Characterization of an aFGF gene expression vector with therapeutic potential.
BACKGROUND
Topical application of growth factors to wounds has proven to be suboptimal in achieving epithelial growth and accelerating healing. We propose transfection of fibroblasts with a gene for acidic fibroblast growth factor (aFGF) which will allow continuous, local delivery of the growth factor to wounds, ulcerative lesions, or healing tissues.


METHODS
We utilized a pMEXneo vector containing the human aFGF gene with a secretory signal sequence from the hst/KS3 gene to obtain continuous secretion of therapeutic doses of aFGF. NIH 3T3 fibroblasts were transfected using a liposomal transfection reagent and grown in selective media.


RESULTS
Dot blot hybridization with labeled complementary DNA probes revealed the presence of plasmid DNA in transfected but not wild type fibroblasts. Intracellular concentrations of aFGF remained low in transfected cells; however, the media contained high levels (32 +/- 7 nM) of aFGF as measured by ELISA. Concentrations of aFGF capable of stimulating cell proliferation were maintained for several weeks.


CONCLUSIONS
The aFGF cDNA was transcribed and translated into a functional polypeptide that is secreted from NIH 3T3 cells at physiologically significant concentrations. Stable transfection with a eukaryotic vector which induces secretion of aFGF at levels promoting cell growth holds promise for clinical application in wounds or healing tissue. Transfection could be achieved by topical or endoscopic injection of this type of vector.
Tchorzewski
M T
MT
Section of Surgical Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland, 21224, USA.
Duncan
M D
MD
Nass
P
P
Qureshi
F G
FG
Gearhart
P J
PJ
Winchurch
R
R
Harmon
J W
JW
D016475:3T3 Cells / Q000166:cytology / Q000378:metabolism; D000818:Animals; D018076:DNA, Complementary; D016220:Fibroblast Growth Factor 1 / Q000235:genetics*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015151:Immunoblotting; D051379:Mice; D010957:Plasmids; D014162:Transfection / Q000379:methods; D014945:Wound Healing / Q000502:physiology*
D018076:DNA, Complementary; D016220:Fibroblast Growth Factor 1