Elucidating melatonin-regulated mechanisms of melanogenesis in human full-thickness skin and human melanocytes exposed to ultraviolet radiation

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Clinical disorders related to skin pigmentation include hypo- or hyperpigmentation, therefore active and safe pigment modulatory agents are strongly desired. Based on the so far described high biological activity of melatonin, it can be a potent substance that effectively controls mechanisms of melanogenesis. Melatonin (N-acetyl-5-methoxytryptamine) is a secretory product of the vertebrate pineal gland modulating various functions including scavenging free radicals, DNA repair or anti-apoptotic activity. It is also implicated in skin functions such as epidermal proliferation/differentiation, hair growth and melanoma/squamous cell carcinoma control. In this research application, it is proposed to evaluate the potential impact of melatonin on regulatory mechanisms of melanogenesis. So far wide-range experience gained by the applicant in the field of melatonin allows him to conduct all experiments enclosed in this proposal. The planed investigations will be carried out in human ex vivo full-thickness skin (PART I) with additional PART II using normal human melanocytes (NHM) in pure culture in comparison to co-culture with normal human keratinocytes (NHEK) that will reflect skin physiology in vivo. Additionally, such approach may describe the keratinocyte-melanocyte interaction as well as the key role of keratinocyte-dependent regulation of melanogenesis. In skin, the role and action of melatonin will be evaluated with respect to UVR-mediated melanogenesis including melatonins impact on tyrosinase (TYR), one of the key enzymes in melanogenesis, controlling the expression of microphtalmia-associated transcription factor (MITF) as well as the effect of melatonin on melanin synthesis and cAMP-PKA-MITF axis. Furthermore, a set of analysis will be performed with the earlier mentioned cell lines, i.e. NHM, NHEK, and this part together with data obtained from the first one is supposed to provide a comprehensive knowledge over the molecular way of action of melatonin-regulated melanogenesis. In both, experimental models, silencing of melanogenesis-dependent genes will be performed in order to investigate whether melatonin may maintain this process in case of alterations within these genes. Resulting findings undoubtedly will put in a brand new insight into the role of melatonin in cutaneous melanogenesis under UVR conditions and provide new principles for treatment of pigmentary disorders.