LPS Inhibits Endothelin-1-Mediated eNOS Translocation to the Cell Membrane in Sinusoidal Endothelial Cells

• Published in: Microcirculation (New York, N.Y. 1994) Vol. 12; no. 5; pp. 433 - 442
• Main Authors: MERKEL, SANDRA M., KAMOUN, WALID, KARAA, AMEL, KORNESZCZUK, KATARZYNA, SCHRUM, LAURA W., CLEMENS, MARK G.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Informa UK Ltd 01.07.2005; Blackwell Publishing Ltd
• Subjects: Animals; caveolin-1; Cell Membrane - metabolism; Cells, Cultured; Endothelial Cells - drug effects; Endothelial Cells - metabolism; endothelin receptors; Endothelin-1 - metabolism; Endothelin-1 - physiology; Endothelium, Vascular - cytology; endotoxin; Inflammation; intracellular trafficking; Lipopolysaccharides - pharmacology; Liver Circulation; liver microcirculation; Male; Nitric Oxide Synthase - analysis; Nitric Oxide Synthase - metabolism; Nitric Oxide Synthase Type III; Protein Transport - drug effects; Rats; Rats, Sprague-Dawley; Vasoconstriction
• ISSN: 1073-9688; 1549-8719
• DOI: 10.1080/10739680590960377

Objective: The objectives of this study were to develop a model for studying endothelin-1-mediated eNOS regulation in cultured sinusoidal endothelial cells and determine the effect of endothelin-1 and endotoxin (LPS) on eNOS localization. Methods: Changes in caveolin-1, calmodulin, and eNOS expression were determined by western blot and densitometric analysis. Endothelin receptor expression and localization and the intracellular localization of eNOS and caveolin-1 were assessed by confocal microscopy. Results: Sinusoidal endothelial cells expressed caveolin-1 and calmodulin, and expression was altered in cultured and passaged cells. eNOS expression decreased significantly in 24-h cultured cells, with expression dropping below the level of detection in passaged cells. Both endothelin A and endothelin B receptors were expressed on the cell surface after 24 h in culture. In 24-h cultured cells, caveolin-1 was localized in the perinuclear region and cell membrane, while eNOS was predominantly localized in the perinuclear region, where it co-localized with caveolin-1. Endothelin-1 stimulated eNOS translocation to the cell membrane. Pretreatment with LPS markedly inhibited the endothelin-1-mediated eNOS translocation. Conclusions: These studies demonstrate an LPS-mediated uncoupling of endothelin receptor activation and eNOS translocation. This functional uncoupling may, in part, account for the hyperconstrictive effects of endothelin-1 during inflammatory conditions. Microcirculation (2005) 12, 433-442. doi:10.1080/10739680590960377