Qing Zhang1, Ting Chen1, Yun Zhang1, Lingxia Lyu1, Bohuan Zhang1, Chengchen Huang1, Xuhao Zhou1, Yutao Wu2, Zhoubin Li3. 1. Department of Cardiology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, 310003, Zhejiang Province, P.R. China. 2. Department of Cardiology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, 310003, Zhejiang Province, P.R. China. wuyutao@zju.edu.cn. 3. Department of Lung Transplantation, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, 310003, Zhejiang Province, P.R. China. 1510023@zju.edu.cn.
Abstract
BACKGROUND: As the most important component of the vascular wall, vascular smooth muscle cells (VSMCs) participate in the pathological process by phenotype transformation or differentiation from stem/progenitor cells. The main purpose of this study was to reveal the role and related molecular mechanism of microRNA-30c-5p (miR-30c-5p) in VSMC differentiation from adventitial progenitor cells expressing stem cell antigen-1(Sca-1). METHODS: In this study, we detected the expression of miR-30c-5p in human normal peripheral arteries and atherosclerotic arteries. In vitro, a stable differentiation model from adventitial Sca-1+ progenitor cells to VSMCs was established using transforming growth factor-β1 (TGF-β1) induction and the expression of miR-30c-5p during the process was observed. Then, we explored the effect of miR-30c-5p overexpression and inhibition on the differentiation from adventitial Sca-1+ progenitor cells to VSMCs. The target genes of miR-30c-5p were identified by protein chip and biological analyses and the expression of these genes in the differentiation process were detected. Further, the relationship between the target gene and miR-30c-5p and its effect on differentiation were evaluated. Finally, the co-transfection of miR-30c-5p inhibitor and small interfering RNA (siRNA) of the target gene was implemented to verify the functional target gene of miR-30c-5p during the differentiation from adventitial Sca-1+ progenitor cells to VSMCs, and the dual-luciferase reporter gene assay was performed to detect whether the mRNA 3'untranslated region (UTR) of the target gene is the direct binding site of miR-30c-5p. RESULTS: The expression of miR-30c-5p in the human atherosclerotic arteries was significantly lower than that in the normal arteries. During the differentiation from adventitial Sca-1+ progenitor cells to VSMCs, the expression of VSMC special markers including smooth muscle α-actin (SMαA), smooth muscle-22α (SM22α), smooth muscle myosin heavy chain (SMMHC), and h1-caponin increased accompanied with cell morphology changing from elliptic to fusiform. Meanwhile, the expression of miR-30c-5p decreased significantly. In functional experiments, overexpression of miR-30c-5p inhibited SMαA, SM22α, SMMHC, and h1-caponin at the mRNA and protein levels. In contrast, inhibition of miR-30c-5p promoted the expression of SMαA, SM22α, SMMHC, and h1-caponin. The target gene, osteoprotegerin (OPG), was predicted through protein chip and bioinformatics analyses. Overexpression of miR-30c-5p inhibited OPG expression while inhibition of miR-30c-5p had an opposite effect. Co-transfection experiments showed that low expression of OPG could weaken the promotion effect of miR-30c-5p inhibitor on the differentiation from adventitial Sca-1+ progenitor cells to VSMCs and the dual-luciferase reporter gene assay demonstrated that miR-30c-5p could target the mRNA 3'UTR of OPG directly. CONCLUSIONS: This study demonstrates that miR-30c-5p expression was significantly decreased in atherosclerotic arteries and miR-30c-5p inhibited VSMC differentiation from adventitial Sca-1+ progenitor cells through targeting OPG, which may provide a new target for the treatment of VSMCs-associated diseases.
BACKGROUND: As the most important component of the vascular wall, vascular smooth muscle cells (VSMCs) participate in the pathological process by phenotype transformation or differentiation from stem/progenitor cells. The main purpose of this study was to reveal the role and related molecular mechanism of microRNA-30c-5p (miR-30c-5p) in VSMC differentiation from adventitial progenitor cells expressing stem cell antigen-1(Sca-1). METHODS: In this study, we detected the expression of miR-30c-5p in human normal peripheral arteries and atherosclerotic arteries. In vitro, a stable differentiation model from adventitial Sca-1+ progenitor cells to VSMCs was established using transforming growth factor-β1 (TGF-β1) induction and the expression of miR-30c-5p during the process was observed. Then, we explored the effect of miR-30c-5p overexpression and inhibition on the differentiation from adventitial Sca-1+ progenitor cells to VSMCs. The target genes of miR-30c-5p were identified by protein chip and biological analyses and the expression of these genes in the differentiation process were detected. Further, the relationship between the target gene and miR-30c-5p and its effect on differentiation were evaluated. Finally, the co-transfection of miR-30c-5p inhibitor and small interfering RNA (siRNA) of the target gene was implemented to verify the functional target gene of miR-30c-5p during the differentiation from adventitial Sca-1+ progenitor cells to VSMCs, and the dual-luciferase reporter gene assay was performed to detect whether the mRNA 3'untranslated region (UTR) of the target gene is the direct binding site of miR-30c-5p. RESULTS: The expression of miR-30c-5p in the human atherosclerotic arteries was significantly lower than that in the normal arteries. During the differentiation from adventitial Sca-1+ progenitor cells to VSMCs, the expression of VSMC special markers including smooth muscle α-actin (SMαA), smooth muscle-22α (SM22α), smooth muscle myosin heavy chain (SMMHC), and h1-caponin increased accompanied with cell morphology changing from elliptic to fusiform. Meanwhile, the expression of miR-30c-5p decreased significantly. In functional experiments, overexpression of miR-30c-5p inhibited SMαA, SM22α, SMMHC, and h1-caponin at the mRNA and protein levels. In contrast, inhibition of miR-30c-5p promoted the expression of SMαA, SM22α, SMMHC, and h1-caponin. The target gene, osteoprotegerin (OPG), was predicted through protein chip and bioinformatics analyses. Overexpression of miR-30c-5p inhibited OPG expression while inhibition of miR-30c-5p had an opposite effect. Co-transfection experiments showed that low expression of OPG could weaken the promotion effect of miR-30c-5p inhibitor on the differentiation from adventitial Sca-1+ progenitor cells to VSMCs and the dual-luciferase reporter gene assay demonstrated that miR-30c-5p could target the mRNA 3'UTR of OPG directly. CONCLUSIONS: This study demonstrates that miR-30c-5p expression was significantly decreased in atherosclerotic arteries and miR-30c-5p inhibited VSMC differentiation from adventitial Sca-1+ progenitor cells through targeting OPG, which may provide a new target for the treatment of VSMCs-associated diseases.
Authors: Simon S Cross; Zhiyong Yang; Nicola J Brown; Sabapathy P Balasubramanian; Clair A Evans; Julia K Woodward; Helen L Neville-Webbe; Jenny M Lippitt; Malcolm W R Reed; Robert E Coleman; Ingunn Holen Journal: Int J Cancer Date: 2006-04-15 Impact factor: 7.396
Authors: Stefan Kiechl; Georg Schett; Gregor Wenning; Kurt Redlich; Martin Oberhollenzer; Agnes Mayr; Peter Santer; Josef Smolen; Werner Poewe; Johann Willeit Journal: Circulation Date: 2004-04-26 Impact factor: 29.690
Authors: N P E Kadoglou; T Gerasimidis; A Moumtzouoglou; A Kapelouzou; N Sailer; G Fotiadis; I Vitta; A Katinios; P Kougias; S Bandios; K Voliotis; P E Karayannacos; C D Liapis Journal: Eur J Vasc Endovasc Surg Date: 2008-04-18 Impact factor: 7.069