Regulation of pancreatic β-cell function by glutathione

活性氧（Reactive oxygen species，ROS）是组织损伤的重要因素，参与细胞内信号转导.谷胱甘肽（GSH）在清除活性氧以保持细胞持久稳定的生理功能方面发挥着核心作用。GSH还通过氧化还原调节调节细胞内信号通路。本研究旨在阐明GSH在β细胞功能调节中的作用。（1）胰岛β细胞的一个显著特点是细胞内GSH含量低，γ-谷氨酰半胱氨酸合成酶（γ-GCS）是合成GSH的关键酶。为了观察进一步降低GSH水平对胰腺β细胞的影响，我们构建了锤头状核酶，分别与γ-GCS重亚基（催化）和轻亚基（调节）的反义核苷酸偶联，并转染大鼠β细胞MIN-6。我们接下来研究了GSH合成减少对β细胞功能的影响。 ...更多信息 转染任何一种核酶均导致GSH合成减少，随后GSH浓度降低至对照水平的40%。B）GSH合成的持续抑制导致β-细胞胰岛素分泌的增强。c）GSH合成的抑制也增加了细胞内钙的浓度，d）转染胰岛素原报告基因的MIN-6细胞显示，抗-γ-GCS核酶可激活细胞中胰岛素原启动子的活性。(2)使用在高葡萄糖（27 mM）或正常葡萄糖（5.5 mM）中孵育的MIN-6细胞研究了对氧化应激的反应。MIN-6细胞高糖暴露于过氧化氢导致P13激酶/Akt介导的抗凋亡信号的损害。用GSH酯处理这些细胞可以恢复过氧化氢引起的细胞损伤，提示GSH在胰腺β细胞防御机制和细胞内信号中起重要作用。少

Reactive oxygen species (ROS) is a factor for tissue injury and participates in intracellular signal pathways. Glutathione (GSH) plays a central role in scavenging ROS to retain persistent and stable physiological functions in cells. GSH also regulates intracellular signal pathways through redox regulation. This study was addressed to clarify a role of GSH in the regulation of beta-cell function. The results obtained in this year are :(1) One of the striking characteristics of pancreatic β-cells is that the cells possess low concentration of GSH.Gamma-Glutamylcysteine synthetase (γ-GCS) is a key enzyme for the GSH synthesis. In order to see the effect of further lowing the levels of GSH on the pancreatic β-cell, we constructed hammerhead ribozymes coupled with antisense nucleotides for γ-GCS heavy subunit (catalytic) and light stubunit (regulatory), respectively, and transfected to rat β-cells, MIN-6. We next studied on the effect of decrease of GSH synthesis on the β-cell function.a) … More The transfection of either of the ribozymes caused of a decrease of the GSH synthesis followed by a decrease of the GSH concentration to 40% of the control levels.b) Persistent suppression of the GSH synthesis led an enhancement of insulin secretion from β-cells.c) The suppression of GSH synthesis also enhanced the intracellular concentration of calcium, which might be a factor of the increase of insulin secretion.d) MIN-6 cells transfected with proinsulin reporter sene showed that the promoter activity was stimulated in the cells with anti-γ-GCS ribozymes.(2) Response against oxidative stress was studied using MIN-6 cells incubated in high-glucose (27 mM) or normal-glucose (5.5 mM). Exposure of MIN-6 cells with high-glucose to hydrogen peroxide caused an impairment of anti-apoptotic signals mediated by P13-kinase/Akt. Treatment of these cells with GSH ester restored the cell damage induced by hydrogen peroxide, suggesting an important role of GSH in defense mechanism and intracellular signals in pancreatic β-cell. Less