Identifying the molecular and genetic basis for non-healing wounds, scarless healing and scarring: study of human fibroblasts

Aims: In contrast to dermal wounds, oral mucosal wounds heal rapidly with minimal scarring and seldom develop fibrosis or contractures. We have investigated, for the first time, differences in cellular phenotype and onset of senescence in oral mucosal fibroblasts (OMF) and dermal fibroblasts (DF), and identified their temporal gene expression profiles in response to serum exposure (simulating wounding response). Methods: Patient-matched OMF and DF were cultured until senescence. Population doublings levels (PDLs) were determined throughout and cellular phenotype investigated. Senescence was determined using SAbeta- galactosidase, cellular morphology and analysing population doublings. At low PDLs, fibroblasts were synchronised through serum starvation for 48 hours, restimulated with serum and RNA extracted at 0, 1, 6 and 24 hours for transcriptional profile analysis using microarray (Affymetrix). Results:OMFdemonstrated increased ability to reorganise extracellularmatrix (fibroblast-populated collagen lattices); OMF proliferated quickly, underwent more PDLs (>90 vs. 55 in DF), and progressed rapidly through the cell cycle (flow cytometry); OMF manifested better ability to migrate and repopulate monolayer wounds. DF senesced earlier (PDLs: 40–55 vs. 75–95 in OMF), with their ability to repopulate wounds and extracellular matrix reorganisation decreasing further. Initial microarray data analyses demonstrated altered expression of numerous genes, with known wound healing roles (e.g. IGF-2, superoxide dismutase-3, HGF, homeobox), between the two sites. Conclusions:Wehave identified phenotypic and genotypic differences in OMF and DF. These candidate genes provide the basis for identifying molecular pathways and alterations in protein expressions to elucidate the differential healing (scarless, scarring, contractures, nonhealing) observed in these tissues