Migration of a human keratinocyte cell line (HACAT) to interstitial collagen type I is mediated by the alpha 2 beta 1-integrin receptor

Abstract

The migratory response of the human keratinocyte cell line HaCaT to collagen type I and the molecular mechanism underlying collagen-mediated migration have been analyzed. The migratory response of HaCaT cells to collagen type I consisted of a dose-dependent migration to insoluble step gradients of substratum-bound collagen (haptotaxis) and to gradients of soluble collagen (chemotaxis). Checkerboard analysis demonstrated a minor chemokinetic component. Denatured collagen type I was less chemoattractive than the native triple-helical form. Pre-treatment of cells with 25-250 micrograms/ml of synthetic peptides containing the fibronectin cell-recognition sequence RGD (Arg-Gly-Asp) resulted in a concentration-dependent inhibition of fibronectin-mediated chemotaxis, whereas chemotaxis to collagen was not affected. We then investigated the role of VLA/collagen-receptors for collagen type I-induced chemotaxis. Monoclonal antibody (MoAb) 5E8, which selectively blocks function of the alpha 2 subunit of the VLA-2/collagen receptor, dose-dependently inhibited the chemotactic response of HaCaT cells to collagen. This effect was specific for collagen-mediated chemotaxis because the chemotactic response to fibronectin remained unaffected. In contrast, a function blocking MoAb directed to the alpha 3 subunit of the coexpressed VLA-3 receptor, which is also capable of binding collagen, had no effect. However, function blocking MoAb directed to the beta 1-chain of integrins completely inhibited chemotaxis to collagen type I. Based on our results, we propose that the chemotactic migration of the human keratinocyte cell line (HaCaT) to collagen type I is specifically mediated by the RGD independent VLA-2/collagen receptor (alpha 2 beta 1) of the integrin family.