Enhanced Keratinocyte Proliferation and Migration in Co-culture with Fibroblasts

Wound healing is primarily controlled by the proliferation and migration of keratinocytes and fibroblasts as well as the complex interactions between these two cell types. To investigate the interactions between keratinocytes and fibroblasts and the effects of direct cell-to-cell contact on the proliferation and migration of keratinocytes, keratinocytes and fibroblasts were stained with different fluorescence dyes and co-cultured with or without transwells. During the early stage (first 5 days) of the culture, the keratinocytes in contact with fibroblasts proliferated significantly faster than those not in contact with fibroblasts, but in the late stage (11th to 15th day), keratinocyte growth slowed down in all cultures unless EGF was added. In addition, keratinocyte migration was enhanced in co-cultures with fibroblasts in direct contact, but not in the transwells. Furthermore, the effects of the fibroblasts on keratinocyte migration and growth at early culture stage correlated with heparin-binding EGF-like growth factor (HB-EGF), IL-1 alpha and TGF-beta 1 levels in the cultures where the cells were grown in direct contact. These effects were inhibited by anti-HB-EGF, anti-IL-1 alpha and anti-TGF-beta 1 antibodies and anti-HB-EGF showed the greatest inhibition. Co-culture of keratinocytes and IL-1 alpha and TGF-beta 1 siRNA-transfected fibroblasts exhibited a significant reduction in HB-EGF production and keratinocyte proliferation. These results suggest that contact with fibroblasts stimulates the migration and proliferation of keratinocytes during wound healing, and that HB-EGF plays a central role in this process and can be up-regulated by IL-1 alpha and TGF-beta 1, which also regulate keratinocyte proliferation differently during the early and late stage.