PDF(Pubmed)
Abstract：
Islets of Langerhans release peptide hormones in controlled amounts and patterns to ensure proper maintenance of blood glucose levels. The overall release of the hormones is shaped by external factors and by autocrine and paracrine interactions occurring within the islets. To better understand what controls the secretion of islet-secreted peptides, and how these processes go awry in diabetes, methods to monitor the release of multiple hormones simultaneously are needed. While antibody-based assays are typically used, they are most often applied to quantification of a single hormone. Mass spectrometry (MS), on the other hand, is well suited for quantifying multiple hormones simultaneously but typically requires time-consuming separation steps with biological samples. In this report, response surface methodology was used to identify a set of optimal solid-phase extraction (SPE) conditions for the islet-secreted peptides, insulin, C-peptide, glucagon, and somatostatin. The optimized SPE method was used with multiple reaction monitoring and isotopically labeled standards to quantify secretion levels. Calibrations were linear from 0.5 to 50 nM with < 15% RSD peak area ratios. A microfluidic system was used to perfuse 30 human islets with different glucose conditions, and fractions were collected every 2 min for SPE-MS analysis. Results showed the release dynamics of the individual peptides, as well as patterns, such as positively and negatively correlated release and oscillations. This rapid SPE-MS method is expected to be useful for examining other peptide and small-molecule secretions from islets and could be applied to a number of other biological systems for investigating cellular communication.
摘要：
胰岛以受控的量和模式释放肽激素以确保血糖水平的适当维持。激素的整体释放由外部因素以及胰岛内发生的自分泌和旁分泌相互作用决定。为了更好地理解是什么控制胰岛分泌肽的分泌，以及这些过程如何在糖尿病中出错，需要同时监测多种激素释放的方法。虽然通常使用基于抗体的测定法，它们最常用于量化单一激素。质谱(MS),另一方面,非常适合同时定量多种激素，但通常需要耗时的生物样品分离步骤。在这份报告中,响应面方法用于确定胰岛分泌肽的一组最佳固相萃取（SPE）条件，胰岛素，C-肽，胰高血糖素,和生长抑素.优化的SPE方法与多个反应监测和同位素标记的标准一起使用以定量分泌水平。校准从0.5至50nM为线性的,具有<15%RSD峰面积比。使用微流体系统以不同的葡萄糖条件灌注30个人类胰岛，并且每2分钟收集级分用于SPE-MS分析。结果显示了单个肽的释放动力学，以及模式，如正相关和负相关的释放和振荡。这种快速的SPE-MS方法有望用于检查胰岛的其他肽和小分子分泌物，并可应用于许多其他生物系统以研究细胞通讯。
