MARKERS AMP; MECHANISMS FOR PRE-ECLAMPSIA IN WOMEN W/TYPE 1 DIABETES

标记放大器；

• 批准号: 7378101
• 负责人: TIMOTHY J LYONS
• 金额: $ 0.32万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2007-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7378101
• 关键词: MARKERS; AMP; MECHANISMS; PRE; ECLAMPSIA

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Introduction: Our primary goal is to define the roles of dyslipidemia and oxidative stress in accelerating macrovascular disease (atherosclerosis) in diabetes. Microvascular disease (nephropathy and retinopathy) will be studied as secondary end-points. The place of our project in the broad scope of the program is shown in Figure 1. We will continue (and expand) our longitudinal and cross-sectional studies of the DCCT/EDIC cohort of Type 1 diabetic patients. This cohort is the largest and best-characterized group of Type 1 diabetic patients in the world. Though still relatively young, it has provided exciting data during the first funding period of the Program Project (1996-present). To obtain a higher "hard" event rate (myocardial infarction, stroke etc.) and to study the more common form of diabetes, we will add a Type 2 diabetes cohort from the VA Cooperative Trial "Glycemia and Complications in Diabetes, Type 2". This prospective trial commences October 2000, and as in the DCCT, patients will be randomized to intensive and standard glycemic control. The VA trial provides us with the opportunity to study patients during the randomized portion of the study (in the DCCT, randomization finished in 1993). We have obtained interim funding to support our collaboration with the VA trial in its first year. We will also perform basic science studies to investigate hypotheses we have developed over the past four years relating to dyslipidemia, oxidative stress and atherogenesis in diabetes. These studies, in collaboration with the other Projects and Cores and the Coordinating Centers of both studies, will greatly increase our understanding of the complex multi-factorial mechanisms underlying vascular disease in both types of diabetes. Our Specific Aims are: 1. To define the role of dyslipidemia in the vascular complications of Type 1 and Type 2 diabetes. We will perform comprehensive lipoprotein analyses, including Nuclear Magnetic Resonance (NMR) lipoprotein profiles, levels of apolipoproteins, Lipoprotein(a), and studies of lipoprotein-associated enzymes. We hypothesize that dyslipidemia, including both quantitative and qualitative lipoprotein abnormalities which can now be assessed in much greater detail than ever before, mediates endothelial cell injury and activation, contributing to accelerated atherosclerosis in diabetes. We will obtain NMR lipoprotein profiles of selected subjects from the DCCT/EDIC Type 1 diabetes cohort ("Type 1 Cohort") on two occasions, completing a total of seven determinations over the entire DCCT/EDIC study duration (1983-2006). We will also obtain NMR analyses prospectively on two occasions in the VA Cooperative Trial Type 2 diabetes cohort ("Type 2 Cohort") during 2000-2006. We will measure apolipoproteins A1, B, E, and Lp(a). We will determine activities of the following lipoprotein-related enzymes: paraoxonase (PON), Lecithin Cholesterol Acyl Transferase (LCAT), and Platelet Activating Factor Acetyl Hydrolase (PAFAH). The data will be related to lipoprotein subclass distribution, and cross-sectionally and prospectively to vascular complication status, randomization groups, and interventions. 