepatocyte-derived extracellular vesicles promote endothelial inflammation and atherogenesis via microRNA-1.

Clinical evidence has indicated a close link between non-alcoholic fatty liver disease (NAFLD) and cardiovascular disease (CVD). However, the underlying mechanism remains to be elucidated. This study aimed to explore a potential role of hepatocyte-derived extracellular vesicles (EVs) in endothelial inflammation and atherogenesis in the context of NAFLD. EVs were isolated, quantified and characterized from steatotic hepatocytes. An endothelial cell-specific PCR array was used to screen the functional properties of EVs. Profiling of global microRNA expression was conducted in EVs. The expression level and biological function of microRNA-1 (miR-1) was determined by quantitative PCR, immunoblot and reporter gene assays, respectively. The in vivo effect of miR-1 on atherogenesis was investigated in apolipoprotein E (ApoE)-deficient mice administered with a miR-1-specific inhibitor, antagomiR-1. Steatotic hepatocytes released more EVs, which had significantly altered miRNA expression profiles compared to the EVs released by control hepatocytes. Endothelial cells co-cultured with steatotic hepatocytes, or treated with their EVs or miR-1, expressed significantly more proinflammatory molecules, as well as exhibiting increased NF-κB activity and reduced Kruppel-like factor 4 (KLF4) expression. EV-induced endothelial inflammation was prevented by either downregulation or inhibition of miR-1. While miR-1 treatment suppressed KLF4 expression and reporter gene activity, overexpression of KLF4 dramatically abolished the miR-1-induced endothelial inflammation. Moreover, not only did the miR-1 inhibitor reduce endothelial inflammation in vitro, but it also attenuated atherogenesis in ApoE-deficient mice. Steatotic hepatocyte-derived EVs promote endothelial inflammation and facilitate atherogenesis by miR-1 delivery, KLF4 suppression and NF-κB activation. The findings illustrate an important role of hepatocyte-derived EVs in distant communications between the liver and vasculature, suggesting a new mechanism underlying the link between NAFLD and CVD. Non-alcoholic fatty liver disease (NAFLD), a condition highly prevalent in obese and/or diabetic patients, is emerging as an independent risk factor of cardiovascular disease. Herein, we demonstrated that extracellular vesicles, released by hepatocytes under NAFLD conditions, cause vascular endothelial inflammation and promote atherosclerosis. Within these toxic vesicles, we identified a small molecular cargo that acted as a potent inducer of endothelial inflammation. By inhibiting this cargo's function, a specific gene-based inhibitor profoundly attenuated atherogenesis in mice, uncovering a novel mechanism which may be used to prevent or treat cardiovascular disease in patients with NAFLD.

Jiang F ，Chen Q ，Wang W ，Ling Y ，Yan Y ，Xia P ... -  《-》

Extracellular-vesicle containing miRNA-503-5p released by macrophages contributes to atherosclerosis.

Wang Y ，Xu Z ，Wang X ，Zheng J ，Peng L ，Zhou Y ，Song Y ，Lu Z ... -  《Aging-US》

Extracellular vesicles derived from M1 macrophages deliver miR-146a-5p and miR-146b-5p to suppress trophoblast migration and invasion by targeting TRAF6 in recurrent spontaneous abortion.

Rationale: Emerging evidence demonstrates that insufficient migration and invasion of trophoblasts play critical roles in the pathogenesis of recurrent spontaneous abortion (RSA). Cell-to-cell communication at the maternal-fetal interface is essential to maintain the invasion and migration of trophoblasts. M1 macrophages, important immune cellular components at the maternal-fetal interface, have been reported to be elevated in decidua tissues from patients with RSA. Recent studies indicate that M1 macrophages modulate trophoblast biological behaviors; however, the underlying mechanisms remain poorly understood. Methods: Extracellular vesicles (EVs) were isolated from the supernatant of M1 macrophages inducted from THP-1 cells (M1-EVs) by ultracentrifugation, identified by transmission electron microscopy, nanoparticle tracking analysis, and western blotting, and their miRNA profile was characterized by miRNA sequencing. Scratch wound healing and transwell assays were used to investigate the effect of M1-EVs on trophoblast migration and invasion. RT-PCR, western blotting, and luciferase reporter assays were conducted to uncover the underlying mechanism. Finally, animal experiments were employed to explore the effect of M1-EVs on embryo absorption in mice. Results: M1 macrophages suppressed trophoblast EMT to reduce their migration and invasion abilities in vitro by secreting EVs. Through miRNA sequencing, miR-146a-5p and miR-146b-5p were identified as the most upregulated miRNAs in trophoblasts treated with M1-EVs. Further functional experiments showed that M1-EVs inhibited trophoblast migration and invasion by transferring miR-146a-5p and miR-146b-5p. Mechanistically, EV miR-146a-5p and miR-146b-5p inhibited EMT of trophoblasts by directly suppressing TNF receptor-associated factor 6 (TRAF6) expression at the post-transcriptional level. Furthermore, M1-EVs aggravated embryo absorption in mice. Clinically, expression of miR-146a-5p, miR-146b-5p, and TRAF6 were aberrant in placental villous tissues from patients with RSA, and negative correlations were found between miR-146a-5p/miR-146b-5p and TRAF6 expression levels. Conclusions: Our findings indicate that miR-146a-5p and miR-146b-5p derived from EVs play important roles in intercellular communication between M1 macrophages and trophoblasts, illuminating a novel mechanism in M1 macrophage regulation of trophoblasts and their role in RSA.

Ding J ，Zhang Y ，Cai X ，Zhang Y ，Yan S ，Wang J ，Zhang S ，Yin T ，Yang C ，Yang J ... -  《Theranostics》

M1 macrophages-derived extracellular vesicles elevate microRNA-185-3p to aggravate the development of atherosclerosis in ApoE(-/-) mice by inhibiting small mothers against decapentaplegic 7.

Extracellular vesicles (EVs) are vital mediators of transferring microRNAs (miRNAs). We focused on effect of miR-185-3p that mediated by macrophages-derived EVs on atherosclerosis (AS) by targeting small mothers against decapentaplegic 7 (Smad7). EVs were extracted from M1 macrophages and identified. ApoE-/- mice were treated with EVs, EVs containing miR-185-3p inhibitor or mimic, then the pathological changes of mouse aorta were observed. The levels of blood lipid, cell adhesion molecules, oxidative stress factors, inflammatory factors, and proliferation and apoptosis of vascular endothelial cells were assessed. Expression of miR-185-3p and Smad7 was detected and the targeting relationship between miR-185-3p and Smad7 was validated. MiR-185-3p was upregulated while Smad7 was downregulated in atherosclerotic mouse aorta. M1 macrophages-derived EVs elevated miR-185-3p to promote development of AS pathology and levels of blood lipid, endothelial cellular adhesion, oxidative stress factors and inflammatory factors, suppressed cell proliferation and promoted cell apoptosis of vascular endothelial cells in atherosclerotic mice through downregulating Smad7. Smad7 was a target gene of miR-185-3p and miR-185-3p could inhibit expression of Smad7. M1 macrophages-derived EVs and upregulated miR-185-3p aggravated the development of AS in ApoE-/- mice by negatively regulating Smad7. This research may further the understanding of AS mechanism.

Li K ，Cui M ，Zhang K ，Wang G ，Zhai S ... -  《-》