Methods: Patient-matched OMF and SF were cultured until senescence. Population doubling (PD) levels were determined throughout and cellular phenotype investigated. Senescence was determined using senescence associated-â galactosidase, cellular morphology and analysis of PDs. At low PD level, fibroblasts were synchronised through serum starvation for 48 hours, re-stimulated with serum and RNA extracted at 0, 1, 6 and 24 hours for transcriptional profile analysis using Affymetrix Microarrays.
Results: Microarray data analysis demonstrated altered expression of 50 genes (greater than 3-fold up or down regulated) between the OF and SF. Cluster analysis and annotation identified numerous pathways and proteins which will form the basis of future investigations. From the PD data it was notable that OMF proliferated more quickly than SF, progressed more rapidly through the cell-cycle (Flow cytometry: 31% of OMF vs. 7% of SF in G2 phase) and underwent many more PDs (>90 vs. 45 in SF); as such SF senesced earlier. Furthermore, compared to SF, OMF demonstrated an increased ability to re-organise their extracellular matrix environment (fibroblast-populated collagen lattices), migrated faster and were better able to repopulate monolayer wounds.
Conclusions: These results demonstrate that distinct genotypic differences exist between OMF and SF which are reflected by distinct differences in their cellular phenotypes and ageing profiles. Ongoing studies are aimed at corroborating this link between the genotypes and phenotypes and delineating how these differences relate to the ability of the cells in these tissues to give rise to differential healing.