Glucose-stimulated insulin secretion fundamentally requires H2O2 signaling by NADPH oxidase 4

Abstract

NADPH facilitates glucose-stimulated insulin secretion (GSIS) in pancreatic islet (PI) b-cells by an as yet unknown mechanism. We found NADPH oxidase, isoform-4 (NOX4), to be the major producer of cytosolic H<sub>2</sub>O<sub>2</sub>, essential for GSIS, while the increase in ATP/ADP alone was insufficient. The fast GSIS phase was absent in PIs from NOX4-null, b-cell-specific knockout mice (NOX4bKO) (not NOX2KO), and NOX4-silenced or catalase-overexpressing INS-1E cells. Lentiviral NOX4 overexpression or H<sub>2</sub>O<sub>2</sub> rescued GSIS in PIs from NOX4bKO mice. NOX4 silencing suppressed Ca<sup>2+</sup> oscillations and the patch-clamped ATP-sensitive potassium channel (K<sub>ATP</sub>) opened more frequently at high glucose. Mitochondrial H<sub>2</sub>O<sub>2</sub>, decreasing upon GSIS, provided an alternative redox signaling when 2-oxo-isocaproate or fatty acid oxidation formed superoxide by electron-transport flavoprotein:Q-oxidoreductase. Unlike GSIS, this ceased with mitochondrial antioxidant SkQ1. Both NOX4KO and NOX4bKO strains exhibited impaired glucose tolerance and peripheral insulin resistance. Thus the redox signaling previously suggested to cause b-cell-self-checking – hypothetically induces insulin resistance when absent. In conclusion, ATP plus H<sub>2</sub>O<sub>2</sub> elevations constitute an essential switch-on signal of insulin exocytosis for glucose and branched-chain oxoacids as secretagogues (partly for fatty acids). Redox signaling could be impaired by cytosolic antioxidants, hence those targeting mitochondria should be preferred for clinical applications to treat (pre)diabetes at any stage.