Regulation of spatial organization and cell-cell communication in the islet of Langerhans

朗格汉斯胰岛中不同内分泌细胞类型的协调激素分泌对葡萄糖稳态至关重要。在肥胖、胰岛素抵抗和糖尿病中，空间胰岛组织和细胞间通讯被破坏，导致协调激素调节的丧失。尽管胰岛组织和细胞间通讯在胰岛功能中起着至关重要的作用，但针对它们作为潜在治疗糖尿病的药物尚未开发出来。环形（Robo）受体是在胰岛中表达的细胞粘附蛋白。在胰岛素分泌过程中，机器人受体可以将细胞间接触与细胞骨架动力学和囊泡运输联系起来。在肥胖和糖尿病的小鼠和人类的胰岛中，Robo的表达严重减少。我们最近发现，小鼠β细胞中Robo1和Robo2的缺失会导致胰岛组织丢失和葡萄糖耐量受损。我们进一步证明，在β细胞选择性删除Robo时所看到的胰岛表型不是由于β细胞向α或δ细胞的转分化，也不是由于β细胞成熟缺失或β细胞死亡的结果。相反，我们的研究结果使我们假设β细胞中Robo的表达促进了正确的胰岛组织，这是内分泌细胞间通信和正确葡萄糖反应所必需的。根据这一假设，肥胖中Robo的下调破坏了胰岛组织，导致细胞间通讯的丧失，从而导致葡萄糖反应功能失调。了解Robo在β细胞中的表达如何调节胰岛组织和激素分泌，将为糖尿病的新药理学方法提供基础。我们将以两个具体目的来检验上述假设：在Aim 1中，我们将检验β细胞中Robo的缺失会损害胰岛素和胰高血糖素分泌的假设。我们将通过同步胰岛素分泌的活体可视化进一步验证Robo缺失破坏β细胞- β细胞功能偶联的假设。我们还将验证Robo通过介导内分泌细胞-细胞粘附调节激素分泌的假设。在Aim 2中，我们将确定β细胞中Robo的表达如何控制胰岛组织，并确定Robo配体Slit在这一过程中的作用。我们将进一步确定β细胞中的信号通路和下游Robo靶点，这可能解释了胰岛组织和葡萄糖耐量的缺陷。这些数据将在三个方面对糖尿病产生重要的转化影响：1)操纵Robo信号可以预防和恢复2型糖尿病患者胰岛组织和内分泌细胞-细胞通讯的恶化；2) Robo信号可用于在体外赋予人类多能干细胞衍生的胰岛样细胞簇正确的3D组织和细胞-细胞接触，以产生用于移植的真正的胰岛的无限来源。3)Robo受体及其细胞外配体Slit可用于生物工程方法，用于开发基质支架，以支持临床胰岛移植环境中尸体胰岛的结构完整性，存活和功能。

Coordinated hormone secretion from the different endocrine cell types in the islets of Langerhans is critical to glucose homeostasis. Spatial islet organization and cell-cell communication are disrupted in obesity, insulin resistance and diabetes, leading to loss of coordinated hormone regulation. Despite the critical roles of islet organization and cell-cell communication in islet function, drugs that target them as potential therapies to diabetes have not yet been developed. Roundabout (Robo) receptors are cell adhesion proteins expressed in the islet. Robo receptors can link cell-cell contact to cytoskeleton dynamic and vesicle transport in insulin secretion. The expression Robo in the islets of mice and humans are severely diminished in obesity and diabetes. We have recently found that deletion of Robo1 and Robo2 in beta cells in mice results in loss of islet organization and impaired glucose tolerance. We further demonstrated that the islet phenotype seen upon beta cell-selective deletion of Robo is not due to transdifferentiation of beta cells to alpha or delta cells, nor is it the result of loss of beta cell maturation or beta cell death. Rather, our results led us to hypothesize that expression of Robo in beta cells facilitates correct islet organization which is required for endocrine cell-cell communication and correct glucose response. According to this hypothesis, the downregulation of Robo in obesity disrupts islet organization, leading to loss of cell-cell communication, thus contributing to dysfunctional glucose response. Understanding how expression of Robo in beta cells regulates islet organization and hormone secretion will provide the basis for new pharmacological approaches to diabetes. We will test the above hypothesis with two specific aims: in Aim 1, we will test the hypothesis that deletion of Robo in beta cells impairs insulin and glucagon secretion. We will further test the hypothesis that deletion of Robo disrupts functional beta cell-beta cell coupling using intravital visualization of synchronized insulin secretion. We will also test the hypothesis that Robo regulates hormone secretion through mediating endocrine cell-cell adhesion. In Aim 2, we will determine how expression of Robo in beta cells controls islet organization, and determine the involvement of the Robo ligand, Slit, in this process. We will further identify signaling pathways and downstream Robo targets in beta cells which could account for the defects in islet organization and glucose tolerance. These data will have important impact on diabetes in three translational aspects: 1) Robo signaling may be manipulated to prevent and restore the deterioration in islet organization and endocrine cell-cell communication in type-2 diabetics; 2) Robo signaling may be used to confer correct 3D organization and cell-cell contact in human pluripotent stem cells-derived islet-like clusters in vitro, to generate an unlimited source of bona fide islets for transplantation, and 3) Robo receptors and their extracellular ligand, Slit, may be used in bioengineering approaches for developing matrix scaffolds that could support structural integrity, survival and function in cadaver islets in clinical islet transplantation settings.