Novel mechanism of smooth muscle phenotypic modulation and vascular remodeling

平滑肌表型调节和血管重塑的新机制

• 批准号: 8653749
• 负责人: Shiyou Chen
• 金额: $ 37.25万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-01 至 2018-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8653749
• 关键词: 3' Untranslated Regions; ADAR1; Adenosine; Angioplasty; Ants; Arterial Disorder; Arteries; Atherosclerosis; Blood Platelets; Blood Vessels; Bypass; Cardiovascular Diseases; Carotid Arteries; Cell physiology; Contractile Proteins; Contracts; DRADA2b protein; Data; Development; Diabetic Angiopathies; Disease; Down-Regulation; Figs - dietary; Gene Expression; Genes; Genetic; Goals; Growth; Hyperplasia; Hypertension; Injury; Inosine; Introns; Knockout Mice; Lead; Mediating; Messenger RNA; Mining; Modeling; Molecular; Mus; Muscle; Myosin ATPase; Myosin Heavy Chains; Pathologic; Phenotype; Platelet-Derived Growth Factor; Play; Proteins; RNA Editing; RNA Splicing; Rattus; Regulation; Role; Seminal; Site; Smooth Muscle; Smooth Muscle Myocytes; Smooth Muscle Myosins; Steno; Testing; Therapeutic; Therapeutic Agents; Transcript; Translations; Transplantation; United States; Vascular Diseases; Vascular Proliferation; Vascular Smooth Muscle; Vascular remodeling; Veins; calponin; dsRNA adenosine deaminase; gain of function; in vivo; injured; loss of function; mRNA Precursor; migration; mortality; mouse model; neointima formation; novel; platelet-derived growth factor BB; protein expression; public health relevance; restenosis; small hairpin RNA; teration

DESCRIPTION (provided by applicant): Vascular smooth muscle (SMC) phenotypic modulation, the transition from a contractile to a proliferative phenotype accompanied by neointima formation following vascular injury, plays a critical role in the development and progression of several proliferative cardiovascular diseases such as atherosclerosis, hypertension, restenosis after angioplasty or bypass, diabetic vascular complications, and transplantation arteriopathy. The regulatory mechanisms underlying SMC phenotypic modulation, however, are poorly understood. A hallmark feature of the phenotypic modulation is the down-regulation of SMC contractile genes. Platelet-derived growth factor-BB (PDGF-BB), a well-known stimulator of SMC phenotypic modulation, down- regulates SMC gene expression and stimulates SMC proliferation via posttranscriptional regulation of the related genes. The post-transcriptional mechanisms involved in SMC phenotype gene expression, however, remain largely unknown. Our exciting preliminary data indicate that the down-regulation of SMC contractile genes is caused by abnormal RNA editing of their precursor mRNAs (pre-mRNAs). This abnormal pre-mRNA editing is facilitated by adenosine deaminase acting on RNA (ADAR), which converts adenosines to inosines (A->I editing). A-to-I RNA editing of the pre-mRNA transcripts from introns or 3'-untranslated regions alters pre- mRNA splicing, leading to decreased mature mRNA levels and abnormal cellular functions. PDGF-BB induces ADAR1 while down-regulating SMC myosin heavy chain (SMMHC) and calponin (CNN). Knockdown of ADAR1 by shRNA restores PDGF-BB-blocked SMMHC and CNN expression, demonstrating that ADAR1 plays an essential role in SMC phenotype modulation. ADAR1 appears to be also important for PDGF-BB-induced SMC proliferation/survival. In vivo studies show that SMMHC and CNN pre-mRNA is accumulated when their mature mRNA is decreased in balloon-injured rat carotid arteries. Moreover, ADAR1 is highly induced in media layer SMCs initially, and neointima SMCs subsequently following the injury. Of importance, knockdown of ADAR1 dramatically inhibits injury-induced neointima formation, demonstrating a critical role of ADAR1 in vascular remodeling in vivo. These seminal findings strongly support a novel hypothesis that ADAR1/abnormal RNA editing mediates PDGF-BB-induced down-regulation of SMC contractile genes and SMC proliferation/survival, leading to SMC phenotypic modulation and vascular remodeling. Using primary culture of SMCs, in vivo rat balloon injury and mouse wire injury models combining with molecular, cellular and histological approaches, we will 1) determine the role and mechanism whereby ADAR1 modulates SMC phenotype; 2) elucidate the molecular mechanisms underlying ADAR1 function in regulating SMC proliferation/survival; and 3) study the role of ADAR1 in SMC phenotypic modulation and vascular remodeling in vivo. Successful completion of these aims will unravel a novel mechanism governing SMC phenotypic modulation, which will ultimately lead to identification of novel targets for developing therapeuti agents to treat proliferative vascular diseases.

