Lipopolysaccharide-induced endothelial barrier breakdown is cyclic adenosine monophosphate dependent in vivo and in vitro

• Published in: Critical care medicine Vol. 37; no. 5; p. 1735
• Main Authors: Schlegel, Nicolas, Baumer, Yvonne, Drenckhahn, Detlev, Waschke, Jens
• Format: Journal Article
• Language: English
• Published: United States 01.05.2009
• Subjects: Animals; Antigens, CD - metabolism; Blotting, Western; Cadherins - metabolism; Capillary Permeability - drug effects; Capillary Permeability - physiology; Cells, Cultured; Cyclic AMP - metabolism; Disease Models, Animal; Endothelial Cells - drug effects; Endothelial Cells - metabolism; Enzyme Activation; Female; Histocytochemistry; Humans; Lipopolysaccharides - pharmacology; Male; Microcirculation - drug effects; Microcirculation - physiology; Probability; rac1 GTP-Binding Protein - metabolism; Random Allocation; Rats; Rats, Wistar; rho GTP-Binding Proteins - drug effects; rho GTP-Binding Proteins - metabolism; Sensitivity and Specificity; Statistics, Nonparametric
• ISSN: 1530-0293
• DOI: 10.1097/CCM.0b013e31819deb6a

To determine whether cyclic adenosine monophosphate (cAMP) is critically involved in lipopolysaccharide (LPS)-induced breakdown of endothelial barrier functions in vivo and in vitro. Experimental laboratory research. Research laboratory. Wistar rats and cultured human microvascular endothelial cells. Permeability measurements in single postcapillary venules in vivo and permeability measurements and cell biology techniques in vitro. We demonstrate that within 120 minutes LPS increased endothelial permeability in rat mesenteric postcapillary venules in vivo and caused a barrier breakdown in human dermal microvascular endothelial cells in vitro. This was associated with the formation of large intercellular gaps and fragmentation of vascular endothelial cadherin immunostaining. Furthermore, claudin 5 immunostaining at cell borders was drastically reduced after LPS treatment. Interestingly, activity of the small GTPase Rho A, which has previously been suggested to mediate the LPS-induced endothelial barrier breakdown, was not increased after 2 hours. However, activity of Rac 1, which is known to be important for maintenance of endothelial barrier functions, was significantly reduced to 64 +/- 8% after 2 hours. All LPS-induced changes of endothelial cells were blocked by a forskolin-mediated or rolipram-mediated increase of cAMP. Consistently, enzyme-linked immunosorbent assay-based measurements demonstrated that LPS significantly decreased intracellular cAMP. In summary, our data demonstrate that LPS disrupts endothelial barrier properties by decreasing intracellular cAMP. This mechanism may involve inactivation of Rac 1 rather than activation of Rho A.