Cytosolic and Plasma Membrane Circuitry of Beta Cell Redox Control

描述（由申请人提供）：质膜电子传递系统（PMET）是一种重要的线粒体外途径，通过NAD（P）H依赖性醌氧化还原酶（NQO 1）参与β细胞氧化还原状态和代谢的调节。PMET/NQO 1通路的功能是再氧化糖酵解衍生的NADH，并从细胞中输出质子和电子，产生第二信使过氧化氢。此外，NQO 1参与葡萄糖依赖的醌氧化还原循环过程，这也在葡萄糖刺激的β细胞胰岛素分泌（GSIS）中发挥重要作用。NAD激酶是一种新的NADPH调节酶，负责NADPH的从头产生并参与保护β细胞免受氧化应激。这些氧化还原控制的线粒体外途径对正常细胞功能至关重要，并且尚未在胰岛中表征。我们的初步数据证明了这些通路对β细胞功能，存活和GSIS的关键作用。我们将使用shRNA和腺病毒载体策略以及NQO 1敲除胰岛来分析这些组分在β细胞中间代谢和胰岛素分泌中的作用。我们将使用定制的集成电化学共聚焦成像平台来分析NQO 1和NAD激酶在胰岛对葡萄糖或其他刺激剂的响应中的作用，以同时研究胰岛代谢、呼吸和信号传导。这些研究将为氧化还原信号在调节胰岛素分泌中的机制提供更深入的了解，并为2型糖尿病治疗药物的开发确定新的靶点。

DESCRIPTION (provided by applicant): The plasma membrane electron transport system (PMET) is an important extra-mitochondrial pathway that participates in the regulation of beta cell redox status and metabolism via NAD (P) H-dependent quinone oxidoreductase, NQO1. The PMET/NQO1 pathway functions to reoxidize glycolytically-derived NADH and to export protons and electrons from the cell, generating the second messenger hydrogen peroxide. Furthermore, NQO1 participates in the glucose-dependent process of quinone redox cycling, which also plays an important role in glucose-stimulated insulin secretion (GSIS) from beta cells. NAD kinase, a novel NADPH- regulating enzyme, is responsible for the de novo production of NADPH and participates in the protection of beta cells from oxidative stress. These extra-mitochondrial pathways of redox control are critical for normal cell function, and have not been characterized in islets. Our Preliminary Data demonstrate the critical role of these pathways for beta cell function, survival, and GSIS. We will employ over-expression or knockdown using shRNA and adenoviral vectors strategies as well as NQO1 knockout pancreatic islets to analyze the role of these components in beta cell intermediary metabolism and insulin secretion. We will analyze the role of NQO1 and NAD kinase in the response of islets to glucose or other stimulatory agents using a custom- made integrated electrochemical-confocal imaging platform to simultaneously study islet metabolism, respiration, and signaling. These studies will provide greater insight into the mechanism of redox signaling in regulating insulin secretion and identify novel targets for the development of therapeutics for type 2 diabetes.