Toll-Like Receptors and Atherosclerosis in Diabetes

Toll 样受体与糖尿病中的动脉粥样硬化

• 批准号: 8198364
• 负责人: Yan Huang
• 金额: --
• 依托单位: RALPH H JOHNSON VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-10-01 至 2014-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8198364
• 关键词: Address; Adult; Aftercare; American; Animal Model; Animals; Arterial Fatty Streak; Atherosclerosis; CD14 Antigen; Cardiovascular Diseases; Cardiovascular system; Cell Line; Cells; Child; Clinical; Coronary; Development; Diabetes Mellitus; Diagnosis; Dyslipidemias; Electrophoretic Mobility Shift Assay; Epidemic; Event; Exhibits; Gene Expression; General Population; Glucose; Health; Healthcare; Immune response; Immunity; In Vitro; Incidence; Infiltration; Inflammation; Inflammatory; Insulin Resistance; Interferon Type II; Interstitial Collagenase; Lesion; Ligands; Lipids; Lipopolysaccharides; Lymphocyte Antigen 96; Matrix Metalloproteinases; Mediating; Metabolic syndrome; Mitogen-Activated Protein Kinases; Molecular Biology; Mononuclear; Mus; Natural Immunity; Newly Diagnosed; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Pathway interactions; Patients; Persons; Plasma; Play; Prediabetes syndrome; Prevalence; Prevention; Proteins; Reporting; Research Project Grants; Risk; Role; Rupture; Signal Pathway; Signal Transduction; TLR2 gene; TLR4 gene; Testing; Thrombosis; Thrombus; Time; Tissues; Toll-like receptors; United States; Veterans; acute coronary syndrome; atherogenesis; base; cardiovascular disorder risk; cytokine; diabetic; diabetic patient; insight; macrophage; monocyte; mortality; non-diabetic; novel; novel therapeutic intervention; novel therapeutics; photosynthetic bacteria; prevent; promoter; public health relevance; receptor; receptor expression; toll-like receptor 4

DESCRIPTION (provided by applicant): Diabetes is an epidemic in USA and Veterans are nearly three times as likely as the general population to have diabetes. Patients with diabetes have an increased risk of developing cardiovascular disease and more than 75 percent of them die of cardiovascular complications. Therefore, it is crucial to develop new therapeutic strategies for cardiovascular disease to reduce mortality in diabetic patients. In recent years, the innate immunity (immunity a person is born with) has been shown to play an important role in atherosclerosis, a major cause of cardiovascular disease. Studies have shown that deficiency of toll-like receptor (TLR)4 that triggers cellular innate immune responses is associated with a significant reduction of atherosclerotic lesions in nondiabetic mice, suggesting that TLR4 is involved in atherosclerosis. However, these studies only demonstrated the effect of "lack of TLR4" on development of atherosclerosis. From clinical point of view, since it is known that stabilization of atherosclerotic plaques is critical to prevent cardiovascular events, and that all patients have TLR4 expression and many of them had developed atherosclerosis when they were diagnosed with type 2 diabetes, it is critical to determine if TLR4 inhibition with TLR4 antagonist in diabetic animal models that already have atherosclerosis changes plaque composition and gene expression that favor plaque stability. Nevertheless, the information is lacking. SPECIFIC OBJECTIVES AND HYPOTHESES: We found that elevated glucose concentrations (high glucose), a hallmark of diabetes, enhanced lipopolysaccharide (LPS)-stimulated matrix metalloproteinase (MMP) and proinflammatory cytokine expression by mononuclear cells. We also found that high glucose enhanced interferon gamma (IFN3)-stimulated MMP expression by upregulating MD-2, a TLR4-associated protein that confers TLR4 signaling. Since TLR4 is the receptor for LPS, and MMPs and proinflammatory cytokines play a crucial role in atherosclerotic plaque instability, the above findings suggest that TLR4 may be involved in diabetes-promoted plaque destabilization. Furthermore, we found that TLR4 antagonist Rs-LPS (isolated from nontoxic photosynthetic bacterium R. sphaeroides) blocked LPS-stimulated MMP-1 expression by mononuclear cells. Recent studies also showed the potency of Rs-LPS as a TLR4 antagonist. Based on these findings, we hypothesized that TLR4 inhibition with Rs-LPS in mice with atherosclerosis and diabetes or metabolic syndrome changes the lesion composition and gene expression that favor plaque stability. In this project, we will conduct animal studies to test our hypothesis. In addition, we will also conduct in vitro studies to further investigate the mechanisms involved in the stimulation of MD-2 expression by high glucose and IFN3. Two specific objectives were proposed: 1. To test our hypothesis that TLR4 inhibition in mice with established atherosclerosis and diabetes changes the lesion composition and gene expression that favor plaque stability. 2. To determine the signaling mechanisms involved in the stimulation of MD-2 expression by high glucose and IFN3, and the role of MD-2 in the IFN3 signaling. METHODS: We will first induce atherosclerotic lesions in mice with diabetes or metabolic syndrome and then treat mice with TLR4 antagonist Rs-LPS. After treatment, the plaque composition and gene expression will be analyzed. In the in vitro study, we will use macrophages and macrophage cell line and perform molecular biology studies such as electrophoretic mobility shift assay and promoter activity analysis. SIGNIFICANCE: This study will establish whether there is a rationale for targeting TLR4 in prevention of cardiovascular event in patients with diabetes or metabolic syndrome. POTENTIAL IMPACT ON VETERANS HEALTH CARE: The incidence of diabetes in Veterans is much higher than that in general population. This research project will provide important information for developing a new therapeutic strategy to prevent fatal cardiovascular events in our Veterans with diabetes. PUBLIC HEALTH RELEVANCE: The prevalence of diabetes in Veterans is nearly three times greater than that in the general population. Most of diabetic patients die of atherosclerosis-related cardiovascular disease. Therefore, it is important to develop new therapeutic interventions to prevent fatal cardiovascular events in diabetic patients. Recent studies have suggested that Toll-like receptor (TLR) 4 plays an important role in atherosclerosis. However, it remains undetermined if TLR4 inhibition in animal models of atherosclerosis and diabetes or metabolic syndrome changes atherosclerotic plaque composition and gene expression that favor plaque stability. In this study, we will investigate the effect of TLR4 antagonist on plaque composition and gene expression in animals with diabetes or metabolic syndrome. We will also conduct in vitro studies on the role of TLR4-associated protein MD-2 in the interferon gamma signaling pathway. Overall, this research project will establish a rationale for targeting TLR4 in treatment of atherosclerosis in diabetic patients. Thus, this study is highly relevant to Veterans' health.

