2000 Fiscal Year Final Research Report Summary
Regulation of pancreatic β-cell function by glutathione
| Project/Area Number |
10470041
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Pathological medical chemistry
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| Research Institution | Nagasaki University |
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Principal Investigator |
KONDO Takahito Nagasaki University of Medicine, Professor, 医学部, 教授 (00158908)
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| Co-Investigator(Kenkyū-buntansha) |
KIJIMA Hiroshi Tokai University, School of Medicine, Research Associate, 医学部, 助手 (90204859)
YAMASHITA Shunichi Nagasaki University, School of Medicine, Professor, 医学部, 教授 (30200679)
AKAZAWA Shouichi Nagasaki University, School of Medicine, Associate Professor, 医学部・附属病院, 助教授 (10145261)
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| Project Period (FY) |
1998 – 2000
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| Keywords | glutathione / pancreatic β-cells / γ-glutamylcysteine synthetase / insulin / hammerhead ribozyme |
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| Research Abstract |
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)
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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
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