Induction of a freeze lesion in human skin is an experimental model of hyperalgesia that allows assessing the antihyperalgesic effects of traditional non-steroidal anti-inflammatory drugs (NSAIDs). We have investigated whether this model is also sensitive to selective cyclooxygenase (COX)-2 inhibitors and have characterized morphological substrates of the generated hyperalgesia in the skin. In eight healthy subjects, a freeze lesion was induced and mechanical pain thresholds (MPT) were tested for 5 h following administration of the non-selective COX inhibitor diclofenac (75 mg), the COX-2-selective inhibitor parecoxib (40 mg) or placebo in a randomized, double-blind cross-over study. In five additional healthy subjects, biopsies were taken from normal skin and the area of freezing injury. Induction of the freeze lesion resulted in hyperalgesia expressed by a decrease of MPT after 24 h. Diclofenac and parecoxib, but not placebo, statistically significantly elevated MPT. Histochemical and Western blot analyses of skin biopsies revealed a strong upregulation of COX-2, a slight decrease of COX-1 and activation of nuclear factor kappa B (NF-κB) in the area of the freezing injury. These findings indicate that the freeze lesion model is sensitive to NSAIDs including selective COX-2 inhibitors, and that NF-κB-dependent COX-2 upregulation contributes to the hyperalgesia in this model.