Lipoproteins and Inflammation in the Development of Diabetic Complications

糖尿病并发症发生过程中的脂蛋白和炎症

• 批准号: 9275407
• 负责人: MARIA F LOPES-VIRELLA
• 金额: --
• 依托单位: RALPH H JOHNSON VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-10-01 至 2017-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9275407
• 关键词: Acute; Address; Age; Animal Model; Animals; Antibodies; Antigen-Antibody Complex; Apolipoprotein E; Apoptosis; Apoptotic; Arterial Fatty Streak; Atherosclerosis; Binding; Biological Assay; Biological Markers; Blood Circulation; Blood Vessels; Cardiovascular Diseases; Cardiovascular system; Cell Culture Techniques; Cell Separation; Cell Survival; Cells; Chronic; Clinical; Collagen; Collagen Gene; Complications of Diabetes Mellitus; Coronary Arteriosclerosis; Data; Development; Diabetes Mellitus; Disease Progression; Endothelial Cells; Event; Fc Receptor; Gelatinase B; Gene Expression; Gene Expression Profiling; Genes; Goals; Growth; Growth Factor; Health; Histologic; Human; Immunoglobulin G; Immunoglobulin M; In Vitro; Inflammation; Inflammation Mediators; Inflammatory; Insulin-Dependent Diabetes Mellitus; Interstitial Collagenase; Knockout Mice; Lead; Lipoproteins; Low Density Lipoprotein Receptor; Low-Density Lipoproteins; Malondialdehyde; Matrix Metalloproteinases; Measures; Mediating; Metalloproteases; Metalloproteinase Gene; Methods; Modification; Mus; Non-Insulin-Dependent Diabetes Mellitus; Pathogenesis; Pathway interactions; Patients; Pattern; Play; Predictive Value; Prevention strategy; Process; Production; Protein Array; Risk; Risk Factors; Role; Sampling; Serum; Testing; Therapeutic; Up-Regulation; Western Blotting; anti-IgM; base; biomarker panel; cardiovascular disorder risk; cohort; collagenase; cytokine; diabetic; enzyme activity; high risk; inhibitor/antagonist; low density lipoprotein inhibitor; macrophage; mesangial cell; mouse model; new therapeutic target; novel; novel marker; overexpression; oxidized low density lipoprotein; prevent; public health relevance; uptake

DESCRIPTION (provided by applicant): In the past few years we have clearly demonstrated in two large diabetic cohorts of type 1 and type 2 diabetes (DCCT/EDIC and VADT cohorts) that high levels of immune complexes (IC) containing oxidized LDL (oxLDL) and advanced glycated end-products-modified LDL (AGE-LDL) are associated with coronary artery disease (CAD) 1-4 and can strongly predict the development and progression of CAD in the early stages of type 1 diabetes3. We have also shown in the VADT cohort, which includes many patients with advanced CAD, that high levels of malondialdehyde-modified (MDA)-LDL in IC are able to predict acute cardiovascular disease (CVD) events4. However, when circulating IC contain both high levels of MDA-LDL and high levels of oxLDL or AGE-LDL the risk to suffer an acute event is reduced4 leading us to conclude that the modified lipoproteins carried by IC play a differential modulating role in plaque progression/stability and plaque destabilization. Our mechanistic studies support the above hypothesis. OxLDL IC induces the transformation of macrophages into foam cells5-7, lead to upregulation of pro-survival genes8 and induces the release of pro-inflammatory cytokines10. OxLDL IC also induces increased collagen production by mesangial cells12. In contrast, MDA-LDL IC induces macrophage apoptosis, stimulate the expression of matrix metalloproteinases (MMPs) by macrophages and by aortic endothelial cells, and reduce the expression of collagen genes in mesangial cells. Hypothesis and Specific Aims: Based on our clinical and in vitro preliminary data, we hypothesize that the type of predominant modifications of LDL in circulating IC (MDA-LDL, oxLDL or AGE-LDL) has a significant impact in plaque progression/destabilization. We postulate that, when MDA is the predominant modification of the LDL carried by IC, the uptake of those IC by macrophages induces cell apoptosis, increased release of MMPs, and increased collagenase activity, thus contributing to plaque destabilization. In contrast, when oxLDL is the predominant modification of LDL in IC, the uptake of these IC induce macrophage survival and stimulate the release of pro-inflammatory mediators and growth factors, thus contributing to plaque expansion/stability. Furthermore we postulate that a biomarker panel including MDA-LDL IC and MMPs will be more able to strongly predict patients at high risk to suffer acute CVD events than a panel of conventional CVD biomarkers. To test the above hypotheses we propose three aims. The first aim will compare the activation patterns of macrophages exposed to IC (MDA-LD IC and oxLDL IC), the effect of these IC on the survival and apoptosis of macrophages, and the pathways involved. In the second aim we will measure novel biomarkers known to be associated with plaque instability in patient samples from the VADT cohort in which MDA-LDL IC and conventional CVD biomarkers have been previously measured. We will determine whether a panel including MDA-LDL IC, MMP-1 and MMP-9 will have a significantly higher predictive power to identify patients at high risk for acute MI. Finally in the third aim we will determine, on an animal mouse model of atherosclerosis and type 2 diabetes whether blocking/reducing engagement of Fc�Rs by oxLDL IC and MDA-LDL IC by treatment with mouse IgM anti-oxLDL and IgM anti-MDA-LDL will prevent plaque progression and instability. Methods: Cell isolation and culture, separation and modification of lipoproteins, isolation of antibodies, protein arrays, cell viability/apoptosis assays, rtPCR, western blots, enzyme-activity assays, immunohistochemical analysis of gene expression and histologic analysis of atherosclerotic lesions. Relevance to VA Health and Significance: The definition of mechanisms responsible for the differences in cell activation, survival and apoptosis induced by different modified LDL-IC may allow to define new therapeutic targets and new prevention strategies that will curtail the progression of cardiovascular complications and the occurrence of acute vascular events in patients with type 2 diabetes.

