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Glutaredoxin-1 Mediates NADPH-Dependent Stimulation of Calcium-Dependent Insulin Secretion

Department of Clinical Sciences (T.M.R., R.I., D.-Q.L., O.N., X.J., E.Z., U.B., E.R.) Islet Pathophysiology, Lund University, Clinical Research Centre 90-11, SE-20205 Malmö, Sweden; Institution of Endocrinology (D.-Q.L.), Tianjin University, 300070 Tianjin, China; Department of Pharmacology (O.N.), Alberta Diabetes Institute (ADI), 6040 Health Research Innovation Facility East, Edmonton, Alberta, Canada; and Department of Experimental Medical Science (L.S.), Cell Signalling, Lund University, BMC C11, SE-22100 Lund, Sweden

Address all correspondence to: Thomas M. Reinbothe, Department of Clinical Sciences, Islet Pathophysiology, Lund University, Clinical Research Centre 90-11, SE-20205 Malmö, Sweden. E-mail: thomas.reinbothe{at}med.lu.se. Address reprint requests to E. Renstrom. E-mail: erik.renstrom{at}med.lu.se.

Nicotinamide adenine dinucleotide phosphate (NADPH) enhances Ca²⁺-induced exocytosis in pancreatic β-cells, an effect suggested to involve the cytosolic redox protein glutaredoxin-1 (GRX-1). We here detail the role of GRX-1 in NADPH-stimulated β-cell exocytosis and glucose-stimulated insulin secretion. Silencing of GRX-1 by RNA interference reduced glucose-stimulated insulin secretion in both clonal INS-1 832/13 cells and primary rat islets. GRX-1 silencing did not affect cell viability or the intracellular redox environment, suggesting that GRX-1 regulates the exocytotic machinery by a local action. By contrast, knockdown of the related protein thioredoxin-1 (TRX-1) was ineffective. Confocal immunocytochemistry revealed that GRX-1 locates to the cell periphery, whereas TRX-1 expression is uniform. These data suggest that the distinct subcellular localizations of TRX-1 and GRX-1 result in differences in substrate specificities and actions on insulin secretion. Single-cell exocytosis was likewise suppressed by GRX-1 knockdown in both rat β-cells and clonal 832/13 cells, whereas after overexpression exocytosis increased by approximately 40%. Intracellular addition of NADPH (0.1 mM) stimulated Ca²⁺-evoked exocytosis in both cell types. Interestingly, the stimulatory action of NADPH on the exocytotic machinery coincided with an approximately 30% inhibition in whole-cell Ca²⁺ currents. After GRX-1 silencing, NADPH failed to amplify insulin release but still inhibited Ca²⁺ currents in 832/13 cells. In conclusion, NADPH stimulates the exocytotic machinery in pancreatic β-cells. This effect is mediated by the NADPH acceptor protein GRX-1 by a local redox reaction that accelerates β-cell exocytosis and, in turn, insulin secretion.