Human skin pigmentation: melanocytes modulate skin color in … · 2017-09-23 · Human skin...

The FASEB Journal • Review

Human skin pigmentation: melanocytes modulate skincolor in response to stress

Gertrude-E. Costin*,1 and Vincent J. Hearing†

*Global R&D, Avon Products, Inc., New Technology Department, Suffern, New York, USA;and †Laboratory of Cell Biology, National Cancer Institute, National Institutes of Health,Bethesda, Maryland, USA

ABSTRACT All organisms, from simple invertebratesto complex human beings, exist in different colors andpatterns, which arise from the unique distribution ofpigments throughout the body. Pigmentation is highlyheritable, being regulated by genetic, environmental,and endocrine factors that modulate the amount, type,and distribution of melanins in the skin, hair, and eyes.In addition to its roles in camouflage, heat regulation,and cosmetic variation, melanin protects against UVradiation and thus is an important defense system inhuman skin against harmful factors. Being the largestorgan of the body that is always under the influence ofinternal and external factors, the skin often reacts tothose agents by modifying the constitutive pigmenta-tion pattern. The focus of this review is to provide anupdated overview of important physiological and bio-logical factors that increase pigmentation and themechanisms by which they do so. We consider endo-crine factors that induce temporary (e.g., during preg-nancy) or permanent (e.g., during aging) changes inskin color, environmental factors (e.g., UV), certaindrugs, and chemical compounds, etc. Understandingthe mechanisms by which different factors and com-pounds induce melanogenesis is of great interest phar-maceutically (as therapy for pigmentary diseases) andcosmeceutically (e.g., to design tanning products withpotential to reduce skin cancer risk).—Costin, G-E.,Hearing, V. J. Human skin pigmentation: melanocytesmodulate skin color in response to stress. FASEB J. 21,976–994 (2007)

Key Words: hyperpigmentation � UV radiation � aging � mela-nocyte photoprotection

Human skin exists in a wide range of different colorsand gradations, ranging from white to brown to black.This is due to the presence of a chemically inert andstable pigment known as melanin, which is produceddeep inside the skin but is displayed as a mosaic at thesurface of the body. Melanin is therefore responsiblefor the most striking polymorphic traits of humans andfor the most obvious and thoroughly discussed aspectof human geographical variability: skin color. Besidesits role in defining ethnicity, melanin plays an essentialrole in defending the body against harmful UV rays andother environmental challenges. Minor changes in the

physiological status of the human body or exposure toharmful external factors can affect pigmentation pat-terns either in transitory (such as in pregnancy) orpermanent (e.g., age spots) manners.

To understand pigmentation of the skin and thefactors that affect it, one must focus on the intricatecellular and molecular interactions between melano-cytes and keratinocytes, which together compose theepidermal melanin unit. All of the other types of cellsdistributed within different layers of the skin and theintracellular signaling pathways often overlapping andinvolving cross-talking must be considered also.

In this review, we provide an update of currentknowledge regarding the effects of endocrine andenvironmental factors on skin pigmentation and themechanisms by which they function. This review em-phasizes that the skin reacts to stress through all itscellular and molecular components, which form acomplicated, sophisticated, and highly sensitive signal-ing network.

SKIN STRUCTURE

The skin plays an extremely important role, providing avast physical barrier against mechanical, chemical, andmicrobial factors that may affect the physiological statusof the body (1). In addition to those functions, the skinalso acts as an immune network and, through itspigments, provides a unique defense system against UVradiation (UV-R) (2). Thus, melanocytes transfer mela-nosomes through their dendrites to keratinocytes,where they form the melanin caps that reduce UV-induced DNA damage in human epidermis. The skin’slayers are represented by the epidermis, the dermis(whose structure will be discussed in more detail be-low), and the hypodermis, the latter consisting of fattytissue that connects the dermis to underlying skeletalcomponents (Fig. 1A).

1 Correspondence: Avon Products, Inc., New TechnologyDepartment, 1 Avon Pl., Suffern, NY 10901, USA. E-mail:[email protected]

doi: 10.1096/fj.06-6649rev

976 0892-6638/07/0021-0976 © FASEB

Epidermis

The epidermis is an external, stratified epitheliumdevoid of blood or nerve supplies of �5–100 �mthickness (which can reach 600 �m on palms and soles)(3). It is composed of several distinct cell populations;keratinocytes and melanocytes are the main constitu-ents, of which the first comprise �95% of the epidermisand are arranged in four layers, as follows (Fig. 1B).