2. To define the roles of oxidative stress, inflammation, and homocysteine in the vascular complications of Type 1 and Type 2 diabetes. We hypothesize that oxidative stress, inflammatory processes, and elevated homocysteine levels mediate endothelial injury and accelerated atherosclerosis in diabetes. In both cohorts, we will determine products of free radical oxidation and antioxidant reserves in plasma to assess oxidative stress, C-reactive protein and serum amyloid A as measures of inflammatory processes, and plasma homocysteine levels. 3. To determine the effect of lipoprotein associated enzymes on lipoprotein subclass distribution. We hypothesize that lipoprotein subclass distribution in serum is affected by LCAT, CETP, and PAFAH activity. Complementing studies of NMR determined lipoprotein subclass distribution and enzyme activities in vivo (Specific Aim 1), we will incubate serum with purified enzymes in vitro and determine effects on lipoprotein subclass distribution by NMR. 4. To determine functional characteristics of LDL, HDL, and their subclasses from Type 1 and Type 2 diabetic patients and control subjects in studies using human aortic endothelial cells (HAEC) and human red blood cell (RBC) membranes. We hypothesize that altered interactions of lipoproteins with vascular cells, related to dyslipidemia and oxidative stress, contribute to endothelial injury and accelerated atherosclerosis in diabetes. Using well- characterized lipoproteins from Type 1 and Type 2 diabetic subjects and from non-diabetic control subjects, we will determine: (i) effects of large, medium, and small LDL on HAEC expression of proteins mediating fibrinolysis, vascular tone, and adhesion molecules, and on matrix binding and intracellular calcium flux. (ii) ability of total HDL, and of large (cardioprotective) HDL2 and small (non-protective) HDL3 to: (a) mitigate oxidized-LDL induced HAEC adhesion molecule expression, tPA suppression, PAI-1 and NO production. (b) mitigate TNF- induced HAEC adhesion molecule expression (c) protect LDL and RBC membranes from in vitro oxidation (d) remove pre-formed lipid peroxides from oxidized RBC membranes B. BACKGROUND CONTEXT WITHIN THE PROGRAM: The place of the project in the program is shown in Figure 1 (previous page). Interactions, both logistic and scientific, of Project 1 with other Projects and Cores within the Program are shown in Figure 2 (next page) and are summarized below. With Project 2: Potential markers identified in Project 1 will be correlated with lipoprotein immune complexes and circulating adhesion molecules determined in Project 2. We hypothesize that patients with adverse lipoprotein profiles or functions, higher levels of inflammation or oxidative stress will have higher levels of injurious lipoprotein immune complexes and adhesion molecules, and a higher prevalence/risk of macrovascular disease (MVD). Effects of LDL and HDL subfractions on adhesion molecule expression by cultured HAEC (Project 1) will be related to circulating adhesion molecules in the study cohorts (Project 2). With Project 3: Potential markers identified in Project 1 will be correlated with levels of kallikreins, kinins and growth factors determined in Project 3. We hypothesize that patients with adverse lipoprotein profiles or functions, or higher oxidative stress will have greater activity of the kallikrein/kinin system, and higher levels of growth factors that promote vascular injury. We hypothesize that large, intermediate, and small LDL, induce different patterns of intracellular Ca++ flux in HAEC, and will evaluate this in collaboration with Project 3. With Project 4: Potential markers identified in Project 1 will be correlated with insulin resistance, CETP and PLT activity and abnormalities of the fibrinolytic system determined in Project 4. We hypothesize that patients with adverse lipoprotein profiles or functions, or higher oxidative stress, will be more insulin resistant, have reduced fibrinolytic activity, and that CETP