Abstract 5196: LAR Deficiency Impairs Endothelial Barrier Function and Enhances Susceptibility to Atherosclerosis
Vascular endothelium forms a selective semipermeable barrier that regulates vital physiological functions such as tissue-fluid hemostasis and host defense. Impairment of endothelial barrier function is an early step in vascular dysfunction, which ultimately leads to atherosclerosis. The endothelial adherens junction (AJ), a multi-protein complex consisting of VE-cadherin and its associated catenins: β-catenin, γ-catenin and p120-catenin, regulates endothelial barrier function. Protein tyrosine kinases and phosphatases (PTPs) regulate AJ integrity and vascular permeability by modulating protein tyrosine phosphorylation. To determine the role of leukocyte antigen-related (LAR) PTP in endothelial barrier function, we investigated whether LAR associates with and/or regulates tyrosine phosphorylation of AJ proteins and the paracellular pathway. LAR expression in human umbilical vein endothelial cells (HUVEC) and brain microvascular endothelial cells was localized to EC-EC boundaries. Knockdown of LAR impaired (by 129.8%), whereas overexpression enhanced (by 48%) barrier function in HUVEC, as measured by flux of 40 KDa FITC-Dextran through cell monolayers. Reciprocal co-immunoprecipitation and co-localization with double-label fluorescence microscopy demonstrated the interaction of LAR with VE-cadherin and β-catenin. When glutathione S-transferase-cLAR (cytosolic LAR) was incubated with HUVEC lysate or with purified recombinant cytosolic VE-cadherin, LAR directly bound to VE-cadherin. Glutathione S-transferase-cLAR also directly bound to β-catenin in HUVEC lysate. Overexpression of LAR decreased tyrosine phosphorylation of VE-cadherin and β-catenin in postconfluent HUVEC. Consistent with these findings, aortic endothelial permeability and bradykinin induced microvascular leakage in pial venules were increased in apoE−/−/LAR−/− mice compared with apoE−/− mice. Furthermore, apoE−/−/LAR−/− mice fed Western diet for 3 month had a significant increase in aortic atherosclerotic lesion formation compared with apoE−/− mice (p = 0.0198). Together these data indicate that LAR regulates endothelial barrier function and dysregulation of this phosphatase may result in vascular pathophysiology.