描述（由申请人提供）： 在过去的几年中1型糖尿病的早期阶段的CAD进展3。我们还显示了包括许多晚期CAD患者的VADT队列中，IC中高水平的丙二醛改性（MDA）-LDL能够预测急性心血管疾病（CVD）事件4。但是，当循环IC包含高水平的MDA-LDL和高水平的OXLDL或年龄LDL急性事件的风险降低了4导致我们包括IC携带的改性脂蛋白在斑块进展/稳定性/稳定性和斑块上的差异中起差异调制作用。我们的机械研究支持上述假设。 OXLDL IC诱导巨噬细胞转化为泡沫细胞5-7，导致促寿使基因8的上调，并诱导促炎性细胞因子的释放。 OxLDL IC还诱导肾小球细胞产生胶原蛋白的增加12。相反，MDA-LDL IC诱导巨噬细胞凋亡，巨噬细胞和主动脉内皮细胞刺激基质金属蛋白酶（MMP）的表达，并减少膜根细胞中胶原蛋白基因的表达。假设和具体目的：基于我们的临床和体外初步数据，我们假设LDL在循环IC（MDA-LDL，OXLDL或Age-LDL）中的主要修饰类型对斑块进展/破坏稳定性产生了重大影响。我们假设，当MDA是IC携带的LDL的主要修饰时，巨噬细胞对这些IC的摄取会诱导细胞凋亡，增加MMP的释放和增加的胶原酶活性，从而导致斑块不稳定。相反，当OXLDL是IC中LDL的主要修饰时，这些IC的摄取会诱导巨噬细胞存活，并刺激促炎性介体和生长因子的释放，从而有助于斑块的扩张/稳定性。假设包括MDA-LIDL IC和MMP在内的生物标志物面板比常规CVD生物标志物更强烈预测遭受急性CVD事件的高风险患者。为了检验上述假设，我们提出了三个目标。第一个目标将比较暴露于IC（MDA-LD IC和OXLDL IC）的巨噬细胞的激活模式，这些IC对巨噬细胞存活和凋亡的影响以及所涉及的途径。在第二个目的中，我们将测量已知与VADT队列中MDA-LIC IC和常规CVD生物标志物的患者样本中已知与斑块不稳定性相关的新型生物标志物。我们将确定包括MDA-LIC，MMP-1和MMP-9在内的面板是否具有明显更高的预测能力来识别急性高风险的患者 mi。最后，在第三个目标中，我们将在动脉粥样硬化的动物小鼠模型和2型糖尿病上确定是否通过用小鼠IgM Anti-OXLDL和IgM抗MDA来阻止/减少FC.RS的互动/减少FC.RS的参与度都将防止斑块的进展和不稳定。方法：细胞分离和培养，脂蛋白的分离和修饰，抗体的分离，蛋白质阵列，细胞活力/凋亡测定法，RTPCR，蛋白质印迹，酶活性测定，免疫组织化学分析基因表达和基因学表达和动脉粥样硬化的组织学分析。与VA健康和意义相关：导致不同修饰的LDL-IC引起的细胞激活，生存和凋亡差异的机制的定义可能允许定义新的治疗靶标和新的预防策略，这些策略将减少心血管复杂性的进展以及急性血管性血管事件的发生，并在2型二次透射症患者中发生急性血管事件。