Epidermal Melanocytes: Regulation of Their Survival, Proliferation, and Function in Human Skin

Melanocytes are cells specialized in the synthesis of the pigment melanin, in the form of eumelanin, the brown/black, and pheomelanin, the red/yellow pigment (Ito and Wakamatsu 2003). Melanocytes reside in the cutaneous epidermis, within hair follicles, in the eye, the leptomeninges, the inner ear, and as has been demonstrated lately, in the heart (Brito and Kos 2008; Goldgeier et al. 1984; Tachibana 1999; Yajima and Larue 2008). Melanin produced by melanocytes provides the skin, hair, and eyes with their distinctive coloration. In this chapter we focus on epidermal melanocytes, since they have been the most investigated due to their importance in photoprotection against sun-induced skin cancers, and for being the precursors for cutaneous melanoma, the deadliest form of skin cancer. We hereby provide a brief summary of the properties of melanocytes, review how cutaneous pigmentation is regulated, and discuss the significance of paracrine and autocrine factors and their signaling pathways in modulating the survival, proliferation, and function of melanocytes, constitutively, and in response to solar ultraviolet radiation (UV), a major environmental stressor and etiological factor for skin cancers, including melanoma. We end by briefly describing how the knowledge gained about the regulation of melanocytes can be translated into preventative and therapeutic strategies for melanoma. 2.1 Unique Properties of Melanocytes In the human epidermis, melanocytes reside on the basement membrane, at the epidermaldermal junction (Jimbow and Fitzpatrick 1975). Melanocytes differ from keratinocytes, the main structural cells of the epidermis, in many respects. Unlike basal keratinocytes that are highly proliferative and capable of regenerating the epidermal layers, melanocytes Z.A. Abdel-Malek (*) Department of Dermatology, University of Cincinnati College of Medicine, Cincinnati, OH 45267–0592, USA e-mail: [email protected] 8 Z.A. Abdel-Malek and V.B. Swope have a low proliferation potential. Keratinocytes undergo a well-defined differentiation program that culminates in their death by an apoptosis-like process (reviewed by Eckert et al. 1997). On the other hand, most melanoblasts, the precursors for melanocytes, become fully differentiated upon reaching their final destination, the epidermis, after their migration during embryonic development from the neural crest (Bronner-Fraser 1993) (see also Chap. 13). Melanocytes are resistant to apoptosis, as they are endowed with antiapoptotic mechanisms, exemplified by constitutive expression of the antiapoptotic protein Bcl2, which enable them to survive (Plettenberg et al. 1995). Contrary to keratinocytes, melanocytes have a very long life span, and survive for decades in the epidermis (Quevedo et al. 1969). However, the longevity of melanocytes and their resistance to apoptosis is a doubleedged sword, since these properties make them vulnerable to mutations that arise over the years, particularly due to repetitive sun exposure, and might culminate in melanoma formation in high-risk individuals. These properties also explain the resistance of melanoma to various chemotherapeutic agents and to radiation. Given the significance of melanocytes in protection of the skin from UV-induced skin cancers, it is critical to maintain genomic stability of these cells to ensure their proper function and ability to maintain epidermal homeostasis. 2.2 Factors that Determine Cutaneous Pigmentation Cutaneous pigmentation is determined by the rate of synthesis of melanins (eumelanin and pheomelanin) by melanocytes, the relative eumelanin and pheomelanin contents, and the rate of transfer of melanin-containing organelles, the melanosomes, from melanocytes to keratinocytes (Pathak et al. 1980). These are the main factors that account for individual differences in skin pigmentation. Melanosomes contain enzymes that are essential for melanin synthesis, namely tyrosinase, the rate-limiting enzyme for melanin synthesis, tyrosinase-related protein (TYRP-1), and dopachrome tautomerase (DCT), also known as tyrosinase-related protein 2 (TRP-2) (reviewed by Hearing 2005). The activity of tyrosinase and the protein levels of these three melanogenic enzymes correlate directly with melanin content of melanocytes (Abdel-Malek et al. 1993; Wakamatsu et al. 2006). Melanosomes also express OA1, a G-protein coupled receptor for L-DOPA, an intermediate in the melanin synthetic pathway, and a substrate for tyrosinase, on their membrane (Hearing 2005). The number of melanocytes does not significantly differ among individuals with different pigmentary phenotypes (Szabo 1954). The difference in pigmentation lies primarily in the rate of melanin synthesis, which is determined by many genes that code for regulatory proteins, including melanogenic enzymes, growth factor receptors, and transcription factors, as well as structural proteins that make up the melanosome. The difference in constitutive pigmentation among individuals is primarily dictated by eumelanin, which correlates directly with the extent of pigmentation (Hennessy et al. 2005; Wakamatsu et al. 2006). Melanocytes