Metformin Modulates Doxorubicin‐induced Senescence Phenotype in Endothelial Cells

Background Doxorubicin (DOX) is a cardiotoxic chemotherapeutic agent. In addition to cardiotoxicity, DOX treatment is associated with endothelial dysfunction and increased markers of vascular aging in cancer survivors, contributing to DOX-induced cardiovascular complications. Metformin, an anti-diabetic drug, has been shown to have anti-aging effects in different models of aging. Therefore, in our current work, we first established an in vitro model of DOX-induced senescence in EA.hy926 human endothelial-derived cells and Human Umbilical Vein Endothelial Cells (HUVECs). Then, we identified the effects of metformin on DOX-induced senescence phenotype. Methods Both HUVECs and EA.hy926 human endothelial-derived cells were treated with increasing DOX concentrations for 24 hours. Thereafter, DOX was removed and cells were incubated in DOX free media for another 72-120 hours. Protein expression of senescence markers were measured using Western blot analysis. Cell cycle analysis was assessed using flow cytometry. Senescence-associated beta-galactosidase (SA-beta GAL) activity was measured by SA-beta GAL assay. Next, cells were incubated with DOX +/- increasing metformin concentrations. Effects of metformin on DOX-induced senescence markers and cell cycle were assessed. To delineate the molecular mechanisms, the effects of metformin on major signaling pathways were determined. Results DOX significantly induced expression of senescence markers in endothelial cells in a concentration-dependent manner. Moreover, DOX caused cell cycle arrest in the G2/M phase and increased SA-beta GAL staining. Metformin corrected DOX-induced upregulation of senescence markers, which was associated with a significant inhibition of the JNK pathway. Intriguingly, metformin had no effect on DOX-induced cell cycle arrest and slightly corrected the increased expression of SA-beta GAL. Conclusions Our current work shows that metformin modulates DOX-induced senescence phenotype in endothelial cells, suggesting it may protect against DOX-induced vascular aging and endothelial dysfunction.