Insulin stimulation of keratinocyte migration is time- and dose-dependent. Cells were assayed for migration using the cloning ring migration assay. (A, B) Cells were treated with 10-6 M insulin at multiple time points, as indicated. Insulin significantly increased keratinocyte migration. (C) Cells were then treated with 10-8 M, 10-7 M, 10-6 M or 10-5 M insulin, and migration distance measured 24 and 48 h after insulin treatment. The effect of insulin on keratinocyte migration was dose-dependent. (D) Cells were pre-treated with 5 μg/ml mitomycin C for 3 h followed by 10-7 M insulin, and migration distance measured 24 and 48 h after insulin treatment. The effect of insulin on keratinocyte migration was not abolished by mitomycin C. Each treatment group was performed in triplicate. Data are shown as the mean value +/- SD. Statistics are shown as comparisons between the treatment and control. *P < 0.05, **P < 0.01. ***P < 0.001.