Extracellular beta-nicotinamide adenine dinucleotide (beta-NAD) promotes the endothelial cell barrier integrity via PKA- and EPAC1/Rac1-dependent actin cytoskeleton rearrangement.
J Cell Physiol
; 223(1): 215-23, 2010 Apr.
Article
en En
| MEDLINE
| ID: mdl-20054824
ABSTRACT
Extracellular beta-NAD is known to elevate intracellular levels of calcium ions, inositol 1,4,5-trisphate and cAMP. Recently, beta-NAD was identified as an agonist for P2Y1 and P2Y11 purinergic receptors. Since beta-NAD can be released extracellularly from endothelial cells (EC), we have proposed its involvement in the regulation of EC permeability. Here we show, for the first time, that endothelial integrity can be enhanced in EC endogenously expressing beta-NAD-activated purinergic receptors upon beta-NAD stimulation. Our data demonstrate that extracellular beta-NAD increases the transendothelial electrical resistance (TER) of human pulmonary artery EC (HPAEC) monolayers in a concentration-dependent manner indicating endothelial barrier enhancement. Importantly, beta-NAD significantly attenuated thrombin-induced EC permeability as well as the barrier-compromising effects of Gram-negative and Gram-positive bacterial toxins representing the barrier-protective function of beta-NAD. Immunofluorescence microscopy reveals more pronounced staining of cell-cell junctional protein VE-cadherin at the cellular periphery signifying increased tightness of the cell-cell contacts after beta-NAD stimulation. Interestingly, inhibitory analysis (pharmacological antagonists and receptor sequence specific siRNAs) indicates the participation of both P2Y1 and P2Y11 receptors in beta-NAD-induced TER increase. beta-NAD-treatment attenuates the lipopolysaccharide (LPS)-induced phosphorylation of myosin light chain (MLC) indicating its involvement in barrier protection. Our studies also show the involvement of cAMP-dependent protein kinase A and EPAC1 pathways as well as small GTPase Rac1 in beta-NAD-induced EC barrier enhancement. With these results, we conclude that beta-NAD regulates the pulmonary EC barrier integrity via small GTPase Rac1- and MLCP- dependent signaling pathways.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Asunto principal:
Arteria Pulmonar
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Citoesqueleto
/
Permeabilidad Capilar
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Actinas
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Proteínas Quinasas Dependientes de AMP Cíclico
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Proteína de Unión al GTP rac1
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Factores de Intercambio de Guanina Nucleótido
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Células Endoteliales
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NAD
Tipo de estudio:
Prognostic_studies
Límite:
Humans
Idioma:
En
Revista:
J Cell Physiol
Año:
2010
Tipo del documento:
Article
País de afiliación:
Estados Unidos