Modulation of monocyte adherence to endothelial cells by endothelin-1 involvement of Src (p60src) and JAK1-like kinases.
OBJECTIVE
The purpose of this study was to determine the transmembrane signaling pathway by which endothelin-1 (ET-1) enhances monocyte adherence to human umbilical vein endothelial cells (HUVECs) and to investigate the role of tyrosine kinases in this mechanism.


METHODS
Adherence of purified human blood monocytes to HUVEC monolayers was assessed with radiolabeled monocytes. Tyrosine kinase activation was examined by immunoprecipitation and Western blotting.


RESULTS
ET-1 potentiated monocyte adherence to HUVECs in a biphasic manner with peaks at 10(-10) mol/L and 10(-7) mol/L. A potent antagonist to ET B receptors, when used alone, had no effect. However, the antagonist, when combined with ET-1, significantly enhanced monocyte adherence to HUVECs. Incubation of ET-1 (10(-12) mol/L to 10(-7) mol/L) with HUVECs activated tyrosine kinases in a biphasic manner as identified by immunoblotting with PY20 antibody to tyrosine phosphorylated proteins. Phosphorylated proteins with Mr 60, 110, and 130 kDa were observed after ET-1 stimulation of HUVECs. Of interest, ET A or ET B receptor antagonists failed to antagonize the effect of ET-1. Rather, these receptor antagonists significantly augmented ET-1 induced tyrosine phosphorylation in HUVECs. Immunoprecipitation with antibodies to p60SRC and JAK1 kinases followed by immunoblotting with PY20 antibody suggested that ET-1 receptor response coupling in HUVECs involves the activation of p60SRC and JAK1-like kinases.


CONCLUSIONS
These data suggest an association between activation of p60SRC and JAK1-like kinases and monocyte adherence in response to ET-1. ET-1-induced monocyte adherence is upregulated by ET B receptor antagonist, suggesting a negative feedback on cell adhesion through this receptor.
Chisholm
L J
LJ
Creighton University School of Medicine, Department of Surgery, Division of Vascular Surgery, Omaha, NE, USA.
Dovgan
P S
PS
Agrawal
D K
DK
McGregor
P E
PE
Edwards
J D
JD
D015153:Blotting, Western; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D065128:Endothelin Receptor Antagonists; D016232:Endothelins / Q000008:administration & dosage / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D009000:Monocytes / Q000187:drug effects* / Q000502:physiology; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects; D014471:Umbilical Veins; D015854:Up-Regulation; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*
D065128:Endothelin Receptor Antagonists; D016232:Endothelins; D004791:Enzyme Inhibitors; D011505:Protein-Tyrosine Kinases; D019061:src-Family Kinases