Expression of Zonula Occludens and Adherens Junctional Proteins in Human Venous and Arterial Endothelial Cells: Role of Occludin in Endothelial Solute Barriers CHRISTOPHER G. KEVIL,* NAOTSUKA OKAYAMA,* STEVEN D. TROCHA, † THEODORE J. KALOGERIS, † LAURA L. COE,* ROBERT D. SPECIAN,* CHRISTOPHER P. DAVIS,* AND J. STEVEN ALEX ANDER* *Department of Molecular and Cellular Physiology, † Department of Surgery, LSU Medical Center, Shreveport, LA, USA ABSTRACT Objective: The purpose of this study was to correlate the expression of occludin and VE-cadherin with the solute barrier properties of arterial and venous en- dothelial monolayers. Methods: Immunofluorescent confocal and traditional microscopy were used to determine junctional protein localization in endothelium in vivo and in vitro respectively, and western and northern analysis used to determine protein and gene expression levels. Permeability of endothelial monolayers was examined under normal, low calcium, and cytochalasin-D treatment conditions. Antisense oligonucleotide experiments for occludin were performed to determine the con- tribution of occludin to solute barrier. Results: Occludin protein in endothelial monolayers is more concentrated in arterial junctions than in venous junctions both in vivo and in vitro. Arterial endothelial cells express 18-fold more occludin protein and nine times more occludin mRNA compared to venous endothelial cells. In vivo, both endothelial cells demonstrate VE-cadherin staining; and in vitro, only venous endothelial cells express VE-cadherin protein and mRNA. Occludin antisense experiments suggest that both arterial and venous barrier properties are due to these different amounts of occludin expression. Venous barrier was remarkably sensitive to low extracellular calcium, while arterial barrier was more sensitive to cytocha- lasin- D. Conclusions: These findings suggest strongly that arterial and venous endothe- lial barrier reflects the level of expression of different adhesion molecules and that modulation of these proteins, especially occludin, may regulate the level of endothelial solute barrier. KEY WORDS: permeability, junctions, occludin, VE-cadherin, gene regulation. INTRODUCTION Maintenance of the vascular solute barrier is a nor- mal homeostatic function of the endothelium, and endothelial pathology is frequently characterized by increased permeability in many forms of tissue in- jury such as inflammation, edema, and ischemia– reperfusion (44). It is widely accepted that solute barrier is established and maintained by proteins, which form the zonula occludens ( ZO-1 and ZO-2) and zonula adherens (cadherins; e.g., VE-cadherin and N-cadherin) junctional complexes (2,14,29). However, the way in which levels of expression of these proteins regulate endothelial barrier is not well understood. Cadherins, in particular VE-cadherin, help mediate intercellular adhesion and barrier in endothelial and Supported by National Institutes of Health Grants Nos. HL47615 and DK43785. For reprints of this article, contact Dr. J. Steven Alexander, De- partment of Molecular and Cellular Physiology, LSU-Medical Center Shreveport, 1501 Kings Highway, Shreveport, LA 71130, USA. Received 8 August 1997; accepted 6 February 1998 Microcirculation (1998) 5, 197–210 © 1998 Stockton Press All rights reserved 1073-9688/98 $12.00 http:/ / www.stockton-press.co.uk