Trimethylamine N-oxide (TMAO) disrupts endothelial junction integrity through VE-cadherin Tyr658 phosphorylation in vitro.

Endothelial dysfunction induced by the accumulation of uremic toxins is a key contributor to cardiovascular disease (CVD) in patients with chronic kidney disease (CKD), as increased endothelial permeability represents an early and critical event in vascular injury and disease progression. This study evaluated the effects of trimethylamine N-oxide (TMAO), a low-molecular-weight uremic toxin, on endothelial junction integrity. Endothelial cells (EA.hy 926) were treated with TMAO at normal (2.83 mg/L) and uremic (7.49 mg/L) concentrations for 24 h. Cell viability was assessed using the MTT assay. VE-cadherin expression and phosphorylation were analyzed by western blotting and confirmed by confocal immunofluorescence. Endothelial permeability was determined by FITC-dextran passage across transwell inserts, and cell adhesion to extracellular matrix proteins was evaluated using fibronectin-coated plates. Our results demonstrate that TMAO reduced cell viability and disrupted endothelial cell-cell junctions by decreasing VE-cadherin protein levels and increasing its phosphorylation at tyrosine 658, resulting in increased endothelial permeability. TMAO also reduced endothelial adhesion to fibronectin, a key extracellular matrix protein. Notably, TMAO did not alter the expression of VE-cadherin-associated proteins, including p120-catenin and β-catenin. These findings provide mechanistic insight into the direct toxic effects of TMAO on endothelial barrier function and identify endothelial junctions as potential therapeutic targets to mitigate cardiovascular risk in CKD.