In vivo detection of human vascular endothelial growth factor promoter activity in transgenic mouse skin.
We have generated transgenic mice expressing green fluorescent protein (GFP) driven by 2.453-kb (-2,362 to +91) of the 5'-upstream region of the human vascular endothelial growth factor (VEGF) promoter to monitor changes of VEGF gene transcription in situ. Neonatal transgenic mice exhibited GFP-derived fluorescence in tissues that have been previously reported to express VEGF mRNA expression, including lung, cartilage, and brain. In normal skin during postnatal development, moderate fluorescence was observed in the upper epidermis and, more prominently, in the outer root sheath keratinocytes of hair follicles. Strong up-regulation of GFP fluorescence was observed in the hyperplastic epidermis of the wound edge at 48 hours after wounding, whereas little GFP fluorescence was detected in the dermis. In situ hybridization confirmed an identical expression pattern of VEGF mRNA in these wounds. Topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA) induced strong VEGF-GFP expression in suprabasal epidermis. Little or no fibroblast-derived fluorescence was seen both in the wound model and after TPA application. By confocal laser microscopy, increased GFP fluorescence was detectable in the epidermis of intact mouse ear skin as early as 6 hours after topical TPA treatment. Importantly, GFP fluorescence was also measurable in the skin of living transgenic mice. These results resolve the present controversy regarding the ability of VEGF-GFP transgenic mouse models to correctly reflect established patterns of VEGF expression, and show the model to be a powerful tool for the in vivo monitoring of VEGF gene expression.
Kishimoto
J
J
Department of Dermatology, the Cutaneous Biology Research Center, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129, USA. jiro.kishimoto@cbrc2.mgh.harvard.edu
Ehama
R
R
Ge
Y
Y
Kobayashi
T
T
Nishiyama
T
T
Detmar
M
M
Burgeson
R E
RE
D000818:Animals; D002478:Cells, Cultured; D020405:Dermis / Q000166:cytology / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000235:genetics*; D000078404:Epidermal Cells; D004817:Epidermis / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015870:Gene Expression; D005786:Gene Expression Regulation / Q000187:drug effects; D049452:Green Fluorescent Proteins; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D008164:Luminescent Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D008222:Lymphokines / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D012867:Skin / Q000166:cytology / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D014945:Wound Healing
D016228:Endothelial Growth Factors; D008164:Luminescent Proteins; D008222:Lymphokines; D011993:Recombinant Fusion Proteins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D049452:Green Fluorescent Proteins; D013755:Tetradecanoylphorbol Acetate