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: 10265348
• 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/10265348
• 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/antagonist; 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 (T2D) 的病理生理学需要胰岛素存在下胰岛 β 细胞功能障碍 胰岛淀粉样蛋白抵抗是一种病理特征，发生在绝大多数 2 型个体中。 糖尿病 (T2D) 并导致 β 细胞功能障碍和损失，这是该疾病的特征。 由于沉积在胰岛中的 β 细胞肽胰岛淀粉样多肽 (IAPP) 的聚集而产生 细胞外基质 (ECM)，位于 β 细胞和内皮细胞之间；后者是胰岛的主要细胞类型 尽管在胰岛淀粉样蛋白领域进行了数十年的研究，但没有研究检验是否（i） 胰岛淀粉样蛋白对胰岛内皮细胞有毒，或 (ii) 胰岛内皮细胞是否会导致毒性作用 我们的初步数据表明这两种情况都会发生，而且 ECM 分子透明质酸 (HA) 也会发生。 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。 具体来说，我们发现体内胰岛淀粉样蛋白沉积会对胰岛内皮细胞造成困扰，而 用淀粉样变性人 IAPP (hIAPP) 肽在体外处理分离的原代胰岛内皮细胞 我们的初步数据还表明，HA 可以增强 hIAPP 原纤维的形成。 体外，这与我们的观察结果一致，即HA沉积也存在于体内淀粉样沉积物中 来自 hIAPP 转基因小鼠和人类 T2D 的胰岛。 我们和其他人已经证明 IAPP 聚集具有促炎作用，可激活巨噬细胞，并且 然而，这些细胞的募集和引发的机制。 巨噬细胞的发生尚未确定。根据文献，我们提出胰岛内皮细胞。 衍生的 HA 提供了这个缺失的环节并介导巨噬细胞趋化、粘附和启动。 基于这些数据，我们发现 hIAPP 聚集诱导胰岛产生 HA 内皮细胞，加剧 hIAPP 原纤维形成和细胞毒性，并起到招募和 主要巨噬细胞。 我们将通过以下具体目标来解决这一假设： 具体目标 1：确定 HA 加剧 hIAPP 聚集，从而加剧 β 细胞的机制 体外毒性 为了这个目的，我们将确定 HA 如何改变 hIAPP 肽构象，增加 hIAPP。 原纤维形成以及 hIAPP 原纤维是否在 HA 存在下形成显示出 β 细胞的细胞毒性增加。 具体目标 2：确定胰岛内皮细胞来源的 HA 是否是迁移、粘附和迁移所必需的 在胰岛淀粉样蛋白形成的条件下单核细胞/巨噬细胞的引发我们将确定HA是否形成。 由 hIAPP 处理的胰岛内皮细胞产生，导致单核细胞趋化、巨噬细胞粘附、 我们还将确定 NLRP3 炎性体的引发和 HA 的碎片是否产生细胞因子。 巨噬细胞或内皮细胞对 hIAPP 治疗作出反应，对于 HA 的产生是必需的，并且 乙酰透明质酸合酶亚型有助于这一过程。 具体目标 3：确定阻断 HA 产生或片段化是否会抑制胰岛淀粉样蛋白 沉积，从而对 hIAPP 转基因和非体内 β 细胞质量和功能产生不良影响。 转基因小鼠将接受 HA 合成抑制剂 4-甲基伞形酮治疗或进行繁殖以引入 然后，这些小鼠将被巨噬细胞特异性删除透明质酸酶 1（以抑制 HA 碎片）。 我们预计减少 HA 合成/断裂将减少胰岛淀粉样蛋白沉积。 和/或其毒性和促炎作用，从而保留体内β细胞的功能和质量。 这些研究将提供有关 HA 介导 β 细胞细胞毒性作用的新数据。 临床相关研究；胰岛淀粉样蛋白是约 90% 的 T2D 受试者的一个特征，这种疾病状态 HA 对退伍军人群体的影响尤为严重，HA 代表了真正的“可吸毒”目标。