Glucocorticoid triggers endothelial cell ferroptosis via NOX4-mediated reactive oxygen species and lipid peroxidation

Abstract

Background: Glucocorticoids (GCs) are widely used in acute and critical illnesses, but long-term and high-dose use of GCs can cause several vascular side effects. However, the underlying mechanisms are not well-understood. Ferroptosis, a novel form of reactive oxygen species (ROS)-dependent cell death, is characterized by intracellular iron accumulation and lipid peroxidation. NADPH oxidase 4 (NOX4) is a major source of ROS. The roles of ferroptosis and NOX4 in GC-induced endothelial injury remain unknown.

Methods: Human umbilical vein endothelial cells (HUVECs) were exposed to varying concentrations of dexamethasone (DEX) to evaluate ferroptosis and NOX4 expression. Further mechanistic studies were conducted using NOX4-overexpressing adenovirus (Ad-NOX4), NOX4 small interfering RNA (siRNA), ferrostatin-1 (FER-1), and erastin.

Results: Our findings demonstrate that DEX induces ferroptosis in HUVECs. Inhibition of ferroptosis with FER-1 prevents DEX-induced reduction in HUVEC viability. Furthermore, DEX treatment increases NOX4 expression in HUVECs, and NOX4 overexpression with Ad-NOX4 promotes ferroptosis. NOX4 knockdown with siRNA suppresses DEX-induced ROS production, lipid peroxidation, and ferroptosis, thereby improving the viability, angiogenesis, and migration capacity of DEX-treated HUVECs. However, the protective effect of NOX4 knockdown is negated by the reactivation of ferroptosis with erastin.

Conclusion: GC-induced endothelial cell ferroptosis occurs through NOX4-mediated ROS production and lipid peroxidation, leading to cell death, impaired angiogenesis, and migration dysfunction. Inhibition of ferroptosis and NOX4 knockdown ameliorate GC-induced endothelial damage and dysfunction.

Keywords

Endothelial cell / Ferroptosis / Glucocorticoid / Lipid peroxidation / NADPH oxidase 4 / Reactive oxygen species

Conflict of interest statement

The authors declare no conflict of interest.

Author contributions

Lyu L and Fang L participated in the research design and wrote the article. Fang L, Chen J, and Li W performed the experiments and data analysis.

Funding

This work was supported by the China Postdoctoral Science Foundation (2019M652399), National Natural Science Foundation of China (No. 81600302), Shandong Natural Science Foundation (ZR2020QH336 and No. ZR2016HB01), Xinxin Green Valley Microcirculation Research Fund (XXLG07), and the Chinese EthnicMedicine Association (2023ZY086-46).

Ethical approval of studies and informed consent

Approval of all experiments was granted by the Ethics Committee of Qilu Hospital of Shandong University (NO. KYLL-202011-222).

Acknowledgments

None.

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