The beneficial effects of dietary glucosylceramide on the barrier function of the skin have been increasingly reported, but the entire mechanism has not been clarified. By DNA microarray, we investigated changes in gene expression in hairless mouse skin when a damage-inducing AD diet and a glucosylceramide diet (GluCer) were imposed. GluCer administration potentially suppressed the upregulation of six genes and the downregulation of four genes in the AD group. Examination of the epidermal and/or dermal expression of Npr3, Cyp17a1, Col1a1, S100a9, Sprr2f, Apol7a, Tppp, and Scd3 revealed responses of various parts of the skin to the diets. In normal hairless mice, GluCer administration induced an increase in the dermal expression of Cyp17a1 and the epidermal expression of Tppp, and a decrease in the epidermal expression of S100a9. Our results provide information on gene expression not only in whole skin but also in the epidermis and dermis that should prove useful in the search for the mechanisms underlying the effects of GluCer on damaged and normal skin.