CD226 deficiency promotes glutaminolysis and alleviates mitochondria damage in vascular endothelial cells under hemorrhagic shock.

Hemorrhagic shock (HS) is common in clinical emergencies, leading to millions of deaths each year globally. CD226 is a costimulatory adhesion molecule expressed on both immune cells and endothelial cells (ECs) to regulate their metabolic activity and function. As endothelial dysfunction occurs after HS, the roles CD226 plays in vascular EC metabolism were investigated. CD226 Tek mice were adopted to achieve vascular EC-specific knockout of CD226, and subjected to HS modelling. Serum levels of crucial intermediate metabolites were evaluated through liquid chromatography-mass spectrometry analysis. Human umbilical vein ECs (HUVECs) were used to study the effects of CD226 under hypoxia in vitro. Seahorse analysis evaluated the cellular glycolysis and mitochondria bioenergetics. Results showed that CD226 deficiency in vascular ECs alleviated HS-induced intestinal damage and inflammatory response in mice. Animal studies indicated an improved energy metabolism when CD226 was knocked out in ECs after HS, as evidenced by enhanced glutamine-glutamate metabolism and decreased lactic acid levels. Glut-1 was upregulated in mouse vascular ECs after HS and HUVECs under hypoxia, combined with decreased CD226. Moreover, HUVECs with CD226 knockdown exhibited relieved mitochondrial damage and early apoptosis under hypoxia, whereas CD226 overexpression showed opposite effects. Seahorse analysis showed that downregulated CD226 significantly increased mitochondrial ATP production and glucose uptake in HUVECs under hypoxia. Additionally, Erk/PHD2 signaling-mediated HIF-1α/Glut-1 and HIF-2α/ASCT2 pathways were involved in CD226 regulation on HUVEC glutaminolysis after hypoxia. Hence, CD226 deficiency promotes bypass energy supply to vascular ECs under ischemic or hypoxic stress, to ameliorate the stress-mediated metabolic disturbance.