Hypoxia induces epidermal keratinocyte matrix metalloproteinase-9 secretion via the protein kinase C pathway.
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.
O'Toole
Edel A
EA
Centre for Cutaneous Research, ICMS, Barts and the London School of Medicine and Dentistry, Queen Mary, University of London, London, United Kingdom. e.a.otoole@qmul.ac.uk
van Koningsveld
Rudolf
R
Chen
Mei
M
Woodley
David T
DT
D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007118:Immunoassay; D007231:Infant, Newborn; D007527:Isoenzymes / Q000378:metabolism; D015603:Keratinocytes / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism*; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012867:Skin / Q000166:cytology; D013347:Subcellular Fractions / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism
C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007527:Isoenzymes; D012333:RNA, Messenger; D019715:Tissue Inhibitor of Metalloproteinase-1; D011493:Protein Kinase C; D020780:Matrix Metalloproteinase 9