The fatty acids are shown to be critical in the maintenance of the water permeability barrier that is ascribed to the lipids in the intracellular milieu of the cornified cell layer in the epidermis. In view of this importance in the skin, we examined the phenotype of epidermal fatty acid binding protein (E-FABP)-deficient mice. In spite of total lack of E-FABP expression in the various tissues of E-FABP deficient mice, these animals appeared normal in gross and histological examination. In Northern blot analysis for other FABPs, the gene expression of heart (H-)-type FABP is specifically elevated in the liver of neonatal heterozygous and homozygous mice, suggesting the functional compensation of H-FABP for E-FABP deficiency during their development. In functional analyses of the skin, the basal transepidermal water loss (TEWL) of the adult homozygous mice showed lower levels compared with the wild-type mice, and the impairment of recovery in TEWL was observed in the homozygous mice when the lipid barrier of the skin was disrupted by acetone. These results demonstrate that E-FABP is responsible for the water permeability barrier of the skin, although the molecular mechanism remains to be further elucidated.