Stratum basale (also known as the stratum germina-tivum) is a single layer of cells attached to a noncellularbasement membrane that separates the epidermis fromthe dermis. The stratum basale consists mostly of basalkeratinocytes, which have stem cell-like properties, andat least two different types of neural crest-derived cells:Merkel cells (neuroendocrine cells responsible for thetransmission of touch sensation through the cutaneousnerves) and melanocytes.

Stratum spinosum contains irregular polyhedral ker-atinocytes with some limited capacity for cell division.Also found here are the bone marrow-derived sentinelcells of the immune system called Langerhans’ cells,which represent the antigen-presenting cells of the skinand play a vital role in immunological reactions such asallergic contact dermatitis.

Stratum granulosum contains flattened, polyhedralnondividing keratinocytes producing granules of a pro-tein called keratinohyalin. These granules increase in

size and number as the cell nuclei gradually degenerateand the cells die. These cells flatten as dividing cellsunderneath them progressively push them toward theskin surface.

Stratum corneum contains nonviable, but biochemi-cally active cells called corneocytes. The keratinocytescontinue to differentiate as they move from the basallayer to the stratum corneum, the result being cornifiedcells that contain abundant keratin and lack cytoplas-mic organelles. It is these cornified cells that provide abarrier against the physical and chemical agents in theenvironment that may adversely affect the body. Morespecifically, this epidermal barrier functions to reducetransepidermal water loss from within and to preventinvasion by infectious agents and noxious substancesfrom without (4).

Dermis

The dermis is a 2 to 4 mm-thick layer of connectivetissue and fibroblasts that houses the neural, vascular,lymphatic, and secretory apparatus of the skin (Fig.1A). The main cell type, fibroblasts, is required forsynthesis and degradation of the extracellular matrix(ECM) (1). This matrix is a complex structure com-posed of highly organized collagen, elastic, and reticu-lar fibers. The dermis also hosts multifunctional cells of

Figure 1. Structure of the skin. A) The differentlayers and components. Collagen, elastin, andother components of the extracellular matrix(ECM) of the dermis are not represented. B)Layers of the epidermis. The stratum lucidumusually present in thick epidermis such as palmsand soles is not included; it is normally locatedbetween the stratum granulosum and stratumcorneum and consists of flattened cells with nonuclei. Merkel cells (stratum basale) and Lang-erhans’ cells (stratum spinosum) are not repre-sented either. Panel B based on and modifiedfrom A. H. Robins (1991) Biological Perspectiveson Human Pigmentation, pp. 2, Cambridge Uni-versity Press, Cambridge, UK.

977HUMAN SKIN PIGMENTATION IN RESPONSE TO STRESS

the immune system such as macrophages and mastcells, the latter being able to trigger allergic reactionsby secreting bioactive mediators such as histamine.Structures within the dermis include: 1) Excretory andsecretory glands (sebaceous, eccrine, and apocrine).Sebaceous glands secrete triglyceride and cholesterol-rich sebum that lubricate the skin and keep it suppleand waterproof. They are often associated with hairshafts. 2) Hair follicles and nails: in addition to gener-ating the hair shaft, the hair follicle provides a protec-tive niche to several stem cell populations in the skin,including keratinocyte stem cells, melanocyte stemcells, a population of epidermal neural crest stem cells,and the dermal stem cell compartment, known as thedermal papilla (5, 6). These stem cells are requiredmost visibly during wound healing. 3) Sensory nervereceptors of Merkel and Meissner’s corpuscles (fortouch), Pacinian corpuscles (for pressure), and Ruffinicorpuscles (mechano-receptors).

In the dermis, collagen provides the skin with tensilestrength and tissue integrity whereas elastin provideselasticity and resiliency. Besides collagen and elasticfibers, the dermis contains the extrafibrillar matrix,which is extracellular and composed of a complexmixture of proteoglycans, glycoproteins, glycosamino-glycans, water, and hyaluronic acid. The most signifi-cant glycosaminoglycans, which bind to proteins toform the proteoglycans of the skin, are chondroitinsulfate, dermatan sulfate, keratin sulfate, heparan sul-fate, and heparin. The most important proteoglycans ofthe skin are versican, which is involved in assuring thetightness of the skin, and perlecan, which is found inbasement membranes. Glycoproteins, such as laminins,matrilins, fibronectin, fibronectin, tenascins, etc., areinvolved in cell adhesion, cell migration, and cell-cellcommunication, which are extremely important pro-cesses taking place in the skin.

