Nuclear factor E2-related factor 2 (Nrf2) is one of the most important redox-sensitive transcription factors regulating expression of antioxidative genes and cytoprotective enzymes, which constitute the cellular response to oxidative stress and xenobiotic damage. In this study, we investigated the functional role of Nrf2 during normal epidermal keratinocyte (NHEK) differentiation. Immunohistochemical staining showed that Nrf2 is expressed from basal to granular layer of epidermis. When cultured NHEKs were treated with 1.2 mM calcium, Nrf2 expression was increased gradually in protein levels and Nrf2 translocated into the nucleus in a differentiation-dependent manner. When Nrf2 was overexpressed in NHEK by adenoviral transduction, the expression of the NHEK differentiation marker loricrin and keratin 10 was increased and overexpression of Nrf2 also increased the luciferase activity of loricrin in the absence of calcium. These results suggest that Nrf2 helps to promote the differentiation of epidermal keratinocytes.