Revisiting the role of melatonin in human melanocyte physiology: A skin context perspective

Abstract

The evolutionarily ancient methoxyindoleamine, melatonin, has long perplexed investigators by its versatility of functions and mechanisms of action, which include the regulation of vertebrate pigmentation. Although first discovered through its potent skin-lightening effects in amphibians, melatonin's role in human skin and hair follicle pigmentation and its impact on melanocyte physiology remain unclear. Synthesizing our limited current understanding of this role, we specifically examine its impact on melanogenesis, oxidative biology, mitochondrial function, melanocyte senescence, and pigmentation-related clock gene activity, with emphasis on human skin, yet without ignoring instructive pointers from nonhuman species. Given the strict dependence of melanocyte functions on the epithelial microenvironment, we underscore that melanocyte responses to melatonin are best interrogated in a physiological tissue context. Current evidence suggests that melatonin and some of its metabolites inhibit both, melanogenesis (via reducing tyrosinase activity) and melanocyte proliferation by stimulating melatonin membrane receptors (MT1, MT2). We discuss whether putative melanogenesis-inhibitory effects of melatonin may occur via activation of Nrf2-mediated PI3K/AKT signaling, estrogen receptor-mediated and/or melanocortin-1 receptor- and cAMP-dependent signaling, and/or via melatonin-regulated changes in peripheral clock genes that regulate human melanogenesis, namely Bmal1 and Per1. Melatonin and its metabolites also accumulate in melanocytes where they exert net cyto- and senescence-protective as well as antioxidative effects by operating as free radical scavengers, stimulating the synthesis and activity of ROS scavenging enzymes and other antioxidants, promoting DNA repair, and enhancing mitochondrial function. We argue that it is clinically and biologically important to definitively clarify whether melanocyte cell culture-based observations translate into melatonin-induced pigmentary changes in a physiological tissue context, that is, in human epidermis and hair follicles ex vivo, and are confirmed by clinical trial results. After defining major open questions in this field, we close by suggesting how to begin answering them in clinically relevant, currently available preclinical in situ research models.

Keywords: hair follicle; human skin; melanocyte; melatonin; pigmentation.

Figure 1.. Schematic summary of melatonin’s effects in human epidermal and HF melanocytes.

Exogenous or endogenously synthesized melatonin can regulate phenotype in these cells through interactions with membrane-bound MT1/2 receptors NQO2, and the calcium/calmodulin complex or through stimulation of Nrf2 (reviewed in). However, it is not fully understood if melatonin activates NQO2, a detoxifying enzyme^,, Noteworthy phenotypic effects of melatonin include melanogenesis inhibition, and stimulation of DNA repair, and expression and activity of antioxidant enzymes (e.g., superoxide dismutase and catalase) (reviewed in). Melatonin may also be transported to different subcellular compartments, but the detailed mechanism is not fully understood. Furthermore, melatonin can be synthesized within these melanocytes. Melatonin and its metabolites, such as cyclic-3-hydroxymelatonin (C-3HOM) and N-acetyl-5-methoxyknuramine (AMK), directly scavenge ROS/RNS^, and help maintain mitochondrial homeostasis through interactions with cytochrome C and enzymes of the electron transport chain. Specifically, cytochrome C within mitochondria is thought to be involved in the conversion of melatonin to its potent antioxidant metabolite, N(1)-acetyl-N(2)-formyl-5-methoxykynuramine (AFMK), and its secondary product, AMK, when in the presence of hydrogen peroxide. Also, melatonin may interact with cytochrome C and electron transport chain (ETC) enzymes within mitochondria to promote mitochondrial homeostasis and decrease free radical formation. Furthermore, melatonin may affect the transcription of peripheral clock genes Bmal1 and Per1 with alterations in melanogenesis and other melanocyte activities^,,. Direct effects are shown by solid lines and multiple reactions and signaling are shown by dashed lines. Melatonin receptors 1 and 2 (MT1/2); hair follicle (HF); reactive oxygen species (ROS); reactive nitrogen species (RNS); nuclear factor erythroid 2-related factor 2 (Nrf2); N-Ribosyldihydronicotinamide:Quinone Reductase 2 (NQO2).

Figure 2.. Schematic summary describing PI3K/AKT pathway modulation and its effects on melanogenesis.

In normal human melanocytes, melatonin stimulates Nrf2, which can activate the PI3K/AKT pathway to phosphorylate (i.e., inactivate) GSK-3. Without GSK-3, MITF remains unphosphorylated (i.e., inactive), leading to decreased transcription of tyrosinase, TRP-1, and TRP-2, thereby decreasing melanogenesis.

Figure 3.. Schematic summary describing antioxidant defense mechanisms by melatonin and its metabolites in human melanocytes.

Melatonin can bind to MT1 and MT2 receptors on the cell membrane, triggering a signaling cascade that leads to expression of antioxidant enzymes (e.g., SOD, GPx, GR, and CAT) for defense against ROS and RNS^,. Melatonin may also be transported to the cytoplasm, but the detailed mechanism is not fully understood. Furthermore, melatonin can be synthesized within these melanocytes. Melatonin and its metabolites, such as C-3HOM and AMK, can directly scavenge ROS/RNS^,. Furthermore, melatonin and its metabolites, including AFMK, 6-OHM, 5-MT, and NAS, protect human epidermal melanocytes from UV-B-induced damage/apoptosis by enhancing phosphorylation of p53 at Serine 15, thereby leading to activated p53 accumulation in the nucleus and stimulation of DNA repair. Melatonin may activate NQO2, thereby reducing oxidative stress^,, however this mechanism’s presence in these melanocytes is not fully understood. Melatonin at concentrations higher than 1 nM within the cell can interact with the calcium/calmodulin complex leading to inhibition of NOS1-mediated generation of RNS, with potential reductions in RNS levels. Melatonin may also inhibit the Keap1-E3 ligase complex and the ubiquitination and proteasomal degradation of Nrf2, thereby preserving high Nrf2 levels that translocate to the nucleus. In the nucleus, Nrf2 may couple with Maf, a transcription factor, allowing Nrf2 to bind ARE on the promoter region of genes encoding antioxidant enzymes (e.g., SOD and GPx), resulting in their increased expression and activity, which then convert ROS and RNS to unreactive products. Direct effects are shown by solid lines and multiple reactions and signaling are shown by dashed lines. Reactive oxygen species (ROS); reactive nitrogen species (RNS); superoxide dismutase (SOD); glutathione peroxidase (GPx); glutathione reductase (GR); catalase (CAT).

Figure 4.. Schematic summary describing hypothesized mechanisms by which melatonin may regulate melanogenesis in normal human epidermal and HF melanocytes.

Melatonin may activate the PI3K/AKT pathway, via Nrf2 activation, to stimulate expression of Bmal1, thereby increasing BMAL1 levels. BMAL1 may increase expression of Nrf2 to further stimulate this PI3K/AKT pathway and PER1 to inhibit MITF downstream. PER1 also translocates to the nucleus and inhibits transcriptional activity of BMAL1^,, thereby preventing BMAL1’s stimulation of MITF transcription. Decreased MITF levels lead to decreased expression of melanogenesis enzymes tyrosinase, TRP-1, and TRP-2 and results in decreased melanogenesis.