title: Quantitative single-cell motility analysis of platelet-rich plasma-treated endothelial cells in vitro.
abstract: Platelet-rich plasma (PRP) has been widely applied in regenerative therapy due to its high concentration of growth factors. Previous in vitro and in vivo studies have provided evidence supporting the angiogenic activity of PRP. To more directly demonstrate how PRP acts on endothelial cells, we examined the PRP-induced changes in the motility of human umbilical vein endothelial cells by examining the involvement of VEGF. Time-lapse quantitative imaging demonstrated that in the initial phase (∼2 h) of treatment, PRP substantially stimulated cell migration in a wound-healing assay. However, this effect of PRP was not sustained at significant levels beyond the initial phase. The average net distance of cell migration at 10 h was 0.45 ± 0.16 mm and 0.82 ± 0.23 mm in control and PRP-stimulated cells, respectively. This effect was also demonstrated with recombinant human VEGF and was significantly attenuated by a neutralizing anti-VEGF antibody. Immunofluorescent examination of paxillin and actin fibers demonstrated that PRP concomitantly up-regulated focal adhesion and cytoskeletal formation. Western blotting analysis of phosphorylated VEGFR2 demonstrated that PRP mainly stimulated the phosphorylation of immature VEGFR2 in a dose- and time-dependent manner, an action that was completely blocked by the neutralizing antibody. Taken together, these data suggest that PRP acts directly on endothelial cells via the activation of VEGFR2 to transiently up-regulate their motility. Thus, the possibility that PRP desensitizes target endothelial cells for a relatively long period of time after short-term activation should be considered when the controlled release system of PRP components is designed.
authors.affiliation: Division of Oral Bioengineering, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.|Advanced Research Center, the Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Kawase
authors.affiliation: Department of Materials Science and Technology, Niigata University, Niigata, Japan.
authors.forename: Takaaki
authors.initials: T
authors.lastname: Tanaka
authors.affiliation: Division of Periodontology, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Okuda
authors.affiliation: Advanced Research Center, the Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan.|Department of Oral and Maxillofacial Radiology, the Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Tsuchimochi
authors.affiliation: Institute of Research Collaboration and Promotion, Niigata University, Niigata, Japan.
authors.forename: Masafumi
authors.initials: M
authors.lastname: Oda
authors.affiliation: Department of Mechanical and Control Engineering, Niigata Institute of Technology, Kashiwazaki, Japan.
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Hara
mesh_terms: D000199:Actins / Q000378:metabolism; D000328:Adult; D002448:Cell Adhesion; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects; D005260:Female; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D008297:Male; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D051419:Paxillin / Q000378:metabolism; D010766:Phosphorylation; D010976:Platelet Count; D053657:Platelet-Rich Plasma / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012038:Regeneration; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism; D014945:Wound Healing
chemical_list: D000199:Actins; D051419:Paxillin; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C000588866:KDR protein, human; D040301:Vascular Endothelial Growth Factor Receptor-2
keywords: cell motility; human umbilical vein endothelial cells (HUVECs); platelet-rich plasma; time-lapse imaging; vascular endothelial growth factor; wound healing assay