Cardiac endothelial ischemia/reperfusion injury-derived protein damage-associated molecular patterns disrupt the integrity of the endothelial barrier

Abstract

Human cardiac microvascular endothelial cells (HCMECs) are sensitive to ischemia and vulnerable to damage during reperfusion. The release of damage-associated molecular patterns (DAMPs) during reperfusion induces additional tissue damage. The current study aimed to identify early protein DAMPs in human cardiac microvascular endothelial cells subjected to ischemia-reperfusion injury (IRI) using a proteomic approach and their effect on endothelial cell injury. HCMECs were subjected to 60 min of simulated ischemia and 6 h of reperfusion which can cause lethal damage. DAMPs in the culture media were subjected to liquid chromatography-tandem mass spectrometry proteomic analysis. The cells were treated with endothelial IRI-derived DAMP medium for 24 h. Endothelial injury was assessed by measuring lactate dehydrogenase activity morphological features and the expression of endothelial cadherin nitric oxide synthase (eNOS) and caveolin-1. The top two upregulated proteins DNAJ homolog subfamily B member 11 and pyrroline-5-carboxylate reductase 2 are promising and sensitive predictors of cardiac microvascular endothelial damage. HCMECs expose to endothelial IRI-derived DAMP the lactate dehydrogenase activity was significantly increased compared with the control group (10.15 ? 1.03 vs 17.67 ? 1.19 respectively). Following treatment with endothelial IRI-derived DAMPs actin-filament dysregulation and downregulation of vascular endothelial cadherin caveolin-1 and eNOS expressions were observed along with cell death. In conclusion the early protein DAMPs released during cardiac microvascular endothelial IRI could serve as novel candidate biomarkers for acute myocardial IRI. Distinct features of impaired plasma membrane integrity can help identify therapeutic targets to mitigate the detrimental consequences mediated of endothelial IRI-derived DAMPs. ? 2024 The Authors