MiR-142-3p Delivered By Platelet Derived Microparticles Modulates Proliferation And Apoptosis Endothelial Cell During Hypertension

Endothelial cells(ECs)play an important role in the maintenance of vascular physiological homeostasis and pathological remodeling.It has been found that interaction between circulating platelets and ECs participates in the pathogenesis of vascular remodeling,but the molecular mechanism remains unclear.Recent studies revealed that the number of platelet-derived microparticles(PMPs)is significantly increased in hypertensive patients;PMPs directively act on ECs,which may be involved in the pathogenesis of EC dysfunction,and then induce vascular remodeling.Detect on the molecular mechanism of PMPs transfer biological information and regulate EC functions,may give a new insight into understanding the cardiovascular disease.Using microRNAs(miRs)array,the differential expression of miRs between platelets and ECs was firstly analyzed.Since the present study focused on the mechanism that platelets affect on ECs,we screened miRs which was highly expressed in platelets but defected in ECs.Real-time PCR revealed that miR-142-3p were the most pronounced molecule among the top 10 miRs showed by miR array.Hence,the further work was focused on miR-142-3p.The in vitro(thrombin activation)and in vivo(hypertensive rat model)experiments were performed to explore the molecular mechanism of EC dysfunction induced by hypertension.Thrombin was used to stimulate freshly extracted rat platelets,and real-time PCR was used to detect the expression of miRs in platelets and PMPs respectively.The results showed that after thrombin stimulation,miR-142-3p expression in platelets and PMPs was both significantly increased.Using live cell membrane dye PKH67,platelet and PMPs were labled,and the adhesion of platelets and PMPs to ECs was observed by fluorescence confocal microscopy.The results showed that there was a large amount of PMPs adhered to ECs surface after 1 hour incubation with platelet or PMPs.Real-time PCR showed that miR-142-3p could be effectively transferred to ECs after PMPs adhesion for 12 h.The potential target of miR-142-3p was identified using the miRs prediction softwares,and then mimics transfection,dual luciferase reporter gene,real-time PCR and western blotting were used to detect the expression of target genes,i.e.BCL2L1 and BCLAF1,in ECs.It revealed that miR-142-3p significantly reduce the expression of BCL2L1 and BCLAF1 in ECs.BrdU ELISA showed that PMPs and miR-142-3p mimics promoted the proliferation of ECs;AnnexinV/PI combined with flow cytometry showed that EC apoptosis was repressed by PMPs and miR-142-3p mimics.Furthermore,the changes of PMPs expression in the circulating blood were detected in the hypertensive animal model induced by abdominal aortic ligation in rats,and the expression of miR-142-3p in PMPs in the blood of hypertensive animals was detected.The results showed that PMPs in the blood were up-regulated and the proliferation of ECs was enhanced which may be caused by the increased expression of miR-142-3p in the ECs in hypertension.These results suggested that platelets release PMPs which may deliver miR-142-3p into ECs,target on BCL2L1 and BCLAF1,and then promote the proliferation and inhibit the apoptosis of ECs.PMPs and the contained miRs play an important role in the crosstalk between platelets and ECs.PMPs and the contained miRs may be the potential targets for the diagnosis,clinical treatment and evaluation of the therapeutic effects of hypertension.