Hepatoma derived growth factor and vascular smooth muscle cell proliferation

肝癌源性生长因子与血管平滑肌细胞增殖

• 批准号: 7482313
• 负责人: ALLEN D EVERETT
• 金额: $ 36.9万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7482313
• 关键词: Acetylation; Address; Alanine; Amino Acids; Antibodies; Arterial Fatty Streak; Arteries; Binding; Binding Proteins; Biological Assay; Blood Vessels; Brain; Cardiovascular Diseases; Cause of Death; Cell Cycle; Cell Nucleus; Cells; Cholesterol; Collaborations; Condition; D-Aspartic Acid; DNA Binding; DNA-Binding Proteins; Data; Development; EMSA; Future; Gene Targeting; Genetic Transcription; Goals; Growth; Growth Factor; Heart; Histone Deacetylase; Histone deacetylase inhibition; Histones; Human; Immune Sera; In Vitro; Injury; Intervention; Knockout Mice; Laboratories; Link; Mediating; Modeling; Mus; Mutation; Nuclear; Numbers; PCNA gene; PKC Phosphorylation Site; PWWP Domain; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Pilot Projects; Positioning Attribute; Post-Translational Regulation; Process; Production; Proliferating; Protein Kinase C; Proteins; Public Health; Reagent; Recombinant Antibody; Relative (related person); Research Personnel; Role; Serine; Serum; Signal Transduction; Smooth Muscle Myocytes; Stents; Structure; Testing; Thinking; Transcription Repressor/Corepressor; United States; Vascular Diseases; base; cell growth; design; gene repression; hepatoma-derived growth factor; in vivo; injured; mutant; novel therapeutics; programs; promoter; protein function; research study; response; response to injury; vascular smooth muscle cell proliferation

DESCRIPTION (provided by applicant): Smooth muscle cell (VSMC) proliferation is a key component of the pathophysiologic response to vascular injury. Central to this process is the production and activation of growth factors. One such VSMC growth factor, hepatoma derived growth factor (HDGF), becomes highly expressed in the nucleus of neointimal cells after balloon injury and is co-expressed with PCNA in proliferating VSMC in human atherosclerotic plaques. However, the mechanisms by which HDGF stimulates VSMC proliferation are unknown. The overall goal of this proposal is to elucidate the nuclear mechanisms by which HDGF stimulates VSMC proliferation in vitro and in response to injury. In our pilot studies, we found that HDGF is phosphorylated by protein kinase C, a key signaling intermediate in vascular injury. We also now have evidence that HDGF is a nuclear binding protein, functioning as a transcriptional repressor. Based upon these studies, we propose to determine the mechanisms of PKC mediated post-translational regulation of HDGF mitogenic function in vitro and in vivo and as a DMA binding, transcriptional repressor. First, we will test the hypothesis that PKC phosphorylation s critical to HDGF mitogenic function in isolated VSMC and in the wire injured mouse carotid using adenoviral expression of single or combination mutations of candidate PKC serines (S) to alanine (A, loss of phosphorylation) or aspartic acid (D, phosphorylation mimic) mutations, PKCa deficient knockout mice, and phospho-specific HDGF antibodies. Second, we will test the hypotheses that HDGF is a DNA binding protein using NMR based structure/function analyses, functions as a transcriptional repressor binding to the promoters of HDGF target genes via the controversial PWWP domain, whether transcriptional repression is mediated by the HDAC pathway and whether these activities are regulated by PKC phosphorylation. The significance of the proposed studies is that by linking HDGF activation with PKC, a critical step in the function of HDGF in vascular injury is revealed and provides the basis for new therapeutic strategies in vascular disease. Relevance to public health: Cardiovascular disease is the number one cause of death in the United States despite interventions with cholesterol lowering agents and the development of endovascular stents. This is because of the continued detrimental growth of vascular cells in the damaged arteries of patients; eventually, leading to vascular insufficiency of the brain or heart. We have identified a protein (HDGF) produced by vascular cells in diseased arteries that increases the growth of vascular cells. We propose here to determine the mechanism regulating the activity of this protein to provide the basis for the future design of new drugs to treat cardiovascular disease.

描述（由申请人提供）：平滑肌细胞（VSMC）增殖是血管损伤病理生理反应的关键组成部分。这个过程的核心是生长因子的产生和激活。其中一种VSMC生长因子，肝癌源性生长因子（HDGF），在球囊损伤后在新内膜细胞的细胞核中高度表达，并在人动脉粥样硬化斑块中增殖的VSMC中与PCNA共表达。然而，HDGF刺激VSMC增殖的机制尚不清楚。本提案的总体目标是阐明HDGF在体外刺激VSMC增殖和对损伤反应的核机制。在我们的初步研究中，我们发现HDGF被蛋白激酶C磷酸化，蛋白激酶C是血管损伤的关键信号传导介质。我们现在也有证据表明HDGF是一种核结合蛋白，作为转录抑制因子发挥作用。基于这些研究，我们建议在体外和体内确定PKC介导的HDGF有丝分裂功能的翻译后调控机制，并作为DMA结合的转录抑制因子。首先，我们将通过腺病毒表达候选PKC丝氨酸(s)到丙氨酸（A，磷酸化缺失）或天冬氨酸（D，磷酸化模拟）突变的单一或组合突变、PKCa缺陷敲除小鼠和磷酸化特异性HDGF抗体，来验证PKC磷酸化对离体VSMC和钢丝损伤小鼠颈动脉中HDGF有丝分裂功能至关重要的假设。其次，我们将使用基于NMR的结构/功能分析来验证HDGF是DNA结合蛋白的假设，通过有争议的PWWP结构域作为转录抑制因子结合HDGF靶基因的启动子，转录抑制是否由HDAC途径介导，以及这些活性是否受PKC磷酸化调节。本研究的意义在于，通过将HDGF激活与PKC联系起来，揭示了HDGF在血管损伤中功能的关键步骤，并为血管疾病的新治疗策略提供了基础。与公众健康相关：尽管使用降胆固醇药物和血管内支架进行干预，心血管疾病仍是美国的头号死亡原因。这是因为患者受损动脉中的血管细胞持续有害生长；最终导致大脑或心脏血管功能不全。我们已经鉴定出病变动脉中血管细胞产生的一种蛋白质（HDGF），它可以促进血管细胞的生长。我们在此建议确定该蛋白活性的调控机制，为今后设计治疗心血管疾病的新药提供依据。