The role of hyaluronan in hIAPP-induced beta cell toxicity

透明质酸在 hIAPP 诱导的 β 细胞毒性中的作用

• 批准号: 10427226
• 负责人: REBECCA LUCY HULL-MEICHLE
• 金额: --
• 依托单位: VA PUGET SOUND HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10427226
• 关键词: 4-methylumbelliferone; Address; Adhesions; Affect; Amyloid; Amyloid deposition; Animal Model; Beta Cell; Blood capillaries; Bone Marrow; Capillary Endothelial Cell; Caring; Cell Adhesion Molecules; Cell Death; Cell Line; Cell physiology; Cells; Chemotaxis; Data; Deposition; Disease; Endothelial Cells; Endothelium; Extracellular Matrix; Functional disorder; General Population; High Fat Diet; Human; Hyaluronan; Hyaluronidase; ITGAM gene; In Vitro; Individual; Inflammasome; Inflammatory; Insulin Resistance; Interleukin-1 beta; Interleukin-6; Intervention; Islets of Langerhans; Link; Literature; Mediating; Mediator of activation protein; Modeling; Molecular Weight; Morphology; Mus; Non-Insulin-Dependent Diabetes Mellitus; Pathologic; Patients; Peptide Conformation; Peptides; Pharmaceutical Preparations; Process; Production; Protein Isoforms; Role; Signal Transduction; System; TNF gene; Therapeutic; Time; Toxic effect; Transgenic Mice; Transgenic Organisms; Work; amyloid formation; base; cell type; clinically relevant; cytokine; cytotoxic; cytotoxicity; druggable target; hyaluronan synthase 1; in vivo; inhibitor; islet; islet amyloid polypeptide; macrophage; migration; military veteran; monocyte; new therapeutic target; novel; preservation; prevent; promoter; recruit; response

The pathophysiology of type 2 diabetes (T2D) requires islet β-cell dysfunction in the presence of insulin resistance. Islet amyloid is a pathological feature which occurs in the vast majority of individuals with type 2 diabetes (T2D) and contributes to the β-cell dysfunction and loss that characterizes the disease. Islet amyloid arises due to aggregation of the β-cell peptide islet amyloid polypeptide (IAPP), which is deposited in the islet extracellular matrix (ECM), between β cells and endothelial cells; the latter being the major cell type of the islet capillary network. Despite decades of work in the field of islet amyloid, no studies have examined whether (i) islet amyloid is toxic to islet endothelial cells or (ii) whether islet endothelial cells contribute to the toxic effects of islet amyloid. Our preliminary data suggest that both occur, and that the ECM molecule hyaluronan (HA) produced by the islet endothelial cell is a major mediator of amyloid-induced toxicity. Specifically, we have found that islet amyloid deposition in vivo is detrimental to islet endothelial cells, while treatment of isolated primary islet endothelial cells in vitro with amyloidogenic human IAPP (hIAPP) peptide is cytotoxic and results in HA production. Our preliminary data also show that HA enhances hIAPP fibril formation in vitro, which is in line with our observation that HA deposition is also present in in vivo amyloid deposits in islets from hIAPP transgenic mice and human T2D. We and others have shown that IAPP aggregation is proinflammatory, activating macrophages, and resulting in IL-1β production. However, the mechanism by which recruitment and priming of these macrophages occurs has not been established. Based on the literature, we propose that islet endothelial- derived HA provides this missing link and mediates macrophage chemotaxis, adhesion and priming. Based on these data, we hypothesize that hIAPP aggregation induces HA production from islet endothelial cells, which exacerbates hIAPP fibril formation and cytotoxicity, and acts to recruit and prime macrophages. We will address this hypothesis in the following specific aims: Specific Aim 1: To determine the mechanism by which HA exacerbates hIAPP aggregation, and thereby β-cell toxicity, in vitro. In this aim, we will determine how HA alters hIAPP peptide conformation, increases hIAPP fibril formation and whether hIAPP fibrils formed in the presence of HA show increased β-cell cytotoxicity. Specific Aim 2: To determine whether islet endothelial cell-derived HA is necessary for migration, adhesion and priming of monocytes/macrophages under conditions of islet amyloid formation. We will determine whether HA produced by hIAPP-treated islet endothelial cells results in monocyte chemotaxis, macrophage adhesion, priming of the NLRP3 inflammasome and fragmentation of HA. We will also determine if cytokines produced by macrophages or endothelial cells in response to hIAPP treatment, are necessary for HA production, and which hyaluronan synthase isoforms contribute to this process. Specific Aim 3: To determine whether blockade of HA production or fragmentation inhibits islet amyloid deposition, and thereby its detrimental effects on β cell mass and function in vivo. hIAPP transgenic and non- transgenic mice will be treated with the HA synthesis inhibitor 4-methylumbelliferone or bred to introduce a macrophage-specific deletion of hyaluronidase 1 (to inhibit HA fragmentation). These mice will then be followed for one year. We expect that reducing HA synthesis/fragmentation will reduce islet amyloid deposition and/or its toxic and proinflammatory effects, thus preserving β-cell function and mass in vivo. These studies will provide novel data regarding the role of HA mediating β-cell cytotoxicity. These are clinically relevant studies; islet amyloid is a feature of ~90% of subjects with T2D, a disease state that disproportionately affects the veteran population, and HA represents a truly “druggable” target.!

