P03. Ex vivo live imaging at high resolution to directly visualize melanin transfer from melanocytes to keratinocytes

Abstract

Melanin pigmentation in the skin is physiologically important to protect our body from UV irradiation. Melanocytes are known as melanin-producing cells in which melanin is synthesized in melanosomes. In the tanning processes, melanocytes extend numerous dendrites that are attached to surrounding keratinocyes, and subsequently melanin granules are transferred through melanocyte dendrites to keratinocytes. Although several models for intercellar melanin-transfer have been proposed by in vitro experiments and electron microscopic observations, the mechanisms of melanin-transfer in vivo remain controversial. We have developed time-lapse imaging techniques that directly visualize the melanin-transfer in an ex vivo cultured skin of chicken embryo, where epidermal cells and melanocytes are expected to behave in a similar way to in vivo . In this system, embryonic melanocytes are genetically and stably labeled with a membrane-bound EGFP using transposon-mediated gene transfer technique (Sato et al., 2007. Dev. Biol.). This manipulation directly visualizes fine structures of dendrites of differentiated melanocytes. At early stages of melanin-synthesis, melanocytes actively elongate and continuously change their shape with dendrites both extending and retracting. As melanin-synthesis proceeds, these dendrites form blebs on the plasma membrane, and subsequently they release small membrane vesicles (0.5–1 μm in diameter) to the extracellular space. Importantly, these vesicles contain melanin granules, and become incorporated into adjacent keratinocytes. Thus, melanin granules are transferred from melanocytes into keratinocytes via membrane vesicles. This is the first demonstration that the process of melanin transfer is directly visualized in the developing skin.