Activating autophagy and ferroptosis of 3‑Chloropropane‑1,2‑diol induces injury of human umbilical vein endothelial cells via AMPK/mTOR/ULK1

3-Chloropropane-1,2-diol (3-MCPD) is an internationally recognized food pollutant. 3-MCPD has reproductive, renal and neurotoxic properties. However, whether 3-MCPD induces human umbilical vein endothelial cell (HUVEC) injury has not been previously reported. In the present study, HUVECs were treated using 2 µg/ml 3-MCPD for 24 h at 37°C. The effects of 3-MCPD on HUVEC proliferation and cell cycle arrest, death and senescence were then assessed using Cell Counting Kit-8 (CCK-8), flow cytometry and β-galactosidase staining, respectively. Whether 3-MCPD induced ferroptosis was evaluated using JC-1 and FerroOrange staining and transmission electron microscopy. A small interfering RNA targeting AMPK was used to assess whether 3-MCPD promoted ferroptosis via AMPK signaling. The results demonstrated that 3-MCPD inhibited HUVEC proliferation in a dose-dependent manner and induced cell cycle arrest. Furthermore, 3-MCPD promoted senescence in HUVECs with elevated DNA damage and cell death. The CCK-8 results demonstrated that ferroptosis and autophagy inhibitors significantly reversed cell death caused by 3-MCPD. Moreover, 3-MCPD increased mitochondrial membrane potential, which indicated that 3-MCPD contributed to mitochondrial dysfunction. 3-MCPD also markedly increased intracellular Fe(2+) levels and lipid peroxidation in HUVECs. The present study assessed the underlying mechanism by which 3-MCPD activated autophagy and ferroptosis in HUVECs. The data demonstrated that 3-MCPD significantly increased phosphorylation levels of AMPK and unc-51 like autophagy activating kinase (ULK1) but significantly decreased phosphorylation of mTOR in HUVECs. Furthermore, silencing of AMPK significantly reversed the increase in autophagy, lipid peroxidation and Fe(2+) induced by 3-MCPD. In conclusion, 3-MCPD demonstrated a significant damaging effect on HUVECs via induction of autophagy and ferroptosis; such effects may be mediated by AMPK/mTOR/ULK1 signaling. To the best of our knowledge, the present study was the first to demonstrate the mechanism of 3-MCPD-induced vascular endothelial cell injury and lays a molecular foundation for the prevention of 3-MCPD-related vascular diseases.