Melanocytes, melanosomes, and melanin

Melanin biosynthesis is a complex pathway that appearsin highly specialized cells, called melanocytes, withinmembrane-bound organelles referred to as melano-somes (7). Melanosomes are transferred via dendritesto surrounding keratinocytes, where they play a criticalrole in photoprotection. The anatomical relationshipbetween keratinocytes and melanocytes is known as“the epidermal melanin unit” and it has been estimatedthat each melanocyte is in contact with �40 keratino-cytes in the basal and suprabasal layers (8).

Several important steps must occur for the propersynthesis and distribution of melanin, as follows (9).

1. The development of melanocyte precursor cells(melanoblasts) and their migration from the neural crest toperipheral sites

Prospective melanocytes, known as melanoblasts, derivefrom the neural crest beginning in the second monthof human embryonic life and migrate throughout the

mesenchyme of the developing embryo. They reachspecific target sites, mainly the dermis, epidermis, andhair follicles, the uveal tract of the eye, the striavasculare, the vestibular organ and the endolymphaticsac of the ear, and leptomeninges of the brain. Inhumans, this migration process takes place between the10th and the 12th wk of development for the dermisand �2 wk later for the epidermis (1).

The survival and migration of neural crest-derivedcells during embryogenesis is highly dependent oninteractions between specific receptors on the cellsurface and their extracellular ligands. For example,steel factor, formerly known as mast cell growth factor,KIT ligand, or stem cell factor (SCF), binds the KITreceptor on melanocytes and melanoblasts. Mutationsin the KIT gene decrease the ability of the KIT receptorto be activated by the steel factor and are responsiblefor at least one type of human piebaldism (10). Seehttp://albinismdb.med.umn.edu for other examples ofgenes that regulate pigmentation and, when mutant,are involved in pigmentary disorders.

2. Differentiation of melanoblasts into melanocytes

Once melanoblasts have reached their final destina-tions, they differentiate into melanocytes, which atabout the sixth month of fetal life are already estab-lished at epidermal-dermal junction sites (1).

3. Survival and proliferation of melanocytes

Melanocytes have been identified within fetal epider-mis as early as 50 days of gestation. Dermal melanocytesdecrease in number during gestation and virtuallydisappear by birth, whereas epidermal melanocytesestablished at the epidermal-dermal junction continueto proliferate and start to produce melanin.

4. Formation of melanosomes and production of melanins

Once established in situ, melanocytes start producingmelanosomes, highly organized elliptic membrane-bound organelles in which melanin synthesis takesplace. They can be detected using electron microscopy(EM) as early as during the fourth month of gestation.

Melanosomes are typically divided into four matura-tion stages (I–IV) determined by their structure and thequantity, quality, and arrangement of the melaninproduced (Fig. 2) (11, 12). Nascent melanosomes areassembled in the perinuclear region near the Golgistacks, receiving all enzymatic and structural proteinsrequired for melanogenesis. Stage I melanosomes arespherical vacuoles lacking tyrosinase (TYR) activity (themain enzyme involved in melanogenesis) and have nointernal structural components. However, TYR can bedetected in the Golgi vesicles, and it has been shownthat it is subsequently trafficked to stage II melano-somes. At this point, the presence and correct process-ing of Pmel17, an important melanosomal structuralprotein, determine the transformation of stage I mela-

978 Vol. 21 April 2007 COSTIN AND HEARINGThe FASEB Journal

nosomes to elongated, fibrillar organelles known asstage II melanosomes (12, 13); they contain tyrosinaseand exhibit minimal deposition of melanin. After this,melanin synthesis starts and the pigment is uniformlydeposited on the internal fibrils, at which time themelanosomes are termed as stage III. Their last devel-opmental stage (IV) is detected in highly pigmentedmelanocytes; these melanosomes are either elliptical orellipsoidal, electron-opaque due to complete melaniza-tion, and have minimal TYR activity. The developmen-tal stages detailed above refer mainly to eu-melano-somes (containing black-brown pigments); however,they are quite similar to pheo-melanosomes (contain-ing yellow-reddish melanin), the only difference beingthat the latter remain round and are not fibrillarduring maturation.

