Inhibition of GSK3α/β promotes increased pulmonary endothelial permeability to albumin by reactive oxygen/nitrogen species.

Glycogen synthase kinase 3α/β (GSK3α/β) is a serine/threonine kinase that participates in numerous processes in many cell types. Importantly, the role of GSK3α/β in homeostatic maintenance of the pulmonary endothelial cell barrier to protein is not known. We tested the hypothesis that GSK3α/β regulates endothelial barrier function by measuring the permeability to albumin of a rat pulmonary microvessel endothelial cell monolayer (PMECM) treated with and without the selective GSK3α/β inhibitor SB 216763 (1.0, 5.0 and 10 uM) for 1 h. The treatment with the inhibitor SB 216763 caused a dose dependent decrease in phospho-β-catenin-Ser(33/37) levels indicating effective suppression of GSK3α/β. SB216763 caused an increase in both permeability to albumin and DCFDA (6-Carboxy-2',7'-Dichlorodihydrofluorescein Diacetate, Di(Acetoxymethyl Ester)) oxidation that were prevented by co-treatment with the anti-oxidant tiron or the nitric oxide synthase inhibitor L-NAME (Nω-nitro-l-arginine-methyl ester). In separate studies PMECMs were treated with the Akt inhibitor triciribine (12.5 uM) for 1 h to unmask Akt dependent constitutive suppression of GSK3α/β. Triciribine decreased phospho-GSK3α/β-Ser(21)/9 (i.e., the product of Akt) which was associated with an increase in phospho-β-catenin-Ser(33/37) (i.e., the product of GSK3α/β) indicating constitutive activity of Akt for GSK3α/β-Ser(21/9). The data indicates GSK3α/β inhibition causes increased endothelial monolayer protein permeability which is mediated by reactive oxygen/nitrogen species.