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

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          Abstract

          Basic fibroblast growth factor (bFGF) exerts angiogenic and mitogenic properties in human tissue. Since changes in ion currents modulate essential Ca<sup>2+</sup>-dependent intracellular pathways in endothelial cells, we have investigated a possible contribution of Ca<sup>2+</sup>-activated K<sup>+</sup> channels (BK<sub>Ca</sub>) on bFGF-induced endothelial cell proliferation. The patch-clamp technique was used to identify BK<sub>Ca</sub> 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<sub>Ca</sub> blocker iberiotoxin (IBX). Using single-channel recordings, we found characteristic BK<sub>Ca</sub> 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<sub>Ca</sub> 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<sub>Ca</sub> within 2 min in cell-attached patches, whereas bFGF (50 ng/ml) was not able to activate BK<sub>Ca</sub>. 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<sub>Ca</sub> 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.

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          Most cited references 5

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          Hypoxia stimulates human endothelial cells to release smooth muscle cell mitogens: role of prostaglandins and bFGF.

          Abnormal smooth muscle cell (SMC) proliferation is observed in several pathological situations such as atherosclerosis, pulmonary hypertension, and venous pathologies, resulting in a thickening of the vessel wall. If endothelial cells have been assumed to play a role in the triggering of this proliferation, no direct evidence is available. As ischemia is often linked to these situations, we exposed human umbilical vein endothelial cells (HUVEC) to hypoxia. The HUVEC-conditioned medium was then added to SMC and the proliferation of these cells was measured. We observed a pro-proliferative activity for SMC of the hypoxic HUVEC-conditioned medium but not of the normoxic HUVEC one. This pro-proliferative activity could not be inhibited if HUVEC were treated with cycloheximide but was blocked if the synthesis of prostaglandins by HUVEC was inhibited during hypoxia. PGD2, and especially PGF2 alpha at the concentration found in the hypoxic HUVEC-conditioned medium, were demonstrated to have a mitogenic effect on SMC. PGE2 also showed a pro-proliferative activity but at higher concentrations. In addition, the kinetics of increase in SMC proliferation induced by a mixture of the four prostaglandins at the corresponding concentrations was the same as the one observed with hypoxic HUVEC-conditioned medium. However, when tested on fibroblasts which do not respond to PGF2 alpha, hypoxic HUVEC-conditioned medium also had a pro-proliferative activity. In addition, anti-bFGF antibodies but not anti-PDGF blocked the mitogenic activity of this conditioned medium for SMC. Finally, the mitogenic effects of PGF2 alpha and of bFGF on SMC are additive. These results indicate that bFGF is probably also involved. These results indicate that these prostaglandins act in synergy with bFGF and are the molecules responsible for the pro-proliferative activity observed in hypoxic HUVEC-conditioned medium. We propose that these findings can explain the excessive growth of SMC in blood vessels following chronic ischemic situations.
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            Role of potassium channels in mitogenesis

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              Tamoxifen blocks both proliferation and voltage-dependent K+ channels of neuroblastoma cells

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                Author and article information

                Journal
                JVR
                J Vasc Res
                10.1159/issn.1018-1172
                Journal of Vascular Research
                S. Karger AG
                1018-1172
                1423-0135
                1998
                October 1998
                28 October 1998
                : 35
                : 5
                : 363-371
                Affiliations
                Departments of a Cardiology and Angiology, b Physiology, Justus Liebig University of Giessen, Germany
                Article
                25606 J Vasc Res 1998;35:363–371
                10.1159/000025606
                9789117
                © 1998 S. Karger AG, Basel

                Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher. Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug. Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.

                Page count
                Figures: 5, References: 39, Pages: 9
                Categories
                Research Paper

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