Lipoxygenase and vascular disease in diabetes

本研究旨在探讨脂氧合酶（LO）激活和氧化应激在高血糖（HG）和胰岛素抵抗引起的动脉粥样硬化和血管损伤中的作用。我们的研究表明，在糖尿病条件下血管平滑肌细胞（VSMC）功能障碍的NO途径。结果表明，HG、生长因子和细胞因子均能诱导12-LO的活性和表达。LO产物导致VSMC的生长和趋化作用。在这次更新中，我们将重点放在i）HG和LO产品介导其在VSMC中的作用的细胞和分子机制，以及ii）在糖尿病和动脉粥样硬化和血管损伤的动物模型中，通过诱导介导VSMC生长，迁移和基质重塑的关键基因的转录调节，12-LO激活的体内相关性。主要假设是LO通路通过诱导调控VSMC生长、迁移和基质重塑的关键基因的转录而在动脉粥样硬化和血管损伤的发病机制中发挥作用。具体目的：1）研究HG和LO产物介导的猪血管平滑肌细胞基因表达的信号转导机制。在这里，我们将研究HG和LO产品对ERK 1/2和p38丝裂原活化蛋白激酶的激活的影响。我们还将研究IkappaB激酶在HG和LO产物诱导的NF-κ B中的作用。12-LO的中介作用将在新型12-LO核酶的帮助下进行测试。2)研究HG和LO产物对致动脉粥样硬化关键基因的转录调控机制，并确定LO激活在HG和血管紧张素II（Ang II）作用中的功能作用。我们将研究炎症基因VCAM-1、基质蛋白纤连蛋白和基质调节MT 1-MMP的调节。将在过表达12-LO cDNA或12-LO核酶的VSMC中检查体外12-LO激活的功能相关性。3)阐明12- LO和MAPK激活在糖尿病血管并发症病理生理学中的体内相关性。我们将评估小鼠模型中的组织学、12-LO表达以及12个LO调节的信号和基因，所述小鼠模型包括与apoE和/或糖尿病背景杂交的白细胞型12-LO敲除小鼠。我们将研究12-LO是否介导胰岛素抵抗和糖尿病大鼠模型的加速新生内膜增厚。该项目的一个独特的方面将是评估血管的变化和12-LO的作用，在猪模型的加速动脉粥样硬化由于糖尿病。在该模型中，我们将测试以下因素的影响：1）胰岛素严格控制血糖，2）使用新型抗炎剂异丙茶碱对脂质过氧化物的药理学抑制，3）冠状动脉支架植入术的影响。该项目将利用两个核心，并与项目2-4密切互动。这些结果应该增加我们对导致糖尿病心血管疾病加速的因素的认识，并导致新的治疗进展。

The role of lipoxygenase (LO) activation and oxidative stress in atherosclerosis and vascular injury due to high glucose (HG) and insulin resistance will be tested in this project. Our studies have implicated the NO pathway in vascular smooth muscle cell (VSMC) dysfunction under diabetic conditions. We showed that HG, growth factors and cytokines could induce 12-LO activity and expression. LO products lead to growth and chemotactic effects in VSMC. In this renewal, we will focus on i) the cellular and molecular mechanisms by which HG and LO products mediate their effects in VSMC and ii) the in vivo relevance of 12-LO activation in animal models of diabetes and atherosclerosis and vascular injury by inducing the transcriptional regulation of key genes that mediate VSMC growth, migration and matrix remodeling. The main hypotheses is that the LO pathway can play a role in the pathogenesis of atherosclerosis and vascular injury by inducing the transcriptional regulation of key genes that mediate VSMC growth, migration and matrix remodeling. The Specific Aims are 1) To examine the signal transduction mechanisms of HG and LO product-mediated gene expression in porcine VSMC. Here we will examine the effects of HG and LO products on the activation of ERK1/2 and p38 mitogen-activated protein kinases. We will also examine the role of IkappaB kinases in HG and LO product-induced of NF-kappaB. The intermediary role of 12-LO will be tested with the aid of novel 12-LO ribozymes. 2) To examine the mechanisms of transcriptional regulation of key atherogenic genes by HG and LO products, and to determine the functional role of LO activation in HG and angiotensin II (Ang II) actions. We will examine the regulation of the inflammatory gene, VCAM-1, the matrix protein fibronectin, and the matrix regulating MT1-MMP. The functional relevance of 12-LO activation in vitro will be examine in VSMC that overexpress 12-LO cDNA or the 12-LO ribozyme. 3) To clarify the in vivo relevance of 12- LO and MAPK activation in the pathophysiology of diabetic vascular complications. We will evaluate histology, 12-LO expression, as well as 12 LO-regulated signals and genes in mouse models including leukocyte- type 12-LO knock-out mouse cross bred to the apoE and/or the diabetic background. We will examine whether 12-LO mediates the accelerated neointimal thickening in a rat model of insulin resistance and diabetes. A unique aspect of this project will be to evaluate vascular changes and the role of 12-LO in a porcine model of accelerated atherosclerosis due to diabetes. In this model we will test the effect of: 1) tight glucose control with insulin, 2) pharmacologic inhibition of lipid peroxides using a novel anti-inflammatory agent lisofylline 3) effects of coronary artery stenting. This project will utilize both Cores and closely interact with Projects 2-4. The results should increase our knowledge of the factors leading to accelerated cardiovascular disease in diabetes and lead to new therapeutic advances.