描述（由申请人提供）：血管平滑肌（SMC）表型调节，即从收缩表型到增殖表型的转变，伴随着血管损伤后新内膜的形成，在多种增殖性心血管疾病（例如动脉粥样硬化）的发生和进展中发挥着关键作用、高血压、血管成形术或搭桥术后再狭窄、糖尿病血管并发症和移植动脉病。然而，人们对 SMC 表型调节的调控机制知之甚少。表型调节的一个标志性特征是 SMC 收缩基因的下调。血小板源性生长因子-BB (PDGF-BB) 是一种众所周知的 SMC 表型调节刺激剂，下调 SMC 基因表达并通过相关基因的转录后调节刺激 SMC 增殖。然而，参与 SMC 表型基因表达的转录后机制仍然很大程度上未知。我们令人兴奋的初步数据表明，SMC 收缩基因的下调是由其前体 mRNA (pre-mRNA) 的异常 RNA 编辑引起的。这种异常的前 mRNA 编辑是通过作用于 RNA (ADAR) 的腺苷脱氨酶促进的，该酶将腺苷转化为肌苷（A->I 编辑）。对内含子或 3'-非翻译区的前 mRNA 转录物进行 A 到 I RNA 编辑会改变前 mRNA 剪接，导致成熟 mRNA 水平降低和细胞功能异常。 PDGF-BB 诱导 ADAR1，同时下调 SMC 肌球蛋白重链 (SMMHC) 和钙调蛋白 (CNN)。 shRNA 敲除 ADAR1 可恢复 PDGF-BB 阻断的 SMMHC 和 CNN 表达，表明 ADAR1 在 SMC 表型调节中发挥重要作用。 ADAR1 似乎对于 PDGF-BB 诱导的 SMC 增殖/存活也很重要。体内研究表明，当球囊损伤的大鼠颈动脉中 SMMHC 和 CNN 的成熟 mRNA 减少时，SMMHC 和 CNN 前体 mRNA 就会积累。此外，ADAR1 最初在中层 SMC 中高度诱导，随后在损伤后在新内膜 SMC 中高度诱导。重要的是，ADAR1 的敲除可显着抑制损伤诱导的新内膜形成，这表明 ADAR1 在体内血管重塑中发挥着关键作用。这些开创性的发现有力地支持了一个新的假设，即 ADAR1/异常 RNA 编辑介导 PDGF-BB 诱导的 SMC 收缩基因和 SMC 增殖/存活的下调，从而导致 SMC 表型调节和血管重塑。利用 SMC 的原代培养、体内大鼠球囊损伤和小鼠丝损伤模型，结合分子、细胞和组织学方法，我们将 1) 确定 ADAR1 调节 SMC 表型的作用和机制； 2）阐明ADAR1调节SMC增殖/存活的分子机制； 3)研究ADAR1在体内SMC表型调节和血管重塑中的作用。这些目标的成功实现将揭示一种控制 SMC 表型调节的新机制，最终将确定新的靶标，用于开发治疗增殖性血管疾病的治疗药物。