Apoptosis signal-regulating kinase-1 regulates thrombin-induced endothelial permeability

Thrombin-induced endothelial permeability is associated with various pathological conditions. Apoptosis signalregulating kinase-1 (ASK1), one of the upstream MAP3K, has been reported to be an important regulator of endothelial stress and apoptosis. Despite this, its role in endothelial permeability is unknown. The aim of this study was to determine the role of ASK1 in thrombin-induced endothelial permeability. To do so, a live cell monitoring system and transwell assay were used to evaluate in vitro endothelial permeability, while a Miles assay was used for in vivo permeability. Immunofluorescence and western blotting were used to visualize integrity of the junctions and phosphorylation of various proteins, respectively. We observed that in vivo thrombin-induced vascular permeability was attenuated in Ask1-/- mice. Pretreatment of human primary endothelial cells (ECs) with GS-4997 (ASK1 inhibitor) and deficiency of ASK1 in primary mouse lung ECs significantly attenuated the thrombin-induced endothelial permeability. Furthermore, in the presence of GS4997, the following were also significantly reduced: thrombin-induced para-cellular gap formation, VEcadherin proteolysis, and dislocation of VE-cadherin, JAM-A, and ZO1 from the junctions. Inhibition of ASK1 restored peripheral location of F-actin, similar to that induced by sphingosine-1-phosphate. These results suggest a unique role for ASK1 in regulating thrombin-induced endothelial permeability.

Automated summary

This study aimed to determine the role of ASK1 in thrombin-induced endothelial permeability. The results showed that the lack of ASK1 attenuated thrombin-induced vascular permeability in vivo. In addition, the presence of ASK1 inhibitor reduced thrombin-induced endothelial permeability and gap formation, and restored the normal location of proteins at the junctions.