Expression, localization and regulation of NADPH oxidases in pancreatic beta cells

Reactive oxygen species (ROS) are short-lived and act in a site-specific manner, underscoring the importance of identifying the subcellular localization of their sources. ROS-generating NADPH oxidases (NOX) regulate pancreatic beta cell (dys)function. However, their subcellular localization and cytokine-mediated regulation in these cells remain largely unknown. We characterized the expression, subcellular localization and time-dependent cytokine-induced regulation of NOX isoforms in beta cells. Isoforms were studied via RT-qPCR, immunoblotting and immunofluorescence in rat islets and beta cell lines. Beta cells express DUOX1 and DUOX2 proteins and <i>Duoxa2</i> transcripts; lacking <i>Duoxa1</i> expression. In INS-1E cells, NOX1 and DUOX1 localize in the endoplasmic reticulum (ER); DUOX2 in insulin vesicles; and NOX2 and NOX4 in vesicles, ER and plasma membrane. In INS-1E, cytokines increased expression of <i>Nox1</i> and <i>Duox1</i> at 4-8 h (returning to baseline at 16 h) and <i>Nox2</i> and <i>p47phox</i> at 8 h (persisting until 24 h). <i>Duox(a)2</i>, <i>p67phox</i> and <i>p40phox</i> were downregulated and DUOX1 upregulated at 16-24 h. The absence of <i>Duoxa1</i> in beta cells might lead to DUOX1 mismatching, impairing its trafficking and activity. NOXs in beta cells are diverse in subcellular localization and cytokine-induced regulation, suggesting their isoform-specific involvement in beta cell function, stress and apoptosis.