and PLT activities will relate to lipoprotein subclass distribution in vivo. Effects of LDL subfractions on expression of modulators of fibrinolysis by cultured HAEC (Project 1) will be related to data concerning tPA and PAI-1 in the study cohorts (Project 4). Project 4 will measure CETP activity in plasma incubated with lipoprotein associated enzymes. With Core A (Administration): Project 1 will receive all samples through Core A, which will coordinate interactions with participating DCCT/EDIC, and VA centers, and facilitate sample transfer and distribution. With Core B (Biostatistics): All data generated by Project 1 will be provided to Core B for statistical analyses. Project 1 will consult Core B on experimental design and data management, including the selection of MVD progressors vs non-progressors for nested case control studies. With Core C (Genetics Core): Project 1 will determine lipoprotein subclass distributions and size, apolipoprotein levels, lipoprotein-related enzyme activities, and measures of oxidative stress and Hcy. This information will be related to genotype data generated in Project 5, including that relating to Lp(a), apoE, CETP, PAFAH, Hepatic Lipase, HDL and small dense LDL, and Hcy.

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。我们的主要目标是确定血脂异常和氧化应激在加速糖尿病大血管疾病（动脉粥样硬化）中的作用。微血管疾病（肾病和视网膜病变）将作为次要终点进行研究。我们的项目在程序的大范围中的位置如图1所示。我们将继续（并扩大）对1型糖尿病患者DCCT/EDIC队列的纵向和横断面研究。该队列是世界上最大和最具特征的1型糖尿病患者组。虽然还比较年轻，但在方案项目的第一个资助期（1996年至今）提供了令人兴奋的数据。为了获得更高的“硬”事件发生率（心肌梗死、中风等）并研究更常见的糖尿病形式，我们将从VA合作试验“2型糖尿病的血糖和并发症”中加入一个2型糖尿病队列。这项前瞻性试验于2000年10月开始，与DCCT一样，患者将随机分为强化和标准血糖控制两组。VA试验为我们提供了在研究的随机部分研究患者的机会（在DCCT中，随机化于1993年完成）。我们已经获得了临时资金，以支持我们在第一年与VA试验的合作。我们还将开展基础科学研究，以调查我们在过去四年中提出的关于糖尿病中血脂异常、氧化应激和动脉粥样硬化的假设。这些研究与其他项目和核心以及这两项研究的协调中心合作，将大大增加我们对两种类型糖尿病中血管疾病的复杂多因素机制的理解。我们的具体目标是：1；目的探讨血脂异常在1型和2型糖尿病血管并发症中的作用。我们将进行全面的脂蛋白分析，包括核磁共振（NMR）脂蛋白谱、载脂蛋白水平、脂蛋白(a)和脂蛋白相关酶的研究。我们假设血脂异常，包括定量和定性脂蛋白异常，现在可以比以往更详细地评估，介导内皮细胞损伤和激活，加速糖尿病动脉粥样硬化。我们将从DCCT/EDIC 1型糖尿病队列（“1型队列”）中两次获得选定受试者的NMR脂蛋白谱，在整个DCCT/EDIC研究期间（1983-2006）共完成7次测定。我们还将在2000-2006年期间在VA合作试验2型糖尿病队列（“2型队列”）中两次获得前瞻性核磁共振分析。我们将测量载脂蛋白A1、B、E和Lp(a)。我们将测定以下脂蛋白相关酶的活性：对氧磷酶（PON）、卵磷脂胆固醇酰基转移酶（LCAT）和血小板活化因子乙酰水解酶（PAFAH）。这些数据将与脂蛋白亚类分布、血管并发症状态、随机分组和干预措施的横断面和前瞻性有关。2. 目的探讨氧化应激、炎症和同型半胱氨酸在1型和2型糖尿病血管并发症中的作用。我们假设氧化应激、炎症过程和升高的同型半胱氨酸水平介导了糖尿病的内皮损伤和加速动脉粥样硬化。在这两个队列中，我们将测定血浆中自由基氧化产物和抗氧化剂储备，以评估氧化应激、c反应蛋白和血清淀粉样蛋白A作为炎症过程的测量指标，以及血浆同型半胱氨酸水平。3. 目的：探讨脂蛋白相关酶对脂蛋白亚类分布的影响。我们假设血清中脂蛋白亚类分布受LCAT、CETP和PAFAH活性的影响。作为核磁共振测定体内脂蛋白亚类分布和酶活性研究的补充（Specific Aim 1），我们将在体外用纯化酶培养血清，并通过核磁共振测定对脂蛋白亚类分布的影响。4. 在使用人主动脉内皮细胞（HAEC）和人红细胞（RBC）膜的研究中，确定1型和2型糖尿病患者和对照受试者的LDL、HDL及其亚类的功能特征。我们假设脂蛋白与血管细胞相互作用的改变，与血脂异常和氧化应激有关，有助于糖尿病内皮损伤和加速动脉粥样硬化。使用从1型和2型糖尿病患者和非糖尿病对照组中提取的特征明确的脂蛋白，我们将确定：(i)大、中、小LDL对介导纤维蛋白溶解、血管张力和粘附分子的HAEC蛋白表达的影响，以及对基质结合和细胞内钙通量的影响。（ii）总HDL、大（心脏保护）HDL2和小（非保护）HDL3的能力：(a)减轻氧化ldl诱导的HAEC粘附分子表达、tPA抑制、PAI-1和NO产生。(b)减轻TNF-诱导的HAEC粘附分子表达(c)保护LDL和RBC膜免受体外氧化(d)从氧化的RBC膜中去除预形成的脂质过氧化物b .项目背景背景：项目在项目中的位置如图1所示（上一页）。图2（下一页）显示了项目1与计划内其他项目和核心的逻辑和科学交互，并总结如下。项目2：项目1中确定的潜在标记物将与项目2中确定的脂蛋白免疫复合物和循环粘附分子相关。我们假设，脂蛋白谱或功能不良、炎症或氧化应激水平较高的患者将具有更高水平的有害脂蛋白免疫复合物和粘附分子，以及更高的大血管疾病（MVD）患病率/风险。LDL和HDL亚组分对培养HAEC粘附分子表达的影响（项目1）将与研究队列中循环粘附分子有关（项目2）。在项目3中：项目1中确定的潜在标记物将与项目3中确定的钾激肽、激肽和生长因子的水平相关。我们假设有不良脂蛋白谱或功能，或较高氧化应激的患者会有更大的钾激肽/激肽系统活性，以及更高水平的促进血管损伤的生长因子。我们假设大、中、小LDL诱导HAEC细胞内Ca++通量的不同模式，并将与Project 3合作对此进行评估。项目4：项目1中发现的潜在标志物将与胰岛素抵抗、CETP和PLT活性以及项目4中确定的纤溶系统异常相关。我们假设，具有不良脂蛋白谱或功能或较高氧化应激的患者将更具有胰岛素抵抗性，纤维蛋白溶解活性降低，并且CETP和PLT活性将与体内脂蛋白亚类分布有关。LDL亚组分对培养HAEC纤维蛋白溶解调节剂表达的影响（项目1）将与研究队列中tPA和PAI-1的数据相关（项目4）。项目4将测量血浆中与脂蛋白相关酶孵育的CETP活性。与核心A（管理）：项目1将通过核心A接收所有样品，核心A将协调与参与DCCT/EDIC和VA中心的互动，并促进样品的转移和分发。Core B（生物统计学）：项目1产生的所有数据将提供给Core B进行统计分析。项目1将在实验设计和数据管理方面咨询核心B，包括在嵌套病例对照研究中选择MVD进展者和非进展者。与核心C（遗传学核心）：项目1将确定脂蛋白亚类分布和大小，载脂蛋白水平，脂蛋白相关酶活性，以及氧化应激和Hcy的测量。该信息将与项目5中生成的基因型数据相关，包括与Lp(a)、apoE、CETP、PAFAH、肝脂肪酶、HDL和小密度LDL以及Hcy相关的基因型数据。