409 Corneoptosis, functional keratinocyte cell death, is tightly associated with spaciotemporal dynamics of epidermal tight junctions

Abstract

Epidermis has two physical barrier components, stratum corneum (SC) and tight junction (TJ). Formation of SC or cornification is initiated by the cell death of the top layer (SG1) of stratum granulosum, termed ‘corneoptosis’ after prolonged intracellular Ca2+([Ca2+]i) elevation (∼60min), followed by rapid intracellular acidification. However, the exact timing of the onset of corneoptorsis remains to be clarified. In this study, we aimed to determine the spatiotemporal relationship between the turnover of TJs and corneoptosis. We performed intravital confocal microscopic imaging of mouse ear co-expressing ZO-1-Venus (TJ strand marker) and GCaMP3 (intracellular Ca2+ marker). Hoechst 33342 was intradermally injected to monitor the change of nuclear pH. Consistent with our previous observation, ZO-1-Venus signals showed two TJ strand-pattens in upper SG. According to the TJ turnover, three phases are observed; TJ-phase 1 which shows single old TJ strand formed in SG2 cells; TJ-phase 2 which shows double TJ strands; TJ-phase 3 which shows new single TJ strand. Time lapse imaging analysis revealed that the elevation of GCaMP3 signal was observed only in TJ-phase 2 which shows double TJs, but not in other TJ-phases. The elevation of [Ca2+]i lasted for approximately 60min. Subsequently, intracellular acidification was induced in these SG1 cells as detected by both the decrease of GCaMP3 signal and the increase of Hoechst 33342 signals. Simultaneously, ‘upper larger TJ strand’ of double-TJ strand in SG1 cells disappeared, resulting in change from TJ-phase 2 to TJ-phase 3. These findings indicated that the onset of corneoptosis is tightly associated with the spaciotemporal dynamics of TJs.