Hypoxia Postconditioning Attenuates Hypoxia-Induced Inflammation and Endothelial Barrier Dysfunction

In recent years, a number of studies have demonstrated that hypoxia reoxygenation (HR) induced by ischemia postconditioning (IPC) reduces endothelial barrier dysfunction and inflammation in various models. When HR occurs, the P38 mitogen-activated protein kinase (P38 MAPK) breaks down the endothelial barrier. But no study has clearly clarified the effect of hypoxia postconditioning (HPC) on P38 MAPK in human dermal microvascular endothelial cells. Therefore, we investigated the function of HPC on P38 MAPK during HR in vitro.

Human dermal microvascular endothelial cells were cultured in a hypoxic incubator for 8 h. Then cells were reperfused for 12 h (reoxygenation) or postconditioned by 5 min of reoxygenation and 5 min of re-hypoxia 3 times followed by 11.5 h reoxygenation. SB203580 was used as an inhibitor of P38 MAPK. Cell counting kit-8 assay kits were employed to detect cell activity. The corresponding levels of IL-6, IL-8 and IL-1β were examined via Enzyme-Linked ImmunoSorbent Assay. The endothelial barrier was evaluated using fluorescein isothiocyanate-dextran leakage assay. Western blot was used to detect claudin-5, phosphorylation of P38 MAPK (P-P38 MAPK) and P38 MAPK expression. Claudin-5 localization was studied by immunofluorescence.

HR induced endothelial barrier hyperpermeability, elevated inflammation levels, and increased the P-P38 MAPK. But HPC reduced cell injury and maintained the integrity of the endothelial barrier while inhibiting P-P38 MAPK and increasing expression of claudin-5. HPC redistributed claudin-5 in a continuous and linear pattern on the cell membrane.

HPC protects against HR induced downregulation and redistribution of claudin-5 by inhibiting P-P38 MAPK.