Mechanisms and Impact of Islet Vascular Inflammation in Diabetes

胰岛素从胰岛β细胞释放不足是2型糖尿病（T2 D）的关键缺陷。 胰岛是高度血管化的，并且这种血管系统对于向胰岛细胞的营养供应是必需的， 胰岛素递送到外周组织，并作为支持β细胞功能的丰富信号源。在人类T2 D中 在动物模型中，胰岛血管病变表现为内皮炎症和毛细血管扩张， 碎片化此外，在体外诱导这些胰岛血管异常足以损害胰岛素 释放和/或诱导β-细胞死亡。 我们已经确定人胰岛淀粉样多肽（hIAPP）聚集作为胰岛细胞增殖的介导剂， 血管病变hIAPP源性淀粉样蛋白沉积在β细胞和胰岛之间的细胞外基质中 几乎所有T2 D患者的毛细血管。我们现在发现hIAPP聚集导致胰岛内皮细胞 细胞毒性和炎症，毛细血管扩张和损失。使用RNA-Seq，我们将Clec 14 a鉴定为潜在的 hIAPP诱导的胰岛内皮损伤的分子介质。Clec 14 a是血管C型 凝集素家族，其胰腺表达仅限于内皮细胞。Clec 14 a缺乏症 导致微血管不稳定，炎症和细胞死亡，但从来没有 在胰腺中进行研究。我们现在将确定Clec 14 a的缺失是否会加剧胰岛内皮对 hIAPP在体外增加对hIAPP诱导的胰岛血管病变的易感性并加重β细胞 体内功能障碍。 毛细血管相关周细胞在正常微血管稳态中起着关键作用。在几种组织中， 包括胰岛，周细胞调节毛细血管张力和血流。周细胞覆盖也是必要的， 内皮完整性，周细胞-内皮相互作用防止内皮炎症和血管渗漏。 在非胰岛组织中，这种相互作用由旁分泌信号和细胞-细胞接触控制，其中 clec 14 a只是一个调解人。内皮-周细胞相互作用的破坏是血管不稳定的基础 以及炎症，这些炎症导致糖尿病视网膜病变和肾纤维化的发展。周细胞脱离/丢失 也发生在T2 D胰岛中。然而，控制胰岛内皮细胞-周细胞附着的机制， 这些在T2 D中被破坏，因此对β细胞功能的影响尚未研究。这项建议 将填补这一重大知识缺口。我们的初步数据表明，hIAPP聚集导致胰岛 周细胞脱落和变性。我们的数据和其他人的数据显示，这包括hIAPP的直接毒性作用 对微血管周细胞的影响，导致控制内皮细胞-周细胞串扰的关键信号丢失。这 该提案将解决以下假设：胰岛内皮细胞-周细胞相互作用的破坏， 部分由Clec 14 a缺陷介导，是hIAPP诱导的胰岛血管病变的基础，并有助于 β细胞功能障碍。我们提出三个目标来解决这个假设： 目标1.为了确定Clec 14 a缺陷是否介导聚集的血管内皮细胞的有害血管效应， 在体外，hIAPP诱导胰岛血管病变和β细胞功能障碍，并在体内加剧hIAPP诱导的胰岛血管病变和β细胞功能障碍。 目标2.为了确定胰岛内皮细胞-周细胞相互作用在介导的有害作用中的作用， 体外hIAPP聚集。 目标3。为了确定聚集的hIAPP对离体胰岛周细胞功能的影响，并鉴定胰岛β细胞功能。 内皮细胞/周细胞衍生的胰岛素释放受损和β细胞死亡的介质。 这些新的研究将是第一个调查Clec 14 a在胰腺中的作用， 在正常条件下和hIAPP聚集后控制胰岛内皮细胞-周细胞相互作用。 这项工作具有重要意义，因为它可能导致开发新的血管靶向疗法， 细胞功能障碍，也可能与影响T2 D中许多组织的微血管疾病有关。

Insufficient release of insulin from the pancreatic islet β cell is a critical defect in type 2 diabetes (T2D). Pancreatic islets are highly vascularized, and this vasculature is essential for nutrient supply to islet cells, insulin delivery to peripheral tissues and as a rich source of signals that support β-cell function. In human T2D and animal models, islet vasculopathy manifests as endothelial inflammation and capillary dilation and fragmentation. Moreover, induction of these islet vascular abnormalities in vitro is sufficient to impair insulin release and/or induce β-cell death. We have identified human islet amyloid polypeptide (hIAPP) aggregation as a mediator of islet vasculopathy. hIAPP-derived amyloid deposits accumulate in the extracellular matrix between β cells and islet capillaries in almost all patients with T2D. We now show that hIAPP aggregation results in islet endothelial cell cytotoxicity and inflammation, capillary dilation and loss. Using, RNA-Seq we identified Clec14a as a potential molecular mediator of hIAPP-induced islet endothelial damage. Clec14a is a member of the vascular C-type lectin family, whose pancreatic expression is restricted to endothelial cells. Clec14a deficiency has been shown to result in microvascular destabilization, inflammation and cell death in several tissues but has never been studied in the pancreas. We will now determine if loss of Clec14a exacerbates the islet endothelial response to hIAPP in vitro and increases susceptibility to hIAPP-induced islet vasculopathy and exacerbates β-cell dysfunction in vivo. Capillary-associated pericytes play a critical role in normal microvascular homeostasis. In several tissues, including islets, pericytes regulate capillary tone and blood flow. Pericyte coverage is also necessary for endothelial integrity, pericyte-endothelial interactions prevent endothelial inflammation and vessel leakage. This interaction, in non-islet tissues, is governed both by paracrine signals and cell-cell contact, of which Clec14a is just one mediator. Disruption of endothelial-pericyte interactions underlies the vascular instability and inflammation that drives development of diabetic retinopathy and renal fibrosis. Pericyte detachment/loss also occurs in T2D islets. However, the mechanisms that govern islet endothelial-pericyte attachment, how these are disrupted in T2D and the consequent impact on β-cell function have not been studied. This proposal will address this major knowledge gap. Our preliminary data show that hIAPP aggregation leads to islet pericyte detachment and degeneration. Our data and others’ show this includes a direct toxic effect of hIAPP on microvascular pericytes, resulting in loss of key signals that govern endothelial cell-pericyte cross-talk. This proposal will address the hypothesis that disruption of islet endothelial cell-pericyte interactions, mediated in part by Clec14a deficiency, underlies hIAPP-induced islet vasculopathy and contributes to β-cell dysfunction. We propose three aims to address this hypothesis: Aim 1. To determine whether Clec14a deficiency mediates the deleterious vascular effects of aggregated IAPP in vitro and exacerbates hIAPP-induced islet vasculopathy and β-cell dysfunction in vivo. Aim 2. To determine the role of islet endothelial cell-pericyte interactions in mediating deleterious effects of hIAPP aggregation in vitro. Aim 3. To determine the impact of aggregated hIAPP on islet pericyte function ex vivo and identify islet endothelial cell/pericyte-derived mediators of impaired insulin release and β-cell death in vitro. These novel studies will be the first to investigate Clec14a in the pancreas, and to interrogate mechanisms governing islet endothelial cell-pericyte interactions under normal conditions and following hIAPP aggregation. This work is significant as it could lead to development of new vascular-targeted therapies which could treat β- cell dysfunction and may also be relevant for the microvascular disease which affects many tissues in T2D.