HIF-2α regulates hypoxia-induced strengthening of the human endothelial barrier by stabilizing adherens junctions

Endothelial cells (ECs) form a tight barrier to prevent vascular leakage. Although several studies reported that vascular leakage is increased during hypoxic conditions, other studies describe a protective effect. This raises the question how hypoxia specifically regulates adherens junction integrity and thus endothelial barrier function. Here, we investigated how hypoxia and the hypoxia-mimetic dimethyloxalylglycine (DMOG) affect adherens junction integrity and barrier function of human endothelial monolayers and which mechanisms are involved. The effect of hypoxia (1% O 2) on the endothelial barrier was measured by transendothelial passage of HRP and electrical impedance (ECIS) of endothelial monolayers subjected to hypoxia. Exposure to hypoxia improved endothelial barrier function, as compared to normoxia (20% O 2). This protective effect on the barrier was also present upon incubation with the hypoxia-mimetic DMOG. Hypoxia improved the endothelial barrier via a HIF-2α-(but not HIF-1α-) dependent process and increased accumulation of the adherens junction protein VE-cadherin at the cell margins. This was accompanied by decreased micromotion and lamellipodia formation by the cells, also pointing to stabilization of the junctions. Baseline contractile traction forces remained unaltered after hypoxia incubation, but were increased upon DMOG treatment, suggesting additional non-hypoxia related effects of DMOG. This study shows that hypoxia strengthens the endothelial barrier through stabilization of VE-cadherin and intercellular junctions. It is mediated by HIF-2α stabilization and is accompanied by reduced lamellipodia formation and motility of the endothelial monolayer. Conflicting data exist presently about the effect of hypoxia on endothelial barrier function. This study shows that hypoxia limits motility and lamellipodia formation in a human setting and strengthens the endothelial barrier through improved cell-cell interactions. The latter process is mediated through HIF-2α, but not HIF-1α, and results from stabilization of adherens junctions. The hypoxia-mimetic DMOG reflected these hypoxia effects partly, but displayed different contractile traction forces suggesting additional non-hypoxic effects.