Within melanosomes, at least three enzymes areabsolutely required to synthesize different types ofmelanin. While tyrosinase is responsible for the criticalsteps of melanogenesis (including the rate-limitinginitial step of tyrosine hydroxylation), tyrosinase-relatedprotein 1 (TYRP1) and DOPAchrome tautomerase(DCT) are further involved in modifying the melanininto different types. Besides these, melanosomes con-tain other melanocyte-specific proteins that have struc-tural functions (e.g., Pmel17, as mentioned above) orprobably are involved in regulating the pH withinmelanosomes, such as P protein- or membrane-associ-ated transporter protein (MATP), or that play as yetunclear roles, such as the melanoma antigen recog-nized by T cells 1 (MART1) or oculocutaneous albi-nism-1 (OA-1) protein (14).

TYR (monophenol, 3,4-�-dihydroxyphenylalanineoxygen oxidoreductase, EC 1.14.18.1) is a single chaintype I membrane glycoprotein catalyzing the hydroxy-lation of tyrosine to �-3,4-dihydroxyphenylalanine(DOPA) (which is the initial rate-limiting step in mel-anogenesis) and the subsequent oxidation of DOPA toDOPAquinone. TYR, TYRP1, and DCT share numerousstructural similarities and follow quite similar biosyn-

thetic, processing, and trafficking pathways (15). Theirmaturation is assisted by chaperones, calnexin beingthe most important one due to its involvement in thecorrect folding of tyrosinase (16–18). The subsequentmetabolism of DOPA and its derivatives by variousmelanocyte-specific enzymes, including TYRP1 andDCT, results in the synthesis of eumelanin, a black-brown pigment. Briefly, 5,6-dihydroxyindole (DHI)melanins are generated from DOPAquinone after sev-eral steps of decarboxylation, oxidation, and polymer-ization. However, in the presence of DCT, the carbox-ylic acid group of 5,6-dihydroxyindole-2-carboxylic acid(DHICA) is retained when derived from DOPAchrome,and therefore the so-called DHICA melanins are pro-duced. The synthesis of pheomelanin involves theproduction of cysteinyldopa conjugates from DOPAqui-none after the production of DOPA from tyrosine.TYRP1 is important for the correct trafficking of tyrosi-nase to melanosomes (19), and DCT also seems to beinvolved in the detoxification processes (20) takingplace within melanosomes.

Melanins are polymorphous and multifunctionalbiopolymers that include eumelanin, pheomelanin,mixed melanins (a combination of the two), and neu-romelanin. Mammalian melanocytes produce twochemically distinct types of melanin pigments: black-brown eumelanin and yellow-reddish pheomelanin(21). Although they contain a common arrangement ofrepeating units linked by carbon-carbon bonds, mela-nin pigments differ from each other with respect totheir chemical, structural, and physical properties. Eu-melanin is a highly heterogeneous polymer consistingof DHI and DHICA units in reduced or oxidized states,as detailed above; pheomelanin consists mainly ofsulfur-containing benzothiazine derivatives (22). Dueto their chemical structure, both eumelanin andpheomelanin are involved in binding to cations, an-ions, drugs, and chemicals, etc., and therefore play animportant protective role within melanocytes (23).Neuromelanin, which is produced in dopaminergic

Figure 2. Schematic of a melanocyte: the en-gine for melanin production. The four devel-opmental stages of melanosomes are shown asthey move toward the periphery of the cellwithin the dendrites. The melanin biosynthesispathway is displayed in the inset showing themain enzymes involved. Asterisks in differentshades of gray represent different stages ofglycosylation for the melanogenic proteins asthey are transported through the ER and Golgiapparatus.


neurons of the human substantia nigra, can also che-late redox active metals (Cu, Mn, Cr) and toxic metals(Cd, Hg, Pb), and thus protects against their ability topromote neurodegeneration (24).

Given their complexity, melanosomes can be used asa model to study organelle biogenesis, protein traffick-ing and processing, organelle movement, and cell-cellinteractions (like those occurring during melanintransfer between melanocytes and keratinocytes) (25).Therefore, even minor changes in the cellular environ-ment affect melanosomes and pigmentation. Numer-ous intrinsic and extrinsic factors, including body dis-tribution, ethnicity/gender differences, variablehormone-responsiveness, genetic defects, hair cycle-dependent changes, age, UV-R, climate/season, toxin,pollutants, chemical exposure and infestations, areresponsible for a whole range of responses in melano-some structure and distribution under different typesof stress.

