title: Lipopolysaccharide-induced endothelial barrier breakdown is cyclic adenosine monophosphate dependent in vivo and in vitro.
abstract: OBJECTIVE
  To determine whether cyclic adenosine monophosphate (cAMP) is critically involved in lipopolysaccharide (LPS)-induced breakdown of endothelial barrier functions in vivo and in vitro.
  
  
  METHODS
  Experimental laboratory research.
  
  
  METHODS
  Research laboratory.
  
  
  METHODS
  Wistar rats and cultured human microvascular endothelial cells.
  
  
  METHODS
  Permeability measurements in single postcapillary venules in vivo and permeability measurements and cell biology techniques in vitro.
  
  
  RESULTS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: University of Würzburg, Institute of Anatomy and Cell Biology, Würzburg, Germany.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Schlegel
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Baumer
authors.forename: Detlev
authors.initials: D
authors.lastname: Drenckhahn
authors.forename: Jens
authors.initials: J
authors.lastname: Waschke