Effect of cAMP derivates on assembly and maintenance of tight junctions in human umbilical vein endothelial cells.
BACKGROUND
Endothelial tight and adherens junctions control a variety of physiological processes like adhesion, paracellular transport of solutes or trafficking of activated leukocytes. Formation and maintenance of endothelial junctions largely depend on the microenvironment of the specific vascular bed and on interactions of the endothelium with adjacent cell types. Consequently, primary cultures of endothelial cells often lose their specific junctional pattern and fail to establish tight monolayer in vitro. This is also true for endothelial cells isolated from the vein of human umbilical cords (HUVEC) which are widely used as model for endothelial cell-related studies.


RESULTS
We here compared the effect of cyclic 3'-5'-adenosine monophosphate (cAMP) and its derivates on formation and stabilization of tight junctions and on alterations in paracellular permeability in HUVEC. We demonstrated by light and confocal laser microscopy that for shorter time periods the sodium salt of 8-bromoadenosine-cAMP (8-Br-cAMP/Na) and for longer incubation periods 8-(4-chlorophenylthio)-cAMP (pCPT-cAMP) exerted the greatest effects of all compounds tested here on formation of continuous tight junction strands in HUVEC. We further demonstrated that although all compounds induced protein kinase A-dependent expression of the tight junction proteins claudin-5 and occludin only pCPT-cAMP slightly enhanced paracellular barrier functions. Moreover, we showed that pCPT-cAMP and 8-Br-cAMP/Na induced expression and membrane translocation of tricellulin.


CONCLUSIONS
pCPT-cAMP and, to a lesser extend, 8-Br-cAMP/Na improved formation of continuous tight junction strands and decreased paracellular permeability in primary HUVEC. We concluded that under these conditions HUVEC represent a feasible in vitro model to study formation and disassembly of endothelial tight junctions and to characterize tight junction-associated proteins.
Beese
Michaela
M
Division of Nephrology, Department of Internal Medicine, Hannover Medical School, Carl-Neuberg-Str 1, 30625 Hannover, Germany.
Wyss
Kristin
K
Haubitz
Marion
M
Kirsch
Torsten
T
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D062507:Claudin-5; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D062794:MARVEL Domain Containing 2 Protein; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D062793:Occludin; D021381:Protein Transport / Q000187:drug effects; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D014471:Umbilical Veins / Q000473:pathology; D015854:Up-Regulation
C107466:CLDN5 protein, human; D062507:Claudin-5; D062794:MARVEL Domain Containing 2 Protein; C516798:MARVELD2 protein, human; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases
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