Cutaneous pigmentation is the outcome of two im-portant events: the synthesis of melanin by melanocytesand the transfer of melanosomes to surrounding kera-tinocytes (26). Although the number of melanocytes inhuman skin of all types is essentially constant, thenumber, size, and manner in which melanosomes aredistributed within keratinocytes vary. The melanin con-tent of human melanocytes is heterogeneous not onlybetween different skin types but also between differentsites of the skin from the same individual. This hetero-geneity is highly regulated by gene expression, whichcontrols the overall activity and expression of melano-somal proteins within individual melanocytes (27). Ithas been shown that melanocytes with a low melanincontent synthesize TYR more slowly and degrade itmore quickly than melanocytes with a higher melanincontent and TYR activity (28). In general, highly pig-mented skin contains numerous single large melanoso-mal particles (0.5–0.8 mm in diameter), which areellipsoidal and intensely melanotic (stage IV). Lighterpigmentation is associated with smaller (0.3–0.5 mm indiameter) and less dense melanosomes (stages II andIII), which are clustered in membrane-bound groups(29). These distinct patterns of melanosome type anddistribution are present at birth and are not deter-mined by external factors (such as sun exposure). They

are responsible for the wide variety of skin complexions(Fig. 3).

Epidermal melanin unit and the involvement ofkeratinocytes in melanin production

The epidermal melanin unit is a functional and struc-tural complex within the epidermis consisting of twocell types: melanocytes and keratinocytes. The variationin skin color among various races is determined mainlyby the number, melanin content, and distribution ofmelanosomes produced and transferred by each mela-nocyte to a cluster of keratinocytes surrounding it (30).Once in keratinocytes, the melanin granules accumu-late above the nuclei and absorb harmful UV-R beforeit can reach the nucleus and damage the DNA. Whenmelanin is produced and distributed properly in theskin, dividing cells are protected at least in part frommutations that might otherwise be caused by harmfulUV (31). The melanocyte-keratinocyte complex re-sponds quickly to a wide range of environmental stim-uli, often in paracrine and/or autocrine manners.Thus, melanocytes respond to UV-R, agouti signalingprotein, melanocyte-stimulating hormone (MSH), en-dothelins, growth factors, cytokines, etc. After UV-Rexposure, melanocytes increase their expression ofproopiomelanocortin (POMC, the precursor of MSH)and its receptor melanocortin 1 receptor (MC1-R), TYRand TYRP1, protein kinase C (PKC), and other signal-ing factors (32, 33). On the other hand, it is known thatUV stimulates the production of endothelin-1 (ET-1)and POMC by keratinocytes and that those factors canthen act in a paracrine manner to stimulate melanocytefunction (34, 35). In addition to keratinocytes, fibro-blasts, and possibly other cells in the skin producecytokines, growth factors, and inflammatory mediatorsthat can increase melanin production and/or stimulatemelanin transfer to keratinocytes by melanocytes. Me-lanocyte growth factors affect not only the growth andpigmentation of melanocytes but also their shape,dendricity, adhesion to matrix proteins, and mobility.

�-MSH, ACTH, basic fibroblast growth factor(bFGF), nerve growth factor (NGF), endothelins, gran-ulocyte-macrophage colony-stimulating factor (GM-

Figure 3. Human pigmentation—the main skin types: African-American, Asian, Caucasian, and Hispanic (left to right).


CSF), steel factor, leukemia inhibitory factor (LIF), andhepatocyte growth factor (HGF) are keratinocyte-de-rived factors that are thought to be involved in theregulation of the proliferation and/or differentiationof melanocytes (36), some acting through receptor-mediated signaling pathways (Fig. 4). It has been shownthat in human epidermis, �-MSH (32, 37) and ACTH(32, 37, 38) are produced in and released by keratino-cytes and are involved in regulating melanogenesisand/or melanocyte dendrite formation. �-MSH andACTH bind to a melanocyte-specific receptor, MC1-R(39), which activates adenylate cyclase through G-protein, which then elevates cAMP from adenosinetriphosphate (40). Cyclic AMP exerts its effect in partthrough protein kinase A (PKA) (41), which phosphor-ylates and activates the cAMP response element bind-ing protein (CREB) that binds to the cAMP responseelement (CRE) present in the M promoter of themicrophthalmia-associated transcription factor (MITF)gene (42, 43). The increase in MITF-M expressioninduces the up-regulation of TYR, TYRP1, and DCT(42, 43), which leads to melanin synthesis.

Keratinocytes also produce and release NGF, which isinvolved in regulating the melanogenesis and/or den-dritogenesis of melanocytes (44). Its expression is up-regulated by UV-R, suggesting yet another paracrineinfluence of keratinocytes on melanocytes with possiblerelevance to the tanning response. It has been shownthat normal human melanocytes express the NGF re-ceptor (45) and also high-affinity receptors for NGF(TRK-A) and NT-3 (TRK-C) (44).

