TRPV2 channels facilitate pulmonary endothelial barrier recovery after ROS-induced permeability

Abstract

Reactive oxygen species (ROS), such as hydrogen peroxide (H₂O₂), are known signaling molecules that increase endothelial barrier permeability. In this study, we investigated the roles of redox-sensitive transient receptor potential (TRP) ion channels, TRPM2, TRPV2 and TRPV4, in H₂O₂-induced endothelial barrier dysfunction. Using primary human pulmonary microvascular endothelial cells (HPMEC), we employed impedance-based resistance measurements, Western blot, and immunofluorescence staining to assess the effects of H₂O₂ on the endothelial barrier. Exposure to sublytic concentrations of H₂O₂ caused an acute loss of endothelial barrier integrity, accompanied by the cleavage of vascular endothelial cadherin (VE-cadherin), which was also apparent after application of the TRPV2 activator cannabidiol. The inhibition of either TRPV2 with tranilast or a disintegrin and metalloprotease domain-containing protein 10 (ADAM10) with GI254023X significantly reduced H₂O₂-induced VE-cadherin cleavage, while TRPM2 inhibition by econazole significantly increased H₂O₂-driven VE-cadherin cleavage and blockage of TRPV4 showed no effect. Although inhibition of either TRPV2 or ADAM10 did not prevent the initial loss of barrier resistance upon H₂O₂ exposure, both were essential for the subsequent recovery of barrier integrity. Time-course immunofluorescence stainings revealed that HPMEC barrier recovery involved a transient localization of N-cadherin proteins at adherens junctions. This process of cadherin-switching did not occur upon inhibition of TRPV2 or ADAM10. Our results highlight a novel role for TRPV2 as a redox sensitive ion channels in the microvascular endothelium and provide insight into the mechanisms underlying pulmonary microvascular endothelial barrier recovery.