WOUNDING UPREGULATES VASCULAR ENDOTHELIAL GROWTH FACTOR PRODUCTION BY KERATINOCYTES OF IN VITRO RECONSTRUCTED HUMAN EPIDERMIS.

Vascular endothelial growth factor (VEGF) is produced by keratinocytes and has been implicated in the past in animal models as the major factor involved in wound healing associated angiogenesis. In the present study we investigated the effects of wounding on the VEGF production by human keratinocytes of in vitro reconstructed epidermis (EpiDerm™). For this purpose these epidermal equivalents, consisting of differential epidermis without dermal and epidermal symbionts, were wounded by a 2-mm biopsy punch. After 4, 8 and 24 h culture supernatants and epidermal lysates were analysed for VEGF by ELISA. At all three time points levels of secreted VEGF were significantly increased after wounding compared to control. Moreover, the rate of VEGF secretion was significantly increased which might be due to increased production of VEGF. Indeed, when transcriptional or translational activity was blocked by addition of actinomycin D or cycloheximide, respectively, VEGF secretion 24 h after wounding was inhibited up to 70%, indicating de novo synthesis rather than release of preformed VEGF after wounding. In addition, as compared to secreted VEGF only low levels of stored VEGF were detectable in lysates of wounded and control epidermal equivalents indicating that upon production most of VEGF is secreted. Our data demonstrate that VEGF production by human keratinocytes of epidermal equivalents is increased after wounding and that this regulation is independent of an interaction with dermal and epidermal symbionts.

Angiogenesis, EpiDerm, VEGF, Vascular endothelial growth factor, Wound healing