2型糖尿病（T2 D）的病理生理学要求在胰岛素存在下胰岛β细胞功能障碍 阻力胰岛淀粉样蛋白是一种病理特征，发生在绝大多数2型糖尿病患者中， 糖尿病（T2 D）是一种严重的疾病，并导致β细胞功能障碍和损失，这是该疾病的特征。胰岛淀粉样 由于β细胞肽胰岛淀粉样多肽（IAPP）的聚集而产生， 细胞外基质（ECM），在β细胞和内皮细胞之间;后者是胰岛的主要细胞类型 毛细血管网尽管在胰岛淀粉样蛋白领域进行了数十年的工作，但没有研究检查（i） 胰岛淀粉样蛋白对胰岛内皮细胞有毒或（ii）胰岛内皮细胞是否促成毒性作用 胰岛淀粉样蛋白我们的初步数据表明，两者都发生，ECM分子透明质酸（HA） 由胰岛内皮细胞产生的β-淀粉样蛋白是淀粉样蛋白诱导的毒性的主要介质。 具体地说，我们发现体内胰岛淀粉样蛋白沉积对胰岛内皮细胞有害， 在体外用淀粉样蛋白生成人IAPP（hIAPP）肽处理分离的原代胰岛内皮细胞， 细胞毒性并导致HA产生。我们的初步数据还表明，HA增强hIAPP原纤维的形成 这与我们观察到的HA沉积也存在于体内淀粉样蛋白沉积中一致， 来自hIAPP转基因小鼠和人T2 D的胰岛。 我们和其他人已经表明，IAPP聚集是促炎性的，激活巨噬细胞， 导致IL-1β产生。然而，这些细胞的募集和启动机制 巨噬细胞的发生尚未建立。基于文献，我们提出胰岛内皮细胞- 衍生的HA提供了该缺失的环节并介导巨噬细胞趋化性、粘附和引发。 基于这些数据，我们假设hIAPP聚集诱导胰岛产生HA， 内皮细胞，其加剧hIAPP原纤维形成和细胞毒性，并用于募集和 原始巨噬细胞 我们将在以下具体目标中讨论这一假设： 具体目的1：确定HA加剧hIAPP聚集的机制，从而加剧β-细胞凋亡。 体外毒性试验。在这个目标中，我们将确定HA如何改变hIAPP肽构象，增加hIAPP 原纤维形成以及在HA存在下形成的hIAPP原纤维是否显示出增加的β-细胞细胞毒性。 具体目的2：确定胰岛内皮细胞衍生的HA是否是迁移、粘附和增殖所必需的。 在胰岛淀粉样蛋白形成的条件下引发单核细胞/巨噬细胞。我们将决定是否HA 由hIAPP处理的胰岛内皮细胞产生的细胞因子导致单核细胞趋化性，巨噬细胞粘附， NLRP 3炎性体的引发和HA的片段化。我们还将确定细胞因子是否由 巨噬细胞或内皮细胞对hIAPP处理的响应是HA产生所必需的， 透明质酸合酶同种型有助于该过程。 具体目标3：确定阻断HA产生或片段化是否抑制胰岛淀粉样蛋白 沉积，从而对体内β细胞质量和功能产生有害影响。hIAPP转基因和非转基因 转基因小鼠将用HA合成抑制剂4-甲基伞形酮处理或繁殖以引入 透明质酸酶1的巨噬细胞特异性缺失（以抑制HA片段化）。然后这些老鼠 跟踪了一年。我们预期减少HA的合成/断裂将减少胰岛淀粉样蛋白的沉积 和/或其毒性和促炎作用，从而在体内保持β-细胞功能和质量。 这些研究将提供关于HA介导β细胞毒性作用的新数据。这些是 临床相关研究;胰岛淀粉样蛋白是约90%的T2 D受试者的特征，T2 D是一种疾病状态， 不成比例地影响退伍军人群体，HA代表了一个真正的“可下药”目标。