MiR-33 regulation of stretch-induced intimal hyperplasia in vein grafts.
MiR-33 regulation of stretch-induced intimal hyperplasia in vein grafts.
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DOI：
10.1093/cvr/cvx038
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发表时间：
2017-04
影响因子：
10.8
通讯作者：
Xinbo Zhang;C. Fernández-Hernando
中科院分区：
文献类型：
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作者：
Xinbo Zhang;C. Fernández-Hernando
静脉搭桥手术已成为冠状动脉疾病患者最常用的血运重建技术，冠状动脉疾病是世界范围内死亡率和发病率的主要原因。静脉移植物适应新的动脉环境，移植物血管壁的结构性血管重构和内膜增厚是血管重建后再狭窄的主要原因。平滑肌细胞（SMCs）的内膜积聚通过促炎细胞因子诱导的细胞迁移和局部细胞增殖，有助于血管腔的增厚和变窄。SMCs的增殖在内膜增生的发病机制中起着至关重要的作用，这被认为是静脉移植适应成功的重要决定因素。虽然疾病过程已被描述，但潜在的机制仍不清楚。过去十年的研究已经揭示了非编码rna在包括静脉移植旁路失败在内的几种心血管疾病中的重要作用。MicroRNAs （miRNAs）是高度保守的非编码小RNA分子，在转录后水平参与基因表达的调控。Huang和同事发现microRNA-33 （miR-33）是静脉移植物中SMC增殖和内膜增生的主要调节因子。Huang及其同事发现miR-33在移植静脉中的表达明显减弱。作者观察到miR-33水平与内膜增厚和SMC增殖增加呈负相关。为了确定miR-33是否直接控制SMC增殖，作者进行了一系列细致的研究，包括BrdU掺入和CCK-8测定。他们发现miR-33过表达明显抑制SMC增殖。相反，拮抗miR-33可增强SMC增殖。这些发现与先前的报道一致，即miR-33是细胞增殖和细胞周期进程的重要调节因子。为了剖析miR-33控制SMCs增殖的分子机制，Huang等人使用多种计算算法分析了miR-33预测的靶mrna。他们发现骨形态发生蛋白3 （BMP3）是一种新的miR-33靶基因。BMP3是转化生长因子β (TGF-b)超家族的成员，通过TGF-b/Activin信号通路促进间充质干细胞增殖。作者发现BMP3在移植静脉中的表达显著上调，而miR-33的表达则相反。他们还利用获得和丧失功能的方法来证明外源性BMP3加速静脉SMC增殖，而BMP3的敲除则表现出相反的效果。最重要的是，外源性BMP3消除了miR-33对SMC增殖的抑制作用。进一步的研究表明，BMP3下游的两个分子SMAD2和SMAD5的磷酸化受miR-33以依赖BMP3的方式调控。总之，这些观察结果支持了miR-33通过抑制BMP3保护SMCs增殖和新生内膜增生的假设。作者还分析了miR-33在静脉SMC增生中响应机械循环拉伸的功能，机械循环拉伸是影响SMC结构组织、功能和基因表达的主要机械力。与体内静脉移植模型一致，体外循环拉伸降低miR-33的表达，同时升高BMP3的表达，增加SMAD2和SMAD5的磷酸化。通过用miR-33模拟物或BMP3特异性siRNA处理SMCs，作者进一步验证了miR-33和BMP3在响应循环拉伸的静脉SMC增殖中的重要作用。值得注意的是，注射agomiR-33通过调节BMP3的表达和SMAD2和SMAD5的磷酸化，减少了移植静脉的新生内膜形成，抑制了细胞增殖。正如预期的那样，使用慢病毒过表达BMP3否定了agomiR-33对静脉移植物内膜增厚的影响，这表明miR-33对静脉SMC增殖和新生内膜增生的影响依赖于BMP3的表达。值得注意的是，每个microRNA可以调控多个靶mRNA，每个靶mRNA也可以被多个microRNA调控。先前，miR-33已被证明在细胞增殖和细胞周期进程中发挥重要作用，通过靶向细胞周期蛋白依赖性激酶6 （CDK6）、细胞周期蛋白D1和p53，控制肝细胞增殖、小鼠胚胎成纤维细胞的复制性衰老和造血干细胞的自我更新。CDK6是一种d -cyclin激活的激酶，在细胞周期的G1期通过与细胞周期蛋白D1、D2和D3的相互作用来驱动细胞周期，而p53通过调节p21的表达来诱导细胞周期的G1停滞。这些靶基因是否参与mir -33依赖性的机械拉伸诱导的增殖调控
Vein graft bypass surgery has become the most commonly performed revascularization technique in patients with coronary artery disease, the leading cause of mortality and morbidity worldwide. Vein grafts adapt to the new arterial environment, and the structural vascular remodelling and intimal thickening in the vein graft wall is the main cause of restenosis after vascular reconstruction. Intimal accumulation of smooth muscle cells (SMCs) contributes to the thickening and narrowing of the vessel lumen through pro-inflammatory cytokine-induced cell migration and local cell proliferation. Proliferation of SMCs is a crucial event in the pathogenesis of intimal hyperplasia, which is thought to be an important determinant of successful vein graft adaptation. Although the disease process has been described, the underlying mechanisms are still unclear. Work over the last decade has uncovered prominent roles for noncoding RNAs in several cardiovascular disorders including the failure of vein graft bypass. MicroRNAs (miRNAs) are highly conserved small non-coding RNA molecules involved in the regulation of gene expression at the post-transcriptional level. Huang and co-workers identified microRNA-33 (miR-33) as a major regulator of SMC proliferation and neointimal hyperplasia in vein grafts. Huang and colleagues found that miR-33 expression was markedly attenuated in grafted veins. The authors observed an inverse correlation between miR-33 levels and increased