DOES MELANIN MATTER IN THE DARK?

Mystery of melanin

In living cells, melanin pigment is formed within melanosomes, which not only protect the cells from autodestruction, but also serve as second messenger organelles regulating important skin functions, with melanocytes acting as primary sensory and regulatory cells of the epidermis (1). Yet, one can argue that skin melanin, which may negatively affect cellular homeostasis in melanoma (2), really exerts protective functions. Consequently, the actual functions of melanin and the melanogenic pathway in skin biology remains enigmatic (3).

Yet, the solution of this riddle seems simple - to check the actual influence of natural melanin on skin cells in the dark. Since many interesting hypotheses and theories put forward in this respect did not survive confrontation with the experiment, a leading pigment research group from Naples was brave to “jump off the cliff” by confronting theory with experimental reality. They showed that, in the dark, human hair-derived melanin promotes inflammation in keratinocytes, lowers their viability, promotes oxidative stress, and that red pheomelanin does so more strongly than black eumelanin (4). Thus, pheomelanin hardly protects red-haired individuals, even when avoiding the sun. And black hairs don't do much better either, unless they undergo graying.

Toxicity of isolated melanin

A stronger action of pheo- than of eumelanin is one of the most important facts established in the study (4). While some consequences of the demonstrated toxicity of isolated melanins have been emphasized explicitly, there are at least two important consequences of the discovery, delivered indirectly: i) Despite their toxicity, melanins must play some adaptative roles, which outweigh their toxicity in the dark. Moreover, this must concern both types of melanin, because otherwise natural selection would long have eliminated pigmented phenotypes, including the pheomelanotic ones. ii) It is probably the naked melanin itself that is toxic or creates a toxic environment, as the study was carried out on “mature”, natural melanins isolated from hair and purified of low-molecular-weight components.

Caveats for melanin toxicity

The paper by Lembo et al. (4) is probably the prelude to a wider project, and its content possesses, in general, a descriptive character. As the facts are not fully discussed, making it too early to reach definitive conclusions, it is important to pinpoint some caveats for future studies. These concern the actual nature of the melanin-related factor which is toxic, the mode of its action, and its target cells:

• i)Melanin adsorbs various low-molecular weight molecules, which might actually be responsible for the observed toxicity and pro-inflammatory effects. Although the authors attempted to separate them from the pigment, some compounds can be separated only by degradation, in which case one cannot be certain whether the action of the degraded melanin or the admixtures is studied.
• ii)Since the authors do not show that melanin is internalized, the effects could be secondary to non-physiological interactions of melanin with media, thus creating a toxic cell environment and/or to recognition of melanin biopolymer by membrane Toll-like receptors as a foreign agent. All this would trigger cytotoxic and pro-inflammatory effects in keratinocytes (5).
• iii)The authors showed the activity towards melanin-not-producing-but-only-storaging cells, namely keratinocytes. They normally receive melanin embedded in the proteinous matrix of melanosomes and covered by membranous lipid layers separating it from the cytosol. It is, therefore, important to test melanin functions in the cell of origin, melanocytes.

Perspectives in melanin research

The attention paid to dealing with the above caveats may be repaid in valuable results and solutions of other long-standing problems of melanin biology (Fig. 1):

• 1)The mechanism of melanin incorporation by keratinocytes and the actual form of the ingested melanin (amorphic, naked grains or melanosomes), relating to the fundamental problem of melanin transfer between various cells in vivo, including e.g. the transfer of melanin from follicular melanocytes to hair matrix keratinocytes exclusively during the anagen phase of the hair cycle (6).
• 2)The fate of epidermal and extra-follicular skin melanin – its biological activity, degradation and internalization outside the so called “pigmentation synapses”. Perhaps keratinocytes do degrade melanin and the authors observe the side-effects?
• 3)Ectopic melanin with its oxido-reductive capabilities making it interact with many extracellular and cell surface components. Such melanin is accumulated e.g. due to chemotherapy of cancer (7) or in the process of post-inflammatory pigment alterations, and may exert its toxicity even without being internalized by keratinocytes.
• 4)Native melanin constituents. The adsorbed metal ions and low-molecular-weight species which the authors attempted to separate are considered as normal (native) melanin constituents. The actual action of melanin may often be reduced to their toxicity (8).

Figure 1.

Selected primary pathways of melanin action in the light and in the dark. In the light, melanin produced by melanocytes (M) in melanosomes is transferred (blue arrows) to keratinocytes and other target cells (K) where it mainly protects them from sun (umbrella). This may outweigh harmful effects (thunderbolts) observed for isolated melanins in the dark (predominantly pheomelanin). Internalization of extracellular melanin by keratinocytes is unknown (?). Melanin may act directly or via releasing adsorbed substances, mainly metal ions (Me), affecting intracellular antioxidants (NADPH) and changing the metabolism, or inducing inflammation (ILs, TNFs, etc.). It can be modulated by phototype and influenced by exogenous protectors and antioxidants. Red arrow indicates possibility of acting as signal transducer (messenger) conveying information (i).

The paper addresses also numerous more distant phenomena:

• 5)Interference with cell metabolism. Melanin affects the pool of natural antioxidants such as NADPH (3). This nucleotide may not only act as an antioxidant, but also deliver the cancer cells the redox equivalents necessary for production of ATP within the pentose cycle.
• 6)Transfer of information. Along with melanin various signaling particles can be absorbed by the target cells (1). The observed effect may be therefore their active response to a signal, not a passive reaction on intoxication.
• 7)Additional skin protection beyond melanin. The discussed paper convinces one that systemic or local administration of antioxidants and/or anti-inflammatory molecules make sense for preventing (photo)oxidative skin damage.
• 8)Phototype. The stress response of keratinocytes may depend not only on the type of melanin, but also on the phototype of the skin of origin and the associated pro-/antioxidant mechanisms (9).

In summary, this important, interesting and painstaking study (4) promises to, paradoxically, shed more light on the activity of melanin in the dark and answers the title question affirmatively – yes, melanin does matter, even in the dark.