Molecular Adaptation of Pancreatic Beta-Cell Function During Endoplasmic Reticulum Stress
Kitiphongspattana, Kajorn
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https://hdl.handle.net/2142/84940
Description
Title
Molecular Adaptation of Pancreatic Beta-Cell Function During Endoplasmic Reticulum Stress
Author(s)
Kitiphongspattana, Kajorn
Issue Date
2004
Doctoral Committee Chair(s)
Gaskins, H. Rex
Department of Study
Nutritional Sciences
Discipline
Nutritional Sciences
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Health Sciences, Pathology
Language
eng
Abstract
Pancreatic beta-cells may balance glucose-stimulated insulin synthesis and secretion with the proteasome activity to regulate protein concentrations in the endoplasmic reticulum (ER). During this process, reactive oxygen species (ROS) may originate as byproducts from the oxidative protein folding machinery in the ER, especially during an unfolded protein response (UPR). Using a beta-cell line (MIN6) and mouse islets, endogenous production of ROS, the GSH/GSSG couple, and expression of genes involved in oxidative protein folding (i.e., ERO1 and PDI) and antioxidant defense were measured. Endogenous ROS production and ERO1 and PDI expression were enhanced during high glucose stimulation and a UPR activated by blocking the ER-associated degradation pathway with the proteasome inhibitor lactacystin or by tunicamycin. Although the GSH/GSSG couple (indicative of redox state) was unaffected after lactacystin exposure in high glucose, lactacystin significantly decreased the GSH/GSSG couple in lower glucose (2.8 and 5.6 mM). Notably, high glucose stimulation decreased the GSH/GSSG couple. Thiol-antioxidants and an enhanced expression of genes involved in thiol metabolism and antioxidant defense protected against ER-derived oxidative stress. Although the direct effects of nitric oxide (NO) are unclear, low concentrations of NO may function as a protective ER stress-activated redox signal. Real-time monitoring of NO provided direct and dynamic measurements of NO generated within MIN6 cells. Endogenous NO production was proportional to increased glucose stimulation and was stimulated further by lactacystin and tunicamycin. NO synthase inhibition prevented the induction of the stress-inducible transcription factor CHOP/GADD153 and repressed induction of ER stress-responsive genes, specifically those involved in ER protein folding, antioxidant defense, and thiol metabolism. NO-induced transcriptional upregulation of gamma-GCS, the rate limiting enzyme for GSH synthesis, likely contributed to the increased total GSH pool observed in lactacystin-treated islets. Electromobility shift assays indicate NO-induced transcriptional gamma-GCS upregulation occurred by the transcription factor Nrf2 binding to an antioxidant response element. Collectively, these data demonstrate beta-cells adapt to ER stress and support a NO-dependent regulated model for adjusting their intracellular redox state in response to increased insulin demand. Constant exposure to high glucose may constitutively activate a UPR, and contribute to the accumulation of ROS and chronic oxidative stress during hyperglycemia.
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