Epidermal protein kinase C-beta(2) is highly sensitive to downregulation and is exclusively expressed in Langerhans cells: Downregulation is associated with attenuated contact hypersensitivity

Treatment of mice with multiple topical applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) or diacylglycerol resulted in a preferential decrease in epidermal protein kinase C-beta(2) (PKC-beta(2)) compared with PKC-alpha as determined by western analysis. When PKC-alpha was decreased by 40%, PKC-beta(2) could no longer be detected, suggesting that PKC-beta(2) is more sensitive to downregulation, and/or specific epidermal cell types that contain PKC-beta(2) are more sensitive to TPA/diacylglycerol. To address this issue, we isolated Langerhans cells (LCs) from epidermal cell suspensions with immunomagnetic beads and an antibody to the class II major histocompatibility complex, Northern blot analysis revealed a PKC-beta(2) signal in isolated LCs that was 40-fold greater than that observed in unfractionated epidermal cells, and no PKC-beta(2) signal was detected in epidermal cells depleted of LCs, indicating that PKC-beta(2) is expressed exclusively in LCs within the epidermis. Western blot analysis confirmed the presence of PKC-beta(2) in LCs, PKC-beta(2), was highly sensitive to downregulation, because a single application of TPA resulted in a 90% loss of PKC-beta(2) within 6 h without a decrease in the number of LCs, To determine whether the decreased level of PKC-beta(2) within LCs was associated with an alteration in contact hypersensitivity, we treated mice with only a single application of TPA, and 6 hours later mice were sensitized with 2,4-dinitrofluorobenzene on the same dorsal area. Subsequent challenge revealed a 60% decrease in contact hypersensitivity in TPA-treated mice, These data indicate that (i) within the epidermis, PKC-beta(2) is highly sensitive to downregulation and is exclusively expressed in LCs, and (ii) the downregulation of PKC-beta(2) is associated with impaired LC function with respect to contact hypersensitivity.