Comparative expression of vascular endothelial growth factor family members, VEGF-B, -C and -D, by normal human keratinocytes and fibroblasts.

The vascular endothelial growth factor (VEGF) family includes the related polypeptides VEGF-B, -C and -D, which contribute to endothelial and lymphatic vessel development. The parental VEGF molecule, VEGF-A, has been widely described in the skin, but the other members of the VEGF family have not yet been reported. The aim of our study was to determine whether the two main skin cells, keratinocytes and fibroblasts, expressed VEGF-B, -C and -D in basal condition and after stimulation by either growth factors or the pro-inflammatory cytokine tumour necrosis factor-alpha (TNF-alpha). Reverse-transcription polymerase chain reaction (RT-PCR) analysis on cultured normal human keratinocytes (NHKs) and normal human fibroblasts (NHFs) allowed the detection of different levels of VEGF-B, -C and -D mRNA, in both cell types with similar RT-PCR products in the skin cells. A semi-quantitative evaluation of the VEGF family proteins by dot blot, using the different human recombinant VEGFs, showed different levels of VEGF-B, -C and -D, in NHKs and NHFs. After cell stimulation by growth factors (epidermal growth factor (EGF) and transforming growth factor-beta1 (TGF-beta1) for NHKs and NHFs, respectively), a significant up-regulation of the VEGF family member proteins was observed in NHFs but not in NHKs. Conversely, TNF-alpha did not exert a significant effect. However, we could not detect any transcriptional modification in stimulated cells, whatever the stimulation duration. The addition of cycloheximide to the cell cultures strongly inhibited the increase of VEGF proteins in TGF-beta1-stimulated NHFs. Taken together, the results underline the major role played by NHFs in the elaboration of the VEGF family proteins known to regulate wound healing, chronic inflammation and tumour angiogenesis and lymphangiogenesis.