The functional expression of costimulatory molecules on antigen-presenting cells may be a key event in the pathogenesis of atopic dermatitis (AD). Recently, the expression of CD86 (B7-2/B70) has been demonstrated on CD1a+ epidermal dendritic cells (DC) in AD lesions by immunohistological and functional analysis. Therefore, we sought to further characterize the in situ expression of costimulatory molecules on these cells, considering the two subpopulations of (1) CD1a+++/CD11b- Langerhans cells (LC) containing Birbeck granules and (2) CD1a+/CD11b+++ inflammatory dendritic epidermal cells (IDEC), devoid of Birbeck granules, from AD and other inflammatory skin diseases. Flow cytometry, skin mixed lymphocyte reactions (SMLR) and immunohistological analysis were performed, and showed that IDEC and not LC are the relevant cells expressing the costimulatory molecules CD80 and CD86 in situ. This expression varied with the underlying diagnosis, with AD showing the highest expression of both CD80 and CD86 in situ. Furthermore, the expression of CD80, CD86 and CD36 were significantly correlated. With short-term culture, both CD80 and CD86 were further upregulated on LC and IDEC. Finally, anti-CD86 antibody reduced the stimulatory activity of epidermal DC. These results indicate that costimulatory molecules on LC and IDEC might play a role in the pathogenesis of AD.