Erythrocytes induce proinflammatory endothelial activation in hypoxia

Area of Science:

• Cardiovascular Biology
• Cellular Physiology
• Respiratory Medicine

Background:

• Ambient hypoxia is known to trigger proinflammatory vascular responses.
• The precise mechanisms initiating these responses in systemic hypoxia remain unclear.
• Understanding these mechanisms is crucial for addressing hypoxia-related vascular dysfunction.

Purpose of the Study:

• To investigate the role of erythrocyte-derived hydrogen peroxide (H2O2) in initiating proinflammatory gene transcription in vascular endothelium during systemic hypoxia.
• To elucidate the signaling pathways involved in hypoxia-induced endothelial activation.
• To identify the cellular source of reactive oxygen species responsible for these vascular changes.

Main Methods:

• Exposure of mice and isolated perfused murine lungs to hypoxia (8% O2) for 4 hours.
• Analysis of bronchoalveolar lavage for leukocyte counts.
• Assessment of leukocyte adhesion receptors, reactive oxygen species, and protein tyrosine phosphorylation in freshly recovered lung endothelial cells (FLECs).
• Inhibition studies using extracellular catalase and erythrocyte removal.
• Evaluation of nuclear factor-kappa B (NF-κB) p65 subunit translocation and hypoxia-inducible factor-1α (HIF-1α) stabilization in FLECs.
• Investigation of hemoglobin binding to erythrocyte band 3 protein and subsequent H2O2 release.

Main Results:

• Hypoxia led to increased leukocyte counts in bronchoalveolar lavage.
• Expression of leukocyte adhesion receptors, reactive oxygen species, and protein tyrosine phosphorylation increased in FLECs.
• These hypoxia-induced effects were significantly inhibited by extracellular catalase and erythrocyte removal.
• Nuclear translocation of NF-κB p65 and HIF-1α stabilization in FLECs were observed only in the presence of erythrocytes.
• Hemoglobin binding to band 3 induced erythrocyte H2O2 release and NF-κB p65 translocation in FLECs.

Conclusions:

• Erythrocytes are a primary source of hydrogen peroxide (H2O2) during systemic hypoxia.
• Erythrocyte-derived H2O2 is responsible for initiating proinflammatory transcriptional responses in vascular endothelium.
• The binding of hemoglobin to band 3 on erythrocytes triggers H2O2 release, leading to endothelial activation via NF-κB signaling.