Role Of MicroRNA-133b Regulated By IGF-1 On Low Shear Stress Induced Vascular Smooth Muscle Cells Proliferation

Vascular remodeling is mainly characterized as the abnormal proliferation,apoptosis and migration of vascular smooth muscle cells(VSMCs).Clinical and pathological studies have shown that atherosclerotic lesions occur preferentially at vessel branch points,bifurcations,and regions of high curvature,which suggests that low and disturbed shear stress is an important inducer for vascular remodeling and atherogenesis.Researches on the molecular mechanism of shear stress-induced vascular remodeling and the cross-talk between endothelial cells(ECs)and VSMCs can help to better understand the pathogenesis of cardiovascular disease.Our previous study showed that ECs modulate the proliferation of co-cultured VSMCs by secreting endothelial insulin-like growth factor-1(IGF-1)under shear stress.However,the role of microRNAs(miRs)in this process is still unclear.In the present study,miRs regulated by the IGF-1 were predicted by Ingenuity pathway analysis(IPA).Real-time PCR was used to examine the expression of miR-133b and miR-378a.After treated with or without recombinant IGF-1 in static situation,the expression of miR-133b and miR-378a in VSMCs was detected.ECs and VSMCs were then co-cultured and exposed to normal shear stress(NSS,15 dyn/cm~2)or low shear stress(LowSS,5 dyn/cm~2)for 12 h with parallel plate flow chamber system.Cell transfection and Real-time PCR were used to examine five putative targets of miR-133b predicted by websites,and finally polypyrimidine tract binding protein 1(Ptbp1)and N-myc downstream regulated 1(Ndrg1)were chosen for the further study.The protein levels of Ptbp1 and Ndrg1 in VSMCs were detected by western blotting after transfected with miR-133b mimics and inhibitor,or treated with recombinant IGF-1.In addition,the mRNA and protein levels of Ptbp1 and Ndrg1 in the co-cultured VSMCs were detected.The VSMC proliferation was detected by EdU flow cytometry assay.The results showed that:?After treated with recombinant IGF-1,the expression of miR-133b and miR-378a in VSMCs was increased.?Compared with NSS,LowSS significantly induced the expression of miR-133b in the co-cultured VSMCs,but had no effect on miR-378a.?In VSMCs,the protein and mRNA levels of Ptbp1 and Ndrg1 were down-regulated by miR-133b mimics.miR-133b inhibitor up-regulated the mRNA levels of Ptbp1 and Ndrg1,but had no remarkable effect on protein level.?After exposed to LowSS or treated with recombinant IGF-1,Ndrg1 expression in VSMCs was significantly depressed,but Ptbp1 had no remarkable change.?miR-133b mimics promoted the proliferation of VSMCs significantly.The present results indicated that miR-133b plays an important role in the proliferation of co-cultured VSMCs.IGF-1 secreted by ECs in response to LowSS may up-regulate the expression of miR-133b in the co-cultured VSMCs,which subsequently depress the expression of Ndrg1,and induce the proliferation of VSMCs eventually.Our research suggested that miR-133b,as a vital molecule,involved in the VSMC function regulated by shear stress and IGF-1,which provided a potential new target for cardiovascular disease therapy.