intimal thickening and SMC proliferation. To determine whether miR-33 directly controls SMC proliferation, the authors performed a series of elegant studies, including BrdU incorporation and CCK-8 assays. They found that miR-33 overexpression markedly inhibited SMC proliferation. By contrast, antagonism miR-33 enhances SMC proliferation. These findings are consistent with previous reports establishing miR-33 as an important regulator of cell proliferation and cell cycle progression. To dissect the molecular mechanisms by which miR-33 controls SMCs proliferation, Huang et al. analysed miR-33-predicted targets mRNAs using a number of computational algorithms. They identified bone morphogenetic protein 3 (BMP3) as a novel miR-33 target-gene. BMP3 is a member of the transforming growth factor beta (TGF-b) superfamily and promotes mesenchymal stem cell proliferation though the TGF-b/Activin signalling pathway. The authors found significantly upregulated BMP3 expression in grafted veins, while miR-33 showed an opposite regulation. They also utilized gainand lossof function approaches to demonstrate that exogenous BMP3 accelerated venous SMC proliferation, whereas knock-down of BMP3 exhibited the opposite effect. Most importantly, exogenous BMP3 abolished the inhibitory effects of miR-33 on SMC proliferation. Further studies showed that the phosphorylation of SMAD2 and SMAD5, two molecules downstream of BMP3, were regulated by miR-33 in a BMP3-dependent manner. Together, these observations support the hypothesis that miR-33 protects SMCs proliferation and neointimal hyperplasia by repressing BMP3. The authors also analysed the function of miR-33 in venous SMC proliferation in response to mechanical cyclic stretch, the predominant mechanical force influencing SMCs structural organization, function, and gene expression. Consistent with the in vivo vein graft model, cyclic stretch decreased the expression of miR-33 accompanied by elevated BMP3 expression and increased phosphorylation of SMAD2 and SMAD5 in vitro. By treating the SMCs with miR-33 mimics or BMP3 specific siRNA, the authors further validated the important role of miR-33 and BMP3 on venous SMC proliferation in response to cyclic stretch. Notably, injection of agomiR-33 attenuated neointimal formation and repressed cell proliferation in grafted veins by regulating BMP3 expression and phosphorylation of SMAD2 and SMAD5. As expected, BMP3 overexpression using lentivirus negated the effects of agomiR-33 on intimal thickening occurring in the vein grafts, suggesting the effects of miR-33 on venous SMC proliferation and neointimal hyperplasia are dependent on BMP3 expression. It is worth noting that each microRNA can regulate multiple target mRNAs and each target mRNA can also be regulated by multiple microRNAs. Previously, miR-33 has been demonstrated to play an important role in the regulation of cell proliferation and cell cycle progression by targeting cyclin-dependent kinase 6 (CDK6), cyclin D1, and p53, by which control hepatocyte proliferation, replicative senescence of mouse embryonic fibroblasts and haematopoietic stem cell self-renewal. CDK6 is a D-cyclin-activated kinase involved in driving the cell cycle through interactions with cyclins D1, D2, and D3 in G1 phase of the cell cycle, while p53 induces G1 arrest in the cell cycle by regulating p21 expression. Whether these target genes are involved in miR-33dependent regulation of mechanical stretch-induced proliferation of