Implantation of VEGF transfected preadipocytes improves vascularization of fibrin implants on the cylinder chorioallantoic membrane (CAM) model.
The successful substitution or augmentation of soft tissues by implantation of three dimensional cell constructs, consisting of human preadipocytes and fibrin glue as a carrier matrix, requires a rapid and homogeneous vascularization of the whole implant in order to provide a sufficient blood supply of centrally situated cells. Previous investigations have shown that under in vivo conditions primary human preadipocytes induce vascularization of fibrin matrices by secretion of several growth factors, such as VEGF and bFGF. The current study investigates whether vascularization of implants can be improved by transplantation of preadipocytes following transfection with a VEGF-vector. Transfection was performed by electroporation with an pCMX-GFP and pCMX-VEGF165 vector. Transfection efficiency (GFP expression) and VEGF expression were determined in vitro by FACS analysis and VEGF immunoassay, respectively. In vivo investigations to determine the vascularization of the implants were performed on the cylinder chorioallantoic membrane (CAM). Four million VEGF transfected cells were transferred within a fibrin matrix onto the CAM on the 7(th) day of incubation and after 8 days the vascularization of the implant was histologically examined and evaluated by means of a computer-assisted image analysis program. Transfection of preadipocytes with the GFP vector by electroporation yielded transfection efficiencies between 12% and 41% of surviving cells. Results of the VEGF immunoassay demonstrated that VEGF expression was significantly higher following transfection. Investigations on the CAM outlined a significantly higher rate of vascularization in the transfected vs. control population. Our investigations demonstrate that primary human preadipocytes can be successfully transfected by electroporation with a VEGF vector. The enhanced VEGF expression on transfected cells results in an increase of vascularization of the cell constructs on the CAM.
Torio-Padron
Nestor
N
Department of Plastic and Hand Surgery, University of Freiburg Medical Center, Freiburg, Germany. torio@ch11.ukl.uni-freiburg.de
Borges
Joerg
J
Momeni
Arash
A
Mueller
Matthias C
MC
Tegtmeier
Florian T
FT
Stark
G Bjoern
GB
D017667:Adipocytes / Q000166:cytology*; D049033:Chorioallantoic Membrane / Q000098:blood supply; D018274:Electroporation; D015718:Fibrin Tissue Adhesive / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D018919:Neovascularization, Physiologic; D023822:Tissue Engineering / Q000379:methods*; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
D015718:Fibrin Tissue Adhesive; D042461:Vascular Endothelial Growth Factor A