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）：该提案的总体目标是建立一种新机制，胞质分裂贡献者2（DOCK2）通过该机制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内膜形成/血管重塑，在动脉粥样硬化、血管成形术或搭桥术后再狭窄、糖尿病血管并发症、移植动脉病、哮喘和癌症的发展中发挥着关键作用。然而，人们对调节 SMC 表型调节和新内膜形成的机制和因素知之甚少。生理条件下，DOCK2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（一种 SMC 表型调节剂）可诱导 SMC 中的 DOCK2 表达。 DOCK2 的敲低可阻断 PDGF-BB 诱导的 SMC 表型调节、增殖和迁移。体内动物研究表明，DOCK2 在正常大鼠颈动脉的 SMC 中检测不到，但最初在中层 SMC 中诱导，随后在球囊导管诱导的血管损伤后在新内膜 SMC 中诱导。重要的是，DOCK2 的敲除显着抑制了损伤诱导的新内膜形成。 DOCK2依赖性血管重塑/新内膜形成最确凿的证据是，敲除DOCK2（DOCK2-/-）可显着阻断小鼠颈动脉中动脉结扎诱导的新内膜增生。有趣的是，DOCK2 定位于 SMC 的细胞膜和细胞核，这表明 DOCK2 除了在细胞骨架/细胞迁移中发挥作用外，还可能参与 SMC 基因转录。事实上，DOCK2 过表达阻断了 SMC 基因 mRNA 表达和心肌素诱导的平滑肌 β-肌动蛋白启动子激活。因此，核心假设是DOCK2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，导致新内膜形成/血管重塑。利用SMC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）研究DOCK2通过调节SMC基因转录来调节SMC表型的分子机制； 2)研究DOCK2是否通过激活Rac/RhoA/Cdc42诱导SMC迁移； 3)确定DOCK2在体内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调节 SMC 表型调节的新机制，并为 DOCK2 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。