Staurosporine promotes endothelial cell assembly and FAK phosphorylation during in vitro angiogenesis.
In the present study, we report that staurosporine, a known PKC inhibitor, enhanced in vitro angiogenesis. Endothelial cells plated in a three-dimensional matrix formed cords and enclosed structures within 4-6 hours. The cells in cord structures became elongated during the subsequent incubation. Tube formation was confirmed by confocal microscopy. Addition of VEGF enhanced the early responses of endothelial cells, leading to enhanced formation of cords. Staurosporine unexpectedly also enhanced the early endothelial responses, leading to faster alignment of cells and assembly into tube-like structures. At concentrations inhibitory to endothelial cell PKC activity, staurosporine produced 91% and 203% increases in the number of cords and the enclosed structures, respectively, as compared to the controls. Other selective inhibitors of PKC did not stimulate in vitro angiogenesis in the absence or presence of VEGF. Further investigation showed that inhibition of PI-3 kinase and Raf-1 significantly reduced the effects of staurosporine. Staurosporine-induced in vitro angiogenesis required integrins alpha2 and alphavbeta3 and was associated with significantly enhanced FAK phosphorylation. These data indicate that staurosporine enhances in vitro angiogenesis by a means unrelated to its PKC inhibition. The data suggest that enhancement of in vitro angiogenesis by staurosporine involves integrin-mediated signaling, including the stimulation of FAK phosphorylation.
Tarzami
Sima T
ST
Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285, USA.
Hsieh
Susie Shao-Hsu
SS
Esterman
Michail A
MA
Singh
Jai Pal
JP
D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D051418:Focal Adhesion Kinase 2; D006801:Humans; D039421:Integrin alpha2 / Q000502:physiology; D039302:Integrin alphaVbeta3 / Q000502:physiology; D018613:Microscopy, Confocal; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf / Q000037:antagonists & inhibitors / Q000502:physiology; D019311:Staurosporine / Q000494:pharmacology*; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D051418:Focal Adhesion Kinase 2; D039421:Integrin alpha2; D039302:Integrin alphaVbeta3; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C; D011505:Protein-Tyrosine Kinases; D019908:Proto-Oncogene Proteins c-raf; D019311:Staurosporine; D042461:Vascular Endothelial Growth Factor A