Protein tyrosine phosphatase activity regulates endothelial cell-cell interactions, the paracellular pathway, and capillary tube stability.

Abstract

Protein tyrosine phosphorylation is tightly regulated through the actions of both protein tyrosine kinases and protein tyrosine phosphatases. In this study, we demonstrate that protein tyrosine phosphatase inhibition promotes tyrosine phosphorylation of endothelial cell-cell adherens junction proteins, opens an endothelial paracellular pathway, and increases both transendothelial albumin flux and neutrophil migration. Tyrosine phosphatase inhibition with sodium orthovanadate or phenylarsine oxide induced dose- and time-dependent increases in [14C]bovine serum albumin flux across postconfluent bovine pulmonary artery endothelial cell monolayers. These increases in albumin flux were coincident with actin reorganization and intercellular gap formation in both postconfluent monolayers and preformed endothelial cell capillary tubes. Vanadate (25 microM) increased tyrosine phosphorylation of endothelial cell proteins 12-fold within 1 h. Tyrosine phosphorylated proteins were immunolocalized to the intercellular boundaries, and several were identified as the endothelial cell-cell adherens junction proteins, vascular-endothelial cadherin, and beta-, gamma-, and p120-catenin as well as platelet endothelial cell adhesion molecule-1. Of note, these tyrosine phosphorylation events were not associated with disassembly of the adherens junction complex or its uncoupling from the actin cytoskeleton. The dose and time requirements for vanadate-induced increases in phosphorylation were comparable with those defined for increments in transendothelial [14C]albumin flux and neutrophil migration, and pretreatment with the tyrosine kinase inhibitor herbimycin A protected against these effects. These data suggest that protein tyrosine phosphatases and their substrates, which localize to the endothelial cell-cell boundaries, regulate adherens junctional integrity, the movement of macromolecules and cells through the endothelial paracellular pathway, and capillary tube stability.