Organ culture of mammalian skin and the effects of ultraviolet light and testosterone on melanocyte morphology and function

Scrotal skin of black Long-Evans rats and human thigh skin were maintained in vitro as organ cultures for as long as 14 days, and examined histologically using the combined skin splitting and Dopa techniques. Selected rat skin cultures received testosterone in the culture medium and/or were irradiated with ultraviolet light (290-320 nm UVL). With increased time in culture, scrotal melanocytes round up and there is an increase in epidermal pigmentation. Human skin behaves similarly; after eight days in vitro human melanocytes also become rounded, but remain strongly Dopa-positive. Addition of exogenous testosterone to cultured rat skin maintains dendritic morphology of melanocytes, but cell body size is still reduced. UVL irradiation stimulates melanocytes in rat skin cultures, maintaining their dendritic morphology and increasing epidermal and dermal pigmentation. Cultured skin receiving both UVL and testosterone illustrates a synergistic effect. Electron microscopic examination of cultured rat skin shows the presence of large melanosome complexes in keratinocytes, much larger than those found in vivo. Melanocytes appear to be active as they contain an extensive Golgi zone, rough endoplasmic reticulum, and melanosomes in various stages of formation. Dermis contained many dermal melanocytes and macrophages laden with melanosomes, correlating with the increased visible dermal pigmentation in vitro. This UVL stimulation of melanocytes in our skin organ cultures contrasts with the lack of melanogenic stimulation found in melanoma cell cultures. Our findings suggest that the intact epidermal melanin unit may be necessary for UVL stimulation of melanocytes.