In this study, we determined whether Langerhans cells (LC) can function as both stimulatory and accessory cells (ACC) in alloreactive and syngeneic modified CTL induction in vitro. LC exhibited potent Ia+ stimulatory function, comparable to spleen cells, in both allo-CTL induction and in TNP-modified CTL induction assays. In contrast to spleen cells, however, when LC were syngeneic to the responders in an Ia+-adherent cell-depleted CTL culture, they could not restore the CTL responses. This was due, in part, to the suppression of CTL induction, presumably via secretion of prostaglandin E by epidermal cells (EC), because addition of indomethacin reversed the suppression. Even in the presence of indomethacin, however, LC were unable to exhibit ACC function. Finally, we tested the possibility that both Ia+ and Ia- cells were required to fulfill ACC functions. However, when spleen cells treated with anti-Ia antibody and complement and EC and indomethacin were added to an Ia+-adherent cell-depleted CTL culture, there was no significant restoration of the CTL response. Thus, LC can clearly act as Ia+ stimulatory cells in CTL induction but cannot serve as ACC in CTL induction, whereas Ia+ spleen cells may subserve both functions. These data suggest a functional heterogeneity of Ia+ antigen-presenting cells in spleen and in epidermis.