Abstract 5548: Blockade of KCa3.1 Calcium-activated Potassium Channels Prevents Endothelial Inflammation

Abstract

Introduction We previously reported that blockade of the intermediate-conductance calcium-activated potassium channel (KCa3.1) prevents the development of atherosclerosis in apoE −/− mice. Endothelial cell (EC) surface CD73, ecto-5′-nucleotidase, extracellularly produces adenosine and prevents EC inflammation. Therefore we hypothesized that KCa3.1 blockade suppresses EC inflammation by upregulating CD73. Methods Human coronary artery ECs were preconditioned by 24-hour exposure to laminar shear stress (LS; 15 dyn/cm 2 ), and were treated with tumor necrosis factor- α (TNF- α , 1 ng/ml) for an additional 4 hours under LS in the presence or absence of the KCa3.1 blocker TRAM-34 (100 nM) or the KCa2.3 blocker apamin (100 nM). Expression of KCa3.1, KCa2.3, CD73 and vascular cell adhesion molecule-1 (VCAM-1) transcripts was determined by real-time PCR. Results Preconditioning ECs with LS resulted in increased KCa3.1 and KCa2.3 expression (6.6±1.8 and 3.3±0.4 fold increase vs. static condition, p<0.05, n=4 –10, respectively). The LS preconditioning also increased CD73 expression (2.4±0.7 fold p<0.05, n=4 –10), whereas it decreased VCAM-1 expression (0.6±0.2 fold, p<0.05, n=7). TNF- α increased VCAM-1 expression (23.5±3.2 fold, p<0.001) with no change in CD73 expression (1.0±0.3 fold, p=ns) under static conditions. LS decreased TNF- α -induced increase in VCAM-1 expression and increased CD73 expression (17.0±1.1 and 1.9±0.5 fold, n=4, respectively). Addition of TRAM-34 enhanced the LS-induced changes in VCAM-1 and CD73 expressions with markedly lower VCAM-1 (9.0±3.0 fold, p<0.05 vs static + TNF- α ) and much higher CD73 expression (3.1±1.0 fold). In contrast, addition of apamin markedly exaggerated the VCAM-1 expression, while it increased CD73 expression as much as TRAM-34 did. Conclusions KCa3.1 blockade suppresses TNF- α -induced EC inflammation by increasing CD73 expression and decreasing VCAM-1 expression. EC KCa3.1 might therefore be one of the targets of anti-atherosclerosis therapy with KCa3.1 blockers.