描述（由申请人提供）： 糖尿病在美国是一种流行病，退伍军人的糖尿病的可能性几乎是糖尿病的三倍。糖尿病患者患心血管疾病的风险增加，其中75％以上死于心血管并发症。因此，为心血管疾病制定新的治疗策略至关重要，以降低糖尿病患者的死亡率。近年来，已经证明先天免疫力（一个人出生）在动脉粥样硬化中起着重要作用，这是心血管疾病的主要原因。研究表明，触发细胞先天免疫反应的通行类受体（TLR）4的缺乏与非糖尿病小鼠动脉粥样硬化病变的显着降低有关，这表明TLR4与动脉粥样硬化有关。但是，这些研究仅证明了“缺乏TLR4”对动脉粥样硬化发展的影响。从临床角度来看，由于众所周知，动脉粥样硬化斑块的稳定对于预防心血管事件至关重要，并且所有患者都具有TLR4表达，并且其中许多患者在被诊断为2型糖尿病时患有动脉粥样硬化，这对于在TLR4抑制TLR4拮抗剂中是否抑制TLR4拮抗剂的构成是至关重要的斑块稳定性。然而，这些信息仍缺乏。 特定目标和假设：我们发现葡萄糖浓度升高（高葡萄糖），糖尿病的标志，增强的脂多糖（LPS）刺激的基质金属蛋白酶（MMP）和一核细胞的促炎细胞因子表达。我们还发现，高葡萄糖增强了干扰素伽马（IFN3）刺激的MMP表达，通过上调MD-2，这是一种赋予TLR4信号传导的TLR4相关蛋白。由于TLR4是LPS的受体，因此MMP和促炎细胞因子在动脉粥样硬化斑块不稳定性中起着至关重要的作用，因此上述发现表明TLR4可能参与糖尿病促进的斑块不稳定。此外，我们发现TLR4拮抗剂RS-LP（从无毒的光合细菌R. sphaeroides分离出来）通过单核细胞阻断了LPS刺激的MMP-1表达。最近的研究还表明，RS-LP作为TLR4拮抗剂的效力。基于这些发现，我们假设在动脉粥样硬化和糖尿病或代谢综合征的小鼠中用RS-LPS抑制TLR4改变了有利于斑块稳定性的病变组成和基因表达。 在这个项目中，我们将进行动物研究以检验我们的假设。此外，我们还将进行体外研究，以进一步研究高葡萄糖和IFN3刺激MD-2表达的机制。提出了两个具体的目标：1。为了测试我们的假设，即具有既定的动脉粥样硬化和糖尿病的小鼠的TLR4抑制改变了有利于斑块稳定性的病变组成和基因表达。 2。确定高葡萄糖和IFN3刺激MD-2表达的信号传导机制，以及MD-2在IFN3信号传导中的作用。 方法：我们将首先在患有糖尿病或代谢综合征的小鼠中诱导动脉粥样硬化病变，然后用TLR4拮抗剂RS-LPs治疗小鼠。处理后，将分析斑块组成和基因表达。在体外研究中，我们将使用巨噬细胞和巨噬细胞系，并进行分子生物学研究，例如电泳迁移率转移测定法和启动子活性分析。 意义：这项研究将确定靶向TLR4在预防糖尿病或代谢综合征患者心血管事件方面是否存在理由。 对退伍军人卫生保健的潜在影响：退伍军人中糖尿病的发病率远高于普通人群。该研究项目将提供重要的信息，以制定一种新的治疗策略，以防止我们的糖尿病退伍军人中致命的心血管事件。 公共卫生相关性： 退伍军人中糖尿病的患病率几乎是普通人群的三倍。大多数糖尿病患者死于动脉粥样硬化相关的心血管疾病。因此，开发新的治疗干预措施以防止糖尿病患者的致命心血管事件非常重要。最近的研究表明，Toll样受体（TLR）4在动脉粥样硬化中起重要作用。然而，如果在动脉粥样硬化和糖尿病或代谢综合征的动物模型中TLR4抑制是否会改变动脉粥样硬化斑块组成和基因表达，这仍然​​不确定。在这项研究中，我们将研究TLR4拮抗剂对糖尿病或代谢综合征动物牙菌斑组成和基因表达的影响。我们还将在体外研究TLR4相关蛋白MD-2在干扰素伽马信号通路中的作用。总体而言，该研究项目将建立靶向TLR4治疗糖尿病患者动脉粥样硬化的基本原理。因此，这项研究与退伍军人的健康高度相关。