Establishment of a functionally active collagen-binding vascular endothelial growth factor fusion protein in situ.
OBJECTIVE
Tissue regeneration requires both growth factor and extracellular matrix such as collagen, serving as a scaffold for cell growth. We established FNCBD-VEGF121, consisting of the fibronectin collagen-binding domain (FNCBD) and vascular endothelial growth factor (VEGF) 121, and investigated its properties.


RESULTS
FNCBD-VEGF121 specifically bound to gelatin and type I, II, III, IV, and V collagen. This collagen-bound FNCBD-VEGF121 captured soluble VEGF receptor 2 (VEGFR-2)/Fc chimeric protein. Cell growth-promoting activity of FNCBD-VEGF121 was almost identical to that of VEGF121. The VEGF fusion protein significantly enhanced the expression of VEGFR-2 (71.6+/-0.8%) on endothelial progenitor cells (EPCs) derived from umbilical cord blood. Expectably, the collagen-bound VEGF fusion protein not only promoted the growth of endothelial cells (ECs) but also induced the expression of VEGFR-2 (63.7+/-0.8%) on non-adherent cells expanded from bone marrow CD34+ cells. Moreover, the VEGF fusion protein enhanced sprout formation of ECs in a matrigel model. In vivo experiments revealed that FNCBD-VEGF121 had local effects but not systemic effect on EPC mobilization.


CONCLUSIONS
These results suggest that FNCBD-VEGF121 stably maintains an optimally high and local concentration of VEGF with collagen matrix and stimulates both ECs and EPCs in situ, supplying a vascular regeneration niche.
Ishikawa
Tetsuya
T
Department of Regenerative Medicine, Tokai University School of Medicine, Bohseidai, Isehara, Kanagawa 259-1193 Japan. tecchan@is.icc.u-tokai.ac.jp
Eguchi
Masamichi
M
Wada
Mika
M
Iwami
Yo
Y
Tono
Kayoko
K
Iwaguro
Hideki
H
Masuda
Haruchika
H
Tamaki
Tetsuro
T
Asahara
Takayuki
T
D018952:Antigens, CD34 / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D003238:Connective Tissue / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology; D005353:Fibronectins / Q000235:genetics*; D005780:Gelatin / Q000378:metabolism; D006801:Humans; D009000:Monocytes / Q000166:cytology; D017434:Protein Structure, Tertiary / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism* / Q000494:pharmacology; D013234:Stem Cells / Q000166:cytology / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
D018952:Antigens, CD34; D005353:Fibronectins; D011993:Recombinant Fusion Proteins; C467484:VEGFA protein, human; C513782:VEGFR-2-Fc chimeric protein; D042461:Vascular Endothelial Growth Factor A; D005780:Gelatin; D003094:Collagen; D040301:Vascular Endothelial Growth Factor Receptor-2