Abstract
Hypoxia promotes keratinocyte migration on wound bed connective tissues and is a profound biological signal that transforms a basal keratinocyte, destined to differentiate, into a motile cell that is essential for re‐epithelialization. In this study, we examined the effect of hypoxia on keratinocyte‐derived collagenases associated with keratinocyte migration. Cells plated on various connective tissue matrices under normoxic and hypoxic conditions, demonstrated a two‐fold increase in the 92 kDa, type IV collagenase (MMP‐9) when examined by quantitative zymography and ELISA. Western blotting and ELISA demonstrated a two‐fold increase in tissue inhibitor of metalloproteinase (TIMP‐1), an enzyme that binds to MMP‐9 and inhibits its activity. The hypoxia‐induced increase in cell motility could be inhibited by a neutralizing antibody to MMP‐9. Northern blotting demonstrated that MMP‐9 and TIMP‐1 mRNA increased 2.5‐ to 4‐fold, 2–12 h after the cells were made hypoxic. The hypoxia‐induced changes in MMP‐9 and TIMP‐1 were inhibited by staurosporine and bisindolylmaleimide, inhibitors of protein kinase C (PKC), but not by inhibitors of tyrosine phosphorylation and the mitogen‐activated protein kinase pathway. Inhibition of PKC also inhibited hypoxia‐induced keratinocyte migration on type I collagen. These data provide evidence that hypoxia‐induced keratinocyte migration is mediated by increased cellular secretion of MMP‐9 via the PKC pathway. J. Cell. Physiol. 214:47–55, 2008. © 2007 Wiley‐Liss, Inc.