Increased induction of Ca 2+-mediated differentiation by gamma ray is mediated by endogeneous activation of the protein kinase C signaling pathways in mouse epidermal cells

Abstract

Purpose: The aim of this study was to determine whether γ-rays can affect Ca 2+-induced differentiation in normal and neoplastic mouse epidermal cells. Methods and Materials: After γ-ray irradiation, primary and v-ras Ha transformed mouse keratinocytes were cultured for 48 h in 0.12 mM Ca 2+-containing media, and cellular translocation from cytosolic to particulated fraction of each PKC isozyme and expressions of differentiation markers were examined. Results: Morphological difference was seen at 48 h after irradiation in both Ca 2+-shifted normal and v-ras Ha transformed cells; v-ras Ha cells were more resistant to the radiation than normal cells. Radiation potentiated granular cell-differentiation marker expressions (filaggrin, loricrin, and SPR-1) in both normal and v-ras Ha transformed cells. In the case of spinous cell markers, the expression of keratins K1 and K10, which are usually blocked in v-ras Ha cells was increased after irradiation. However, there was no change of K8 expression level, which can be seen only after v-ras Ha transfection. Cellular fractionation and immunoblot analysis with antibodies against PKCα, δ, ϵ, η, and ξ revealed that PKCα was responsible for the differentiation marker expression. Conclusions: These findings suggest that PKCα is an important component of the signaling pathway regulating radiation-induced differentiation in both normal and neoplastic epidermal cells.