Thrombopoietin stimulates endothelial cell motility and neoangiogenesis by a platelet-activating factor-dependent mechanism.
In this study, we demonstrate that human umbilical cord vein-derived endothelial cells (HUVECs) expressed c-Mpl, the thrombopoietin (TPO) receptor, and that TPO activates HUVECs in vitro, as indicated by directional migration, synthesis of 1-alkyl-/1-acyl-platelet-activating factor (PAF) and interleukin-8 (IL-8), and phosphorylation of the signal transducers and activators of transcription (STAT) STAT1 and STAT5B. The observation that WEB 2170 and CV3988, 2 structurally unrelated PAF receptor antagonists, prevented the motility of HUVECs induced by TPO suggests a role of PAF as secondary mediator. Moreover, kinetic analysis of TPO-induced tyrosine phosphorylation of STAT demonstrated that STAT5B activation temporally correlated with the synthesis of PAF. PAF, in turn, induced a rapid tyrosine phosphorylation of STAT5B and PAF receptor blockade, by WEB 2170, preventing both TPO- and PAF-mediated STAT5B activation. The in vivo angiogenic effect of TPO, studied in a mouse model of Matrigel implantation, demonstrated that TPO induced a dose-dependent angiogenic response that required the presence of heparin. Moreover, the in vivo angiogenic effect of TPO was inhibited by the PAF receptor antagonist WEB 2170 but not by the anti-basic fibroblast growth factor neutralizing antibody. These results indicate that the effects of TPO are not restricted to cells of hematopoietic lineages, because TPO is able to activate endothelial cells and to induce an angiogenic response in which the recruitment of endothelial cells is mediated by the synthesis of PAF. Moreover, biochemical analysis supports the hypothesis that STAT5B may be involved in the signaling pathway leading to PAF-dependent angiogenesis.
Brizzi
M F
MF
Dipartimento di Medicina Interna, Università di Torino and Dipartimento di Scienze Cliniche e Biologiche, Torino, Italy.
Battaglia
E
E
Montrucchio
G
G
Dentelli
P
P
Del Sorbo
L
L
Garbarino
G
G
Pegoraro
L
L
Camussi
G
G
D000818:Animals; D001381:Azepines / Q000494:pharmacology; D001672:Biocompatible Materials; D001692:Biological Transport / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D003094:Collagen; D004268:DNA-Binding Proteins / Q000378:metabolism; D004338:Drug Combinations; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics; D007700:Kinetics; D007797:Laminin; D051379:Mice; D008894:Milk Proteins; D009363:Neoplasm Proteins; D018919:Neovascularization, Physiologic / Q000187:drug effects* / Q000502:physiology; D010766:Phosphorylation; D010972:Platelet Activating Factor / Q000235:genetics*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011509:Proteoglycans; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D018121:Receptors, Cytokine; D053628:Receptors, Thrombopoietin; D050794:STAT1 Transcription Factor; D050799:STAT5 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects*; D013926:Thrombopoietin / Q000494:pharmacology*; D015534:Trans-Activators / Q000378:metabolism; D014230:Triazoles / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
D001381:Azepines; D001672:Biocompatible Materials; D004268:DNA-Binding Proteins; D004338:Drug Combinations; D016209:Interleukin-8; D007797:Laminin; D008894:Milk Proteins; D009363:Neoplasm Proteins; D010972:Platelet Activating Factor; D010975:Platelet Aggregation Inhibitors; D011509:Proteoglycans; D011518:Proto-Oncogene Proteins; D018121:Receptors, Cytokine; D053628:Receptors, Thrombopoietin; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050799:STAT5 Transcription Factor; C494095:STAT5B protein, human; C494080:Stat1 protein, mouse; C494096:Stat5b protein, mouse; D015534:Trans-Activators; D014230:Triazoles; D016222:Fibroblast Growth Factor 2; C059485:matrigel; C082218:MPL protein, human; D014443:Tyrosine; D003094:Collagen; D013926:Thrombopoietin; C063964:bepafant