A new signaling mechanism of hepatocyte growth factor-induced endothelial proliferation.
BACKGROUND
The hepatocyte growth factor (HGF) has been shown to promote endothelial cell proliferation. In this study, the signaling cascade responsible for the HGF-induced proliferation was examined.


METHODS
The proliferation of human umbilical cord vein endothelial cells (HUCVEC) was determined using cell counts. Changes of the membrane potential were analyzed using the fluorescence dye DiBAC. Intracellular cGMP-levels were measured by means of [3H]-cGMP-radioimmunoassay. Phosphorylation of the p42/p44 MAP-kinase (MAPK) and the endothelial nitric oxide synthase (eNOS) was analyzed by immunocytochemistry.


RESULTS
A dose-dependent (1-30 ng mL(-1)) increase of HUCVEC proliferation with a maximum at a concentration of 15 ng mL(-1) was induced by HGF. This effect was significantly reduced by the addition of the K+ channel blocker iberiotoxin (100 nmol L(-1)), eNOS inhibitor L-NMMA (300 micromol L(-1)), or the MEK inhibitor PD 98059 (20 micromol L(-1)). A HGF-induced hyperpolarization that was blocked by iberiotoxin was observed. In addition, HGF-induced activation of the eNOS was blocked by the K+ channel inhibitor. An increase of +101% MAPK phosphorylation was induced by HGF, which was blocked, if the cells were treated with L-NMMA (n = 20; P < 0.05), whereas HGF-induced phosphorylation of the eNOS was not affected by MEK inhibition.


CONCLUSIONS
Hepatocyte growth factor modulates endothelial K+ channels causing an activation of the eNOS; the increase of nitric oxide is necessary for the phosphorylation of the MAPK inducing the proliferation of HUCVEC.
Kuhlmann
C R W
CR
Department of Cardiology and Angiology, Justus-Liebig-University of Giessen, Klinikstr, Giessen, Germany. chr_kuhlmann@web.de
Schaefer
C A
CA
Fehsecke
A
A
Most
A K
AK
Tillmanns
H
H
Erdogan
A
A
D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology*; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D008564:Membrane Potentials; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis; D010766:Phosphorylation; D015221:Potassium Channels / Q000502:physiology; D015398:Signal Transduction; D014471:Umbilical Veins / Q000166:cytology
D015221:Potassium Channels; D009569:Nitric Oxide; D017228:Hepatocyte Growth Factor; D020928:Mitogen-Activated Protein Kinases