Dedicator of Cytokinesis 2 in smooth muscle phenotype modulation

细胞分裂 2 在平滑肌表型调节中的奉献者

• 批准号: 8998055
• 负责人: Shiyou Chen
• 金额: $ 44.87万
• 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: 2014-04-01 至 2019-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8998055
• 关键词: Actins; Angioplasty; Animals; Arterial Disorder; Arteries; Asthma; Atherosclerosis; Attenuated; Binding; Blood Vessels; Bypass; Cardiovascular Diseases; Carotid Arteries; Catheters; Cell Nucleus; Cell Proliferation; Cell membrane; Cells; Contractile Proteins; Cytokinesis; Cytoskeleton; Data; Development; Diabetes Mellitus; Diabetic Angiopathies; Disease; Etiology; Family; Figs - dietary; Focal Adhesions; GKLF protein; Genes; Genetic Transcription; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate Phosphohydrolases; Health; Hematopoietic; Hyperplasia; Hypertension; Injury; Knock-out; Knockout Mice; Lead; Ligation; Lymphocyte; Malignant Neoplasms; Mediating; Modeling; Molecular; Mus; Pathologic; Phenotype; Physiological; Play; Process; Rattus; Role; Serum Response Factor; Smooth Muscle; Smooth Muscle Actin Staining Method; Smooth Muscle Myocytes; Stenosis; System; Testing; Therapeutic Agents; Time; Transcriptional Activation; Transcriptional Regulation; Transplantation; United States; Vascular Diseases; Vascular Smooth Muscle; Vascular remodeling; Vein graft; cell motility; gain of function; genetic approach; human disease; in vivo; insight; knock-down; loss of function; mRNA Expression; migration; mortality; mouse model; myocardin; neointima formation; new therapeutic target; novel; overexpression; platelet-derived growth factor BB; promoter; protein expression; restenosis; rho; small hairpin RNA; therapeutic target

DESCRIPTION (provided by applicant): The overall goal of this proposal is to establish a novel mechanism by which dedicator of cytokinesis 2 (DOCK2) regulates the phenotypic modulation of vascular smooth muscle cells (SMC). Transition of SMC from a differentiated phenotype to a dedifferentiated state accompanied by neointima formation/vascular remodeling plays a critical role in the development of atherosclerosis, restenosis after angioplasty or bypass, diabetic vascular complications, transplantation arteriopathy, asthma, and cancer. The mechanisms and factors that regulate SMC phenotypic modulation and neointima formation, however, are poorly understood. Under physiological conditions, DOCK2 is only expressed in hematopoietic cells and controls lymphocyte migration and activation by regulating actin cytoskeleton through Rac activation. Our exciting preliminary data demonstrate that platelet derived growth factor (PDGF)-BB, a SMC phenotype modulator, induced DOCK2 expression in SMC. Knockdown of DOCK2 blocked PDGF-BB-induced SMC phenotypic modulation, proliferation, and migration. In vivo animal studies showed that DOCK2 was undetectable in SMC of normal rat carotid arteries, but was induced in the media layer SMC initially and neointimal SMC subsequently following balloon catheter-induced vascular injury. Importantly, knockdown of DOCK2 dramatically inhibited the injury- induced neointima formation. The most conclusive evidence for DOCK2-dependent vascular remodeling/ neointima formation was that knockout of DOCK2 (DOCK2-/-) dramatically blocked artery ligation-induced neointima hyperplasia in mouse carotid artery. Interestingly, DOCK2 is localized in both cell membrane and nuclei of SMC, suggesting that in addition to its role in cytoskeleton/cell migration, DOCK2 may be involved in SMC gene transcription. Indeed, DOCK2 overexpression blocked both SMC gene mRNA expression and myocardin-induced activation of smooth muscle ¿-actin promoter. Thus, the central hypothesis is that DOCK2 regulates SMC phenotypic modulation by suppressing SMC gene transcription and stimulating SMC migration/ proliferation, leading to neointima formation/vascular remodeling. Using primary culture of SMC, in vivo rat balloon injury and mouse wire injury models combining with molecular, cellular and histological approaches, we will 1) study the molecular mechanisms by which DOCK2 modulates SMC phenotype through regulating SMC gene transcription; 2) investigate if DOCK2 induces SMC migration through activation of Rac/RhoA/ Cdc42; and 3) determine the essential role of DOCK2 in SMC phenotypic modulation and vascular remodeling in vivo. The completion of this project will unravel a novel mechanism regulating SMC phenotypic modulation and provide novel insights into whether DOCK2 is a potential therapeutic target for countering vascular damage associated with common diseases including diabetes, restenosis, atherosclerosis, and cancer.

描述（由申请人提供）：本提案的总体目标是建立一种新的机制，通过该机制，胞质分裂专用因子2（DOCK 2）调节血管平滑肌细胞（SMC）的表型调节。SMC从分化表型向去分化状态的转变伴随着新生内膜形成/血管重塑在动脉粥样硬化、血管成形术或旁路术后再狭窄、糖尿病血管并发症、移植动脉病、哮喘和癌症的发展中起关键作用。然而，调节SMC表型调节和新生内膜形成的机制和因素知之甚少。在生理条件下，DOCK 2仅在造血细胞中表达，并通过Rac激活调节肌动蛋白细胞骨架来控制淋巴细胞的迁移和激活。我们令人兴奋的初步数据表明，血小板衍生生长因子（PDGF）-BB，SMC表型调节剂，诱导DOCK 2在SMC中的表达。DOCK 2的敲低阻断PDGF-BB诱导的SMC表型调节、增殖和迁移。体内动物研究表明，DOCK 2在正常大鼠颈动脉SMC中检测不到，但在球囊导管诱导的血管损伤后，在最初的中膜SMC和随后的新生内膜SMC中诱导表达。重要的是，DOCK 2的敲低显著抑制损伤诱导的新生内膜形成。DOCK 2依赖性血管重塑/新生内膜形成的最确凿证据是DOCK 2敲除（DOCK 2-/-）显著阻断小鼠颈动脉中动脉结扎诱导的新生内膜增生。有趣的是，DOCK 2定位于SMC的细胞膜和细胞核中，这表明除了其在细胞骨架/细胞迁移中的作用之外，DOCK 2可能参与SMC基因转录。事实上，DOCK 2过表达阻断了SMC基因mRNA表达和myocardin诱导的平滑肌肌动蛋白启动子的激活。因此，中心假设是DOCK 2通过抑制SMC基因转录和刺激SMC迁移/增殖来调节SMC表型调节，导致新生内膜形成/血管重塑。本研究采用SMC原代培养、大鼠球囊损伤模型和小鼠钢丝损伤模型，结合分子、细胞和组织学方法，研究DOCK 2通过调控SMC基因转录调节SMC表型的分子机制，探讨DOCK 2是否通过激活Rac/RhoA/Cdc 42诱导SMC迁移，以及DOCK 2是否通过激活Rac/RhoA/Cdc 42诱导SMC迁移。3）确定DOCK 2在SMC表型调节和血管重塑中的重要作用。该项目的完成将揭示一种调节SMC表型调节的新机制，并为DOCK 2是否是对抗与糖尿病、再狭窄、动脉粥样硬化和癌症等常见疾病相关的血管损伤的潜在治疗靶点提供新的见解。