Abstract
In insulin-secreting cells, expression of NADPH oxidase (NOX), a potent source of ROS, has been reported, along with controversial findings regarding its function. Here, the role of NOXs was investigated: first by expression and cellular localization in mouse and human pancreatic islets, and then by functional studies in islets isolated from Nox isoform-specific knockout mice. Both human and mouse β-cells express NOX, in particular NOX2. With use of Nox isoform-specific knockout mice, functional analysis revealed Nox2 as the predominant isoform. In human islets, NOX2 colocalized with both insulin granules and endosome/lysosome membranes. Nox2-deficient islets stimulated with 22.8 mmol/L glucose exhibited potentiation of insulin release compared with controls, an effect confirmed with in vitro knockdown of Nox2. The enhanced secretory function in Nox2-deficient islets was associated with both lower superoxide levels and elevated cAMP concentrations. In control islets, GLP-1 and other cAMP inducers suppressed glucose-induced ROS production similarly to Nox2 deficiency. Inhibiting cAMP-dependent protein kinase reduced the secretory response in Nox2-null islets, although not in control islets. This study ascribes a new role for NOX2 in pancreatic β-cells as negative modulator of the secretory response, reducing cAMP/PKA signaling secondary to ROS generation. Results also show reciprocal inhibition between the cAMP/PKA pathway and ROS.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Cells, Cultured
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Cyclic AMP / agonists
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Cyclic AMP / antagonists & inhibitors
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Cyclic AMP / metabolism*
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / metabolism
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Gene Silencing
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Glucagon-Like Peptide 1 / metabolism
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Humans
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Insulin / metabolism*
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Insulin Secretion
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Insulin-Secreting Cells / cytology
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Insulin-Secreting Cells / drug effects
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Insulin-Secreting Cells / metabolism*
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Islets of Langerhans / cytology
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Islets of Langerhans / drug effects
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Islets of Langerhans / metabolism
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Isoenzymes / antagonists & inhibitors
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Isoenzymes / genetics
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Isoenzymes / metabolism
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Membrane Glycoproteins / antagonists & inhibitors
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Membrane Glycoproteins / genetics
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Membrane Glycoproteins / metabolism*
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Mice
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Mice, Inbred C57BL
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Mice, Knockout
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NADPH Oxidase 2
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NADPH Oxidases / antagonists & inhibitors
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NADPH Oxidases / genetics
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NADPH Oxidases / metabolism*
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Protein Kinase Inhibitors / pharmacology
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RNA, Small Interfering
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Reactive Oxygen Species / metabolism*
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Second Messenger Systems* / drug effects
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Secretory Vesicles / drug effects
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Secretory Vesicles / metabolism
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Tissue Culture Techniques
Substances
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Insulin
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Isoenzymes
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Membrane Glycoproteins
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Protein Kinase Inhibitors
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RNA, Small Interfering
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Reactive Oxygen Species
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Glucagon-Like Peptide 1
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Cyclic AMP
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CYBB protein, human
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Cybb protein, mouse
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NADPH Oxidase 2
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NADPH Oxidases
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Cyclic AMP-Dependent Protein Kinases