NMDARs activation regulates endothelial ferroptosis via the PP2A-AMPK-HMGB1 axis

Area of Science:

• Cell Biology
• Neuroscience
• Biochemistry

Background:

• N-methyl-D-aspartate receptors (NMDARs) are critical ionotropic glutamate receptors involved in synaptic plasticity and neuronal function.
• Ferroptosis is a regulated form of cell death characterized by iron accumulation and lipid peroxidation, implicated in various pathologies.
• Vascular endothelial cells play a crucial role in maintaining vascular integrity and are susceptible to various forms of injury.

Purpose of the Study:

• To investigate whether NMDAR activation induces ferroptosis in vascular endothelial cells.
• To elucidate the underlying molecular mechanisms, including specific signaling pathways involved in NMDAR-mediated ferroptosis.
• To validate the findings both in vitro using human umbilical vein endothelial cells (HUVECs) and in vivo.

Main Methods:

• Utilized various detection approaches including western blotting, RNA-sequencing, and measurement of cellular iron content, lipid reactive oxygen species (LOS), MDA, and GSH.
• Employed pharmacological inhibitors (MK801, LB-100, glycyrrhizin) and activators (AICAR) to probe signaling pathways.
• Validated findings using ferroptosis inducers (errasin, RSL3) and inhibitors (Ferrostatin-1, Liproxstatin-1, Deferoxamine).

Main Results:

• NMDAR activation with L-glutamic acid (GLU) or NMDA significantly promoted ferroptosis in HUVECs, evidenced by increased iron content, MDA, PTGS2 expression, and decreased GPX4 expression and GSH.
• RNA-sequencing identified ferroptosis and SLC7A11 as key players in NMDA/GLU-mediated NMDAR activation.
• The PP2A-AMPK-HMGB1 pathway was identified as a crucial mediator of NMDAR activation-induced ferroptosis, confirmed through inhibitor and activator studies.
• In vivo studies corroborated that NMDAR activation induces ferroptosis and LOS production, which can be reversed by targeting this pathway.

Conclusions:

• N-methyl-D-aspartate receptor activation is a novel inducer of ferroptosis in vascular endothelial cells.
• The PP2A-AMPK-HMGB1 signaling pathway is critically involved in mediating NMDAR-induced endothelial cell ferroptosis.
• These findings reveal a new mechanism of endothelial cell injury regulated by NMDARs and ferroptosis, with potential therapeutic implications.