7, 8-Dihydroxycoumarin (daphnetin) protects INS-1 pancreatic β-cells against streptozotocin-induced apoptosis

Background and objective: Daphnetin (7, 8-dihydroxycoumarin), a natural coumarin compound, is known to exhibit antioxidant and anti-inflammatrory effects. However, the underlying mechanisms of its antiapoptotic and antidiabetic effects yet not been examined. Therefore, the present work studied the antiapoptotic and anti-diabetic effects of daphnetin by in vitro experiments. Methods: The rat insulinoma (INS-1) cells were pre-treated with daphnetin at different concentrations (1, 10, 20 and 40 mu M) for 24 h followed by exposition to streptozotocin (STZ) (3 mM) for 12 h. Effects of daphnetin and STZ on INS-1 cells were determined by MTT assay, glucose stimulated insulin secretion (GSIS) assay, lipid peroxidation, antioxidant status (SOD, CAT, GPx, and GST) Apoptosis staining (DAPI, Hoechst 33342, AO/EB and ROS) was performed by fluorescence microscopy, and Bcl-2, Bax and NF-kappa B protein expression was detected by Western blotting. Results: MTT assay indicated that the viability of INS-1 cells was significantly reduced with exposure to STZ for 12 h as compared to control cells, while pre-treated with daphnetin for 24 h resulted in a significant improvement of cell viability. The effects daphnetin treatment in INS-1 cells on insulin secretion was tested and results showed that the pre-treatment of daphnetin could improve GSIS. Further, daphnetin pre-treatment significantly reduced the levels of lipid peroxidation markers and also improved antioxidant enzymes' activities in STZ-induced INS-1 cells. Western blotting assay revealed that daphnetin could suppress apoptosis through up-regulation of anti-apoptotic Bcl-2 protein expression and the down-regulation of pro-apoptotic Bax and nuclear factor NF-kappa B protein levels. Conclusion: The results showed that daphnetin might be used in treating diabetes due to its insulin stimulating property and subsequent regulation of apoptotic pathway. (C) 2016 Elsevier GmbH. All rights reserved.