interact with keratinocytes by donating fully melanized (mature) melanosomes (Pathak et al. 1980). Melanocytes are dendritic cells, and their dendrites serve as 9 2 Epidermal Melanocytes: Regulation of Their Survival, Proliferation, and Function in Human Skin conduits for the transport of melanosomes to surrounding keratinocytes. In turn, keratinocytes participate in regulating the transfer of melanosomes by expressing protease-activated receptor 2 (PAR-2), a G-protein coupled receptor that is activated upon proteolytic cleavage by trypsin, or by binding of its agonist, SLIGRL, resulting in increased melanosome phagocytosis in a Rho-dependent manner (Scott et al. 2003; Seiberg et al. 2000). Expression of PAR-2 by keratinocytes is upregulated by UV exposure in vitro and in vivo (Scott et al. 2001). In the epidermis, the ratio of melanocytes to keratinocytes is 1:34, and the interaction of these cells via transfer of melanosomes has been termed “epidermal melanin unit” (see also Chap. 14). Melanosome transfer is important for normal and uniform skin pigmentation, is increased upon stimulation of melanogenesis, and is critical for optimal photoprotection. 2.3 Pigmentation, the Main Photoprotective Mechanism in the Skin Against Solar UV Solar UV is the main environmental factor that affects skin pigmentation and the main etiological factor for skin cancers, including melanoma (Epstein 1983; Gilchrest et al. 1999; Pathak 1991). Melanin synthesized by melanocytes is the main photoprotective mechanism in the skin (Gilchrest et al. 1999; Halder and Bridgeman-Shah 1995; Pathak et al. 1980). Melanosomes transferred to keratinocytes form supranuclear caps that protect the nucleus from impinging UV rays (Kobayashi et al. 1998). Also, eumelanin acts as a scavenger of reactive oxygen species produced upon exposure to UV, and thus reduces the oxidative damage to DNA, proteins, and lipids (Bustamante et al. 1993). Melanin in the epidermis is also photoprotective for dermal fibroblasts, preventing photoaging caused by UV, particularly long wavelength UVA (Gilchrest and Rogers 1993). An interesting paradigm is that increased melanin synthesis is part of the DNA damage response of melanocytes, as treatment of human skin with DNA oligonucleotides that are homologous to the telomere 3¢ overhang sequence (T-oligos) enhanced nucleotide excision repair and subsequently increased epidermal melanin content (Arad et al. 2006). There is overwhelming clinical and epidemiological evidence for the role of melanin in prevention of sun-induced skin cancers (Epstein 1983; Halder and Bridgeman-Shah 1995; Newton Bishop and Bishop 2005). The incidence of these cancers is by far higher in individuals with fair skin, with low melanin content, than in individuals with dark skin containing high levels of melanin, mainly eumelanin. Experimental evidence shows that exposure to UV results in less DNA photoproducts in dark-skinned individuals with high melanin (mainly eumelanin) content than in light-skinned individuals with low melanin content (Tadokoro et al. 2003). Similarly, an inverse relationship between eumelanin content and the induction of DNA photoproducts was found in cultured human melanocytes derived from donors with different pigmentary phenotypes, with DNA photoproducts being lowest in melanocytes with the highest eumelanin content, and highest in melanocytes that have least eumelanin content (Hauser et al. 2006; Smit et al. 2001). 10 Z.A. Abdel-Malek and V.B. Swope 2.4 Evidence for a Paracrine/Autocrine Network in Human Skin A symbiotic relationship exists between cutaneous melanocytes, keratinocytes, and fibroblasts. It is well established that a complex and well-regulated paracrine/autocrine network is present in human skin, and that this network is upregulated in response to stress, such as in response to UV or inflammation. In turn, the paracrine/autocrine factors mediate many of the stress responses of epidermal cells. Many of the cytokines and growth factors synthesized by keratinocytes and fibroblasts play important roles in regulating melanocyte function and survival (summarized in Table 2.1, Fig. 2.1). The first evidence for keratinocyte-derived paracrine factors that affect melanocytes came from the observation that medium conditioned by cultured human keratinocytes stimulated the proliferation and melanogenesis of cultured normal human melanocytes (Gordon et al. 1989). Additional evidence came from the observation that melanocytes cocultured with keratinocytes exhibited a dose-dependent increase in melanogenesis following irradiation with very low doses of UVB, while melanocytes in monoculture required irradiation with at least a tenfold higher dose of UVB in order to stimulate melanogenesis (Duval et al. 2001). These latter results implicated keratinocyte-derived factors in the melanogenic response of melanocytes to UVB. Medium conditioned with human fibroblasts also stimulated the proliferation of cultured human melanocytes (Imokawa et al. 1998). Collectively, these results provide evidence for the existence of a paracrine network in human skin that modulates melanocyte function, proliferation, and survival under constitutive conditions and in response to UV.