Integrin receptors and RGD sequences in human keratinocyte migration: unique anti-migratory function of alpha 3 beta...

The migration of keratinocytes over the wound bed plays an important role in the re-epithelialization of cutaneous wounds. However, the mechanisms by which keratinocytes migrate over extracellular matrix components are unknown. In this study, we sought to determine if the RGD sequences in matrix molecules and recognition of these sequences by keratinocytes played a role in the locomotion of keratinocytes. After allowing the cells to attach to the matrix, RGD-containing peptides or control peptides were added to a keratinocyte migration assay. The addition of RGD-containing peptide dramatically inhibited keratinocyte locomotion on a matrix of fibronectin but not on collagen matrices. Therefore, RGD recognition is a critical step for fibronectin-mediated migration but not for collagen-mediated migration. Because the RGD sequences are recognized by cell-surface integrin receptors in a number of cell types, we next examined the roles of integrin receptors in human keratinocyte migration. Using monospecific antibodies that recognize integrin subunits, we found that blocking the beta 1 subunit inhibited the migration of keratinocytes on matrices of fibronectin, interstitial collagen, and basement membrane collagen. Blocking the alpha 5 beta 1 receptor significantly inhibited migration on fibronectin but not on collagen matrices. Conversely, blocking the alpha 2 beta 1 receptor inhibited migration on collagen matrices but not on fibronectin. Blocking the alpha 3 beta 1 receptor uniquely enhanced migration on fibronectin and collagen matrices. In contrast to cells apposed to matrices without the receptor blocked, the enhanced migration in the presence of anti-alpha 3 beta 1 antibody occurred at the later time points of the migration assay. The enhancement of migration by blocking the alpha 3 beta 1 integrin receptor suggests that the interaction of the alpha 3 beta 1 receptor with matrices is associated with immobility.