Effect Of Monocytic Extracellular Vesicles On Proliferation Of Human Vascular Smooth Muscle Cells

Recently,cardiovascular diseases are severely threatening the health of human beingswith a noticeable increase in its incidence. However, the pathology of the cardiovascularsystem remained elusive. It is widely accepted proliferation of vascular smooth musclecells(VSMCs) is crucial in pathophysiological of cardiovascular diseases such asatherosclerosis（AS） and essential hypertension. In normal physiological conditions,VSMCs exsit in media of vessel wall.While in patients with AS and other related diseases,VSMCs migrated to the intima from the media and then began to proliferate, leading tovarious pathological consequenses including intimal thickening and vascular stenosis. Abetter understanding of pathogenesis of VSMCs proliferation may assist in developingbetter strategies in treating AS and reduce cardiovascular diseases risk.In recent years, more and more researches have shown that inflammation is closelyassociated with cardiovascular diseases, which are considered as chronic low-gradeinflammation or subclinical inflammation diseases. Inflammation plays an important role inthe occurrence and development of cardiovascular diseases and related target-organdamages in patients. Monocyte/macrophage cells are the main sources of inflammatorymediators. Studies show that monocyte/macrophage cells are involved in the proliferationof VSMCs. In inflammation, the activated circulatory monocytes/macrophage cells cangenerate a variety of cytotoxins, which lead to vascular endothelial injury, and then inducedthe platelet adhesion, aggregation and activation. The activated platelets release variouscytokines and growth factors (such as:PDGF,TGF-β), which initiate the final proliferation ofVSMCs. Subsequent studies have found that VSMCs migrate to the intima from the mediaand proliferate taken place before platelet adhesion and endothelial cell damage in thesepsis model of rabbits. Except for cytokines and growth factors, whether monocyte/macrophage cells promote VSMCs’proliferationbyother means?Nowadays, extracellular vesicles(EVs) was found played an important role in the cell- cell communication. EVs, a kind of microvesicles derived from various cells inphysiological and pathological conditions, containing a various proteins and nucleic acidseven DNA, have a potential to selectively interact with specific cells. The signal molecularsin EVs are relatively stable in normal consition. However, in certain diseases, EVs wassignificantly secreted more from different cells. In patients with hypertension, endothelialcells and monocyte-derived EVs were highly increased, and in the mean while, themoleculars in EVs also rised and experienced a positive relative with the evlation of bloodpressure. The contents in EVs increased in blood vessel wall of of atherosclerotic rat modeland accompanied with a markedly vascular inflammation, oxidative stress and endothelialdysfunction. Monocytic cells is an important source of EVs in the body. As the VSMCsis involing in the pathogenesis of cardiovascular disease, whether the monocyte-derivedEVs promote VSMCs proliferation is still poorly understood. We speculate that EVssecreted from monocyte cells may act on VSMCs and promote VSMCs proliferation, andthen participate in the development and progression of cardiovascular diseases.In this study, we are to investigate the pro-proliferation effect of monocyte-derivedEVs on human aortic smooth muscle cells (HA-VSMCs), and its potential mechanism.Objective：To investigate the effect of EVs derived from monocytes on the the proliferation ofVSMCs and the mechanism of this effect.Method：1. The culture of human aortic smooth muscle cells (HA-VSMCs) and human acutemonocytic leukemia cell line(THP-1).2. EVs derived from monocyte cell culture supernatants were isolated through a seriesof ultracentrifugation steps as previously described(1,2).3. Electron microscopic, immunoblotting, and flow cytometry (FCM)(3)analyses wereused to ensure EVs were correctly identified.4. Stimulated the monocytes for lipopolysaccharide（LPS）to trigger the immunologicresponse and then isolated EVs from monocytes.5. The proliferation of VSMCs were examined by [3H]-TdR and CCK-8experimentsafter EVs were co-cultured with VSMCs.6. Using immunoblotting to detect the machanism of the proliferation effect caused by EVs derived from monocyte cell. The mitogen-activated protein kinases(MAPKs) signalingpathway phosphorylation and the expression of c-jun and c-fos were tested.Results：1. Monocyte-derived promoted the proliferation of VSMCs in a concentration-andtime-dependent manner. EVs from monocytes pretreated with LPS have significantlystronger effects on VSMCs proliferation than from untreated monocytes.2. The mechanism of EVs induced proliferation of VSMCs is mediated by activatingthe phosphorylation of ERK, JNK and P38, and increase the expressions of c-fos and c-junin VSMCs.Conclusions：EVs from monocytes can promote the proliferation of VSMCs. EVs fromLPS-pretreated monocytes have significantly stronger effects on VSMCs proliferation thanfrom unpretreated monocytes, which act through the activation of MAPKs pathways.