Blockade of Ca ²⁺-Activated K ⁺ Channels Inhibits Proliferation of Human Endothelial Cells Induced by Basic Fibroblast Growth Factor

Abstract

Basic fibroblast growth factor (bFGF) exerts angiogenic and mitogenic properties in human tissue. Since changes in ion currents modulate essential Ca²⁺-dependent intracellular pathways in endothelial cells, we have investigated a possible contribution of Ca²⁺-activated K⁺ channels (BK_Ca) on bFGF-induced endothelial cell proliferation. The patch-clamp technique was used to identify BK_Ca and to study their modulation by bFGF in cultured endothelial cells of human umbilical cord veins (HUVEC). Cell counts of HUVEC were carried out on different days to analyze bFGF-induced cell proliferation and its influence by the specific BK_Ca blocker iberiotoxin (IBX). Using single-channel recordings, we found characteristic BK_Ca with a single-channel slope conductance of 170.3 ± 2.1 pS (n = 7), half-maximal activation at internal pCa = 5.7 (n = 5; test potential: 80 mV), and dose-dependent block by IBX (25–100 nmol/l). In cell-attached patches bFGF (50 ng/ml) caused a significant increase in the open-state probability (NPo) after 6 min at test potentials of 80 and 100 mV (n = 28; p < 0.001), respectively, which lasted up to 30 min. After preincubation with pertussis toxin (100 ng/ml; 4 h) bFGF superfusion did not cause a significant increase in BK_Ca activity until 25 min had passed (n = 20; p < 0.01). Addition of 100 nmol/l IBX to the pipette solution caused a total block of BK_Ca within 2 min in cell-attached patches, whereas bFGF (50 ng/ml) was not able to activate BK_Ca. When incubated with IBX (25–100 nmol/l) every 2 days, bFGF-induced proliferation of HUVEC was significantly decreased by 50 (–41%) and 100 nmol/l (–50%) IBX (n = 5; p < 0.001) after 7 days. We conclude that activation of BK_Ca by bFGF may play an important role in bFGF-induced proliferation of human endothelial cells and thus might be important in the process of angiogenesis and vascular remodelling.