Insulin Secretion in Hyperinsulinism Human Islets

项目摘要 从先天性高胰岛素血症（HI）婴儿胰腺中分离的人胰岛的研究 上一个资助周期为我们提供了独特的机会来检查KATPHI中的胰岛表型 整合功能、代谢组学和基因组学。我们的发现揭示了一个复杂的病理生理学， 初级KATP通道缺陷导致影响基因表达，燃料代谢， 以及胰岛素分泌的触发和放大途径。然而，许多关键问题， 解决HI治疗和理解正常生理机制的未满足需求 胰岛素分泌的问题仍然没有答案。我们特别感兴趣的是研究两种离子的作用 在正常的胰岛素分泌调节中，在HI胰岛中差异表达的通道及其 在HI的病理生理学中的潜在作用：TMEM 16 A，一种由ANO 1编码的Ca 2+激活的Cl-通道， Kv7.1，由KCNQ 1编码，其表达在KATPHI胰岛中显著上调， 从Beckwith Wiedemann综合征和HI儿童胰腺中分离的胰岛减少。在 初步研究我们发现这些通道的药理学调节改变胰岛素分泌。我们 总体假设是Kv7.1和TMEM 16 A在胰岛素的终止中起关键作用。 通过在静息时保持β细胞Vm超极化并在静息后促进β细胞Vm复极化来分泌 刺激.为了验证这一假设，我们提出了两个目的来研究Kv7.1和TMEM 16 A在细胞凋亡中的作用。 调节正常和HI胰岛中的胰岛素分泌。为了实现这些目标，我们将使用遗传和 调节正常和HI人和小鼠中这些通道活性的药理学方法 小岛我们将研究：1）TMEM 16 A和Kv7.1在静息和刺激时对β细胞Vm的贡献 2）TMEM 16 A和Kv7.1的激活和抑制对细胞溶质钙和胰岛素分泌的影响 3）TMEM 16 A和Kv7.1基因失活对葡萄糖代谢的影响 体内稳态和体内燃料刺激的胰岛素分泌以及使用遗传修饰的胰岛的分离的胰岛 小鼠模型。这项研究将扩大我们对HI病理生理学的理解， 确定这种毁灭性疾病的新遗传原因和潜在的新治疗靶点。的 这项研究也可能对理解进行性β细胞凋亡的机制有影响。 2型糖尿病的治疗方法

PROJECT SUMMARY The study of human islets isolated from the pancreas of infants with congenital hyperinsulinism (HI) during the previous funding cycle has afforded us the unique opportunity to examine the islet phenotype in KATPHI integrating function, metabolomics, and genomics. Our findings revealed a complex pathophysiology in which the primary KATP channel defect leads to secondary consequences affecting gene expression, fuel metabolism, and both the triggering and amplifying pathways of insulin secretion. However, many critical questions for addressing unmet needs for the treatment of HI and for the understanding of normal physiological mechanisms of insulin secretion remain unanswered. We are particularly interested in examining the role of two ion channels that are differentially expressed in HI islets in the normal regulation of insulin secretion and their potential role in the pathophysiology of HI: TMEM16A, a Ca2+-activated Cl– channel encoded by ANO1 which is markedly upregulated in KATPHI islets, and Kv7.1, encoded by KCNQ1, whose expression is markedly decreased in islets isolated from the pancreases of children with Beckwith Wiedemann syndrome and HI. In preliminary studies we found that pharmacological modulation of these channels alters insulin secretion. Our overall hypothesis is that both Kv7.1 and TMEM16A play critical roles in the termination of insulin secretion by keeping β-cell Vm hyperpolarized at rest and facilitating β-cell Vm repolarization after stimulation. To test this hypothesis, we propose two aims to examine the role of Kv7.1 and TMEM16A in the regulation of insulin secretion in normal and HI islets. To accomplish these aims we will use genetic and pharmacological approaches to modulate the activity of these channels in normal and HI human and mouse islets. We will examine: 1) the contribution of TMEM16A and Kv7.1 to β-cell Vm at resting and stimulated states; 2) the effect of TMEM16A and Kv7.1 activation and inhibition on cytosolic calcium and insulin secretion in normal human and mouse islets; 3) the effect of genetic inactivation of TMEM16A and Kv7.1 on glucose homeostasis in vivo and fuel-stimulated insulin secretion in vivo and in isolated islets using genetically modified mouse models. This study will expand our understanding of the pathophysiology of HI and will facilitate the identification of new genetic causes and potential new therapeutic targets for this devastating disease. The study may also have implications for the understanding of the mechanisms implicated in the progressive β-cell failure that leads to type 2 diabetes.