Evidence for apoptosis signal-regulating kinase 1 in the regenerating palatal epithelium upon acute injury

Apoptosis signal-regulating kinase 1 (ASK1), a recently identified mitogen-activated protein (MAP) kinase kinase kinase, is a key element in the mechanism of stress-and cytokine-induced apoptosis. However, pathophysiologic roles of ASK1 in vivo are poorly understood. In the present study, we analyzed the ASK1 expression in injured rat palate using an immunohistochemical approach to investigate the roles of ASK1 during the process of wound healing. In the normal rat palatal epithelium, a weak cytoplasmic staining of ASK1 was observed in keratinocytes of the prickle cell layer. After mucoperiosteal injury of the palate, ASK1 was clearly observed in the suprabasal keratinocytes surrounding the wound. ASK1 expression was most evident at Day 2 after injury in the edge of the migrating epithelium. Thereafter, the intensity of ASK1 staining decreased gradually until the re-epithelialization was completed at Day 10 to 14. A staining with the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end-labeling method identified a number of apoptotic keratinocytes in the suprabasal layers at the healing edge. Active induction of epithelial apoptosis was readily detectable from Day 5 after injury. In double-staining analysis, the temporal and spatial pattern of ASK1 expression correlated well with the appearance of apoptotic keratinocytes. p38 MAP kinase, a downstream component of ASK1, was found to be activated at the sites of ASK1 expression, suggesting that increased expression of ASK1 leads to activation of downstream MAP kinase signaling pathway in vivo. These results suggest a significant contribution of ASK1 to the epithelial apoptosis in the process of mucoepithelial wound repair.