High-Throughput Luminescent Reporter of Insulin Secretion for Discovering Regulators of Pancreatic Beta-Cell Function
High-Throughput Luminescent Reporter of Insulin Secretion for Discovering Regulators of Pancreatic Beta-Cell Function
复制标题
DOI：
10.1016/j.cmet.2014.12.010
复制
发表时间：
2015-01-06
期刊：
影响因子：
29
通讯作者：
Altshuler, David
中科院分区：
文献类型：
--
作者：
Burns, Sean M.;Vetere, Amedeo;Altshuler, David
胰岛素分泌缺陷在2型糖尿病的发病机制中起着核心作用，然而驱动β细胞功能障碍的机制仍然知之甚少，并且保护葡萄糖依赖性胰岛素释放的治疗方法也不充分。我们报告了一种发光胰岛素分泌试验，通过将Gaussia荧光素酶插入胰岛素原的c肽部分，可以大规模研究β细胞功能。表达这种结构的β细胞系在标准条件下或受到细胞因子、脂肪酸或内质网毒素的胁迫时，都能密切相关地共分泌荧光素酶和胰岛素。我们对报告进行了高通量分析，并进行了1600种化合物的先导筛选，确定了几种抑制分泌的药物，以及葡萄糖增强分泌剂，这些药物已被证实在初级人类胰岛中具有活性。与传统的酶联免疫吸附试验相比，该试验所需的时间和费用减少了40倍，可以加速识别控制胰岛素分泌的途径和安全增强糖尿病β细胞功能的化合物。
Defects in insulin secretion play a central role in the pathogenesis of type 2 diabetes, yet the mechanisms driving beta-cell dysfunction remain poorly understood, and therapies to preserve glucose-dependent insulin release are inadequate. We report a luminescent insulin secretion assay that enables large-scale investigations of beta-cell function, created by inserting Gaussia luciferase into the C-peptide portion of proinsulin. Beta-cell lines expressing this construct cosecrete luciferase and insulin in close correlation, under both standard conditions or when stressed by cytokines, fatty acids, or ER toxins. We adapted the reporter for high-throughput assays and performed a 1,600-compound pilot screen, which identified several classes of drugs inhibiting secretion, as well as glucose-potentiated secretagogues that were confirmed to have activity in primary human islets. Requiring 40-fold less time and expense than the traditional ELISA, this assay may accelerate the identification of pathways governing insulin secretion and compounds that safely augment beta-cell function in diabetes.