Hypoxia increases reepithelialization via an alphavbeta6-dependent pathway

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Reepithelialization is an essential step in successful cutaneous wound healing. Human keratinocytes, integral in this process, have been shown to have increased motility in the hypoxic healing edge of wounds correlating with the clinical success of semiocclusive hypoxic dressings, although the underlying mechanisms remain poorly understood. Subconfluent human keratinocyte cell monolayers were exposed to 1% hypoxia for up to 24 hours or control conditions. Re-oxygenation studies were performed up to 72 hours. Cellular alphav subunit and alphavbeta6 integrin expression was measured by flow cytometry. Migration scratch assays on fibronectin following hypoxic exposure were performed over 24 hours. Relative matrix metallo-proteinase (MMP)-2, 9 activity was determined by gelatin zymography with TIMP-1 levels assayed by enzyme-linked immunoassay. Sustained increases in alphav and alphavbeta6 expression were shown up to 48 hours in re-oxygenation studies (P < 0.001). Standardized scratch assays confirmed increased migration in the hypoxic group (P < 0.05). This effect was attenuated by the addition of a specific inhibitor of the alphavbeta6 integrin. MMP-2 and -9 activity was up-regulated following hypoxic exposure (P < 0.001; P < 0.05, respectively), whereas increased MMP expression was significantly retarded by addition of an alphavbeta6 inhibitor (P < 0.05). Migration on fibronectin was attenuated by a specific gelatinase inhibitor. We conclude that integrin alphavbeta6-dependent MMP-2 and -9 up-regulation is an important feature of increased migration in hypoxic human keratinocytes.