Transforming growth factor-beta1 and ultraviolet A1 radiation increase production of vascular endothelial growth factor but not endothelin-1 in human dermal fibroblasts.
BACKGROUND
Normal and dysregulated wound healing involves fibroblast activation and angiogenesis, in which polypeptide factors such as transforming growth factor (TGF)-beta, vascular endothelial growth factor (VEGF) and endothelin-1 (ET-1) play an important part. Ultraviolet (UV) A1 (365 nm) has recently received attention as a possible treatment for some dermal fibrotic disorders.


OBJECTIVE
The aim of this study was to evaluate the effects of TGF-beta1 and UVA1 radiation, as well as that of cobalt chloride, reported to mimic hypoxia both in vivo and in vitro, on the expression of VEGF and ET-1 by cultured human dermal fibroblasts.


METHODS
Levels of VEGF and ET-1 were measured by enzyme-linked immunosorbent assay and expression of neutral endopeptidase (NEP, CD10), known to degrade ET-1, was quantified by flow cytometric analysis after cell trypsinization.


RESULTS
Our results showed that the cells released minor amounts of VEGF and ET-1. Both TGF-beta1 and UVA1 strongly increased VEGF secretion in a dose- and time-dependent manner, without significantly affecting ET-1 release. Irradiation of TGF-beta1-stimulated fibroblasts resulted in a synergistic effect on increasing levels of VEGF but not ET-1 after 48 h. Cobalt chloride stimulated the secretion of VEGF by fibroblasts; the effects of TGF-beta1 and cobalt were additive. However, no significant effect of cobalt chloride on ET-1 secretion was observed, suggesting that ET-1 production in fibroblasts is not oxygen-sensitive. The expression of NEP was not modified by TGF-beta1 or UVA1 radiation. Addition of a neutralizing anti-CD10 antibody to fibroblast cultures downregulated CD10 expression at the cell surface without changing ET-1 levels in cell supernatants after 24 or 48 h. This suggests that membrane-bound NEP has minimal or no activity against secreted ET-1.


CONCLUSIONS
Taken together, these results underline the major role played by TGF-beta1 in increasing VEGF secretion by fibroblasts. This, as well as the documented effect of UVA1 on increasing VEGF production, may have implications for wound healing in vivo.
Trompezinski
S
S
INSERM U346, Clinique Dermatologique, Hôpital E.Herriot, 69437 Lyon 03, France.
Pernet
I
I
Mayoux
C
C
Schmitt
D
D
Viac
J
J
D015415:Biomarkers; D002478:Cells, Cultured; D003035:Cobalt / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D016228:Endothelial Growth Factors / Q000096:biosynthesis*; D019332:Endothelin-1 / Q000096:biosynthesis*; D004730:Endothelium, Vascular / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000528:radiation effects*; D005434:Flow Cytometry; D006801:Humans; D008222:Lymphokines / Q000096:biosynthesis*; D015260:Neprilysin / Q000378:metabolism / Q000528:radiation effects; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014466:Ultraviolet Rays; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D015415:Biomarkers; D016228:Endothelial Growth Factors; D019332:Endothelin-1; D008222:Lymphokines; D016212:Transforming Growth Factor beta; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D003035:Cobalt; D015260:Neprilysin