Background: Oxidative stress is a central mechanism of beta-cell glucotoxicity, but the underlying signaling pathways are only partially understood. Inhibitor of Differentiation (ID) proteins are transcriptional regulators induced by hyperglycemia in islets, but the mechanisms involved and their role in beta-cells are not clear. Here we investigated: 1) whether oxidative stress regulates ID expression in beta-cells, and 2) the role of ID expression in beta-cell pathophysiology under conditions of oxidative stress.

Methods: Insulin-secreting MIN6 beta-cells were cultured for 0-48h in the presence or absence of H₂O₂ (100-300 µM) or ribose (5-50 mM) to induce oxidative stress. RNA interference was used to silence the expression of Id1 and/or Id3. mRNA and protein levels were measured by real-time RT-PCR, western blot and immunocytochemistry. H₂O₂ levels were assessed by DCFDA probe and apoptosis by DNA fragmentation ELISA.

Results: Ribose and H₂O₂ treatment increased the mRNA levels of Id1-4 in a time- and concentration-dependent manner with parallel changes in the expression of antioxidant genes. Furthermore, immunostaining showed that ribose treatment increased ID1 and ID3 nuclear localisation. In ribose-treated cells, siRNA-mediated inhibition of Id1 and/or Id3 reduced the expression of multiple antioxidant genes, including heme oxygenase, glutathione peroxidase, superoxide dismutase, peroxiredoxins, sulfiredoxin and G6pdx. Additive effects were observed when both isoforms were inhibited. Glutathione peroxidase activity was also reduced after Id1/3 knockdown. These effects were accompanied by significant increases in H₂O₂ levels and beta-cell apoptosis.

Conclusion: We have identified Ids as a novel family of oxidative stress-responsive genes in beta-cells as well as an unexpected role for Id1 and Id3 in the modulation of redox status. The maintenance of an adequate antioxidant response by Ids may promote beta-cell survival under conditions of oxidative stress.