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，即血管损伤后由收缩表型向增生性表型转变并伴有新内膜形成，在一些增生性心血管疾病的发生和发展中起着关键作用，如动脉粥样硬化、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变。然而，SMC表型调节的调控机制尚不清楚。表型调节的一个显著特征是SMC收缩基因的下调。血小板衍生生长因子bb （platelet derived growth factor-BB, PDGF-BB）是一种众所周知的SMC表型调节刺激因子，它通过下调SMC基因表达，并通过转录后调控相关基因刺激SMC增殖。然而，SMC表型基因表达的转录后机制在很大程度上仍然未知。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pre-mRNA编辑是由作用于RNA的腺苷脱氨酶（ADAR）促进的，它将腺苷转化为肌苷（A->I编辑）。对内含子或3'-非翻译区pre-mRNA转录物的A-to-I RNA编辑会改变pre-mRNA剪接，导致成熟mRNA水平下降和细胞功能异常。PDGF-BB诱导ADAR1，同时下调SMC肌球蛋白重链（SMMHC）和钙钙蛋白（CNN）。shRNA敲低ADAR1可恢复pdgf - bb阻断的SMMHC和CNN的表达，表明ADAR1在SMC表型调节中起重要作用。ADAR1似乎对pdgf - bb诱导的SMC增殖/存活也很重要。体内研究表明，在球囊损伤大鼠颈动脉中，SMMHC和CNN pre-mRNA的成熟mRNA减少时会积累。此外，ADAR1最初在介质层SMCs中高度诱导，随后在损伤后的新生内膜SMCs中高度诱导。重要的是，敲低ADAR1可显著抑制损伤诱导的新生内膜形成，表明ADAR1在体内血管重构中起着关键作用。这些开创性的发现有力地支持了一种新的假设，即ADAR1/异常RNA编辑介导pdgf - bb诱导的SMC收缩基因下调和SMC增殖/存活，从而导致SMC表型调节和血管重塑。通过SMCs原代培养、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调控SMC增殖/存活的分子机制；3)在体内研究ADAR1在SMC表型调节和血管重构中的作用。这些目标的成功完成将揭示SMC表型调节的新机制，这将最终导致开发治疗增生性血管疾病的治疗药物的新靶点的鉴定。