ET-1 is a 2l amino acid peptide with vasoactiveproperties first isolated from endothelial cells and laterfound to be synthesized and secreted by keratinocytesas well (46–48), particularly after exposure to UV-R(46–48). The overall effect of ET-1 is the increase ofmelanocyte dendricity and the enhancement of mela-nocyte migration and melanization (48). Binding ofET-1 to its G protein-coupled receptor (ETBR) on

melanocytes activates a cascade of signaling pathways,resulting in mobilization of intracellular calcium, acti-vation of PKC, elevation of cAMP levels, and activationof mitogen-activated protein kinase (MAPK) (49, 50).UV-R stimulates keratinocytes to produce ET-1 and alsoinduces interleukin-1 (IL-1) production in these cells.IL-l is known to induce ET-1 in keratinocytes in anautocrine manner. Therefore, it has been suggestedthat these intracellular events in keratinocytes lead toincreased TYR mRNA, protein, and enzymatic activityin neighboring melanocytes as well as to an increase inmelanocyte number (51).

Prostaglandin (PG) E2 and PGF2� are known to beproduced and released from human keratinocytes bythe stimulation of proteinase-activated receptor 2(PAR-2). PGE2 and PGF2� stimulate the dendritogen-esis of human epidermal melanocytes in culture (52)through EP1, EP3, and FP receptors. Their influenceon melanocyte dendricity has been suggested to becAMP-independent and might be mediated throughphospholipase C (PLC) (52).

bFGF (53) and SCF (54) are also expressed bykeratinocytes. Those secreted factors are involved inregulating the proliferation and melanogenesis/den-dritogenesis of human epidermal melanocytes in nor-mal skin (38, 44, 55) and/or in UV-A (56)/UV-B (32,46, 48, 53, 54) -irradiated skin.

HGF binds to its specific receptor, c-Met (57), acti-vates MAPK, and elicits the up-regulation of proteinsrequired for melanocyte proliferation (58, 59).

GM-CSF binds to its specific receptor, GM-CSFR(60), activates the signal transducer and activator oftranscription (STAT-1, STAT-3, and STAT-5) (61, 62)or MAPK (63), and induces the up-regulation of pro-teins required for the proliferation of melanocytes aswell as of TYR, TYRP1, and DCT.

The epidermis has a complex network that secretesas well as responds to autocrine and paracrine cytokinesproduced by keratinocytes and melanocytes, respec-

Figure 4. Scheme of signaling pathways withinthe epidermal melanin unit and mechanisms bywhich keratinocyte-derived factors act on hu-man melanocyte proliferation and differentia-tion. Based on and modified from T. Hirobe(2005) Role of keratinocyte-derived factors in-volved in regulating the proliferation and dif-ferentiation of mammalian epidermal melano-cytes. Pigment Cell Res., vol. 18, pp. 2–12.


tively. Human melanocyte proliferation requires thecross-talking of several signaling pathways including thecAMP/PKA, PKC, and tyrosine kinase pathways. There-fore, the mechanisms by which various factors increaseskin pigmentation are closely inter-related and will befurther discussed below.

There are numerous internal and external stressesthat affect human skin pigmentation. The list is fairlylong, so for this review we decided to focus on just acouple of the more common stresses whose mecha-nisms of action are known to some extent or arecurrently under investigation and whose use may affectthe discovery of new approaches to reduce hyperpig-mentation. We consider external factors (UV-R: tan-ning and photoaging; drugs, chemicals, etc.) and inter-nal factors (hormonal influences; inflammation:postinflammatory hyperpigmentation).

HYPERPIGMENTATION INDUCED BYEXTERNAL FACTORS

UV influence on human pigmentation

The skin responds to UV-R exposure by developing twodefensive barriers: thickening of the stratum corneumand the elaboration of a melanin filter in cells of theepidermis. The palms and soles are the regions with thethickest stratum corneum, and they are exceptionallyresistant to UV damage. The keratins and proteinswithin the stratum corneum act mainly by scatteringand absorbing the UV. UV-R sets in action an inte-grated mechanism for the formation and delivery ofmelanin within melanosomes from melanocytes to ker-atinocytes. This mechanism is probably triggered bykeratinocytes, which respond to UV-R with bursts ofmitoses and with increased production of ET-1 andPOMC, thus creating a new demand for melanosomes.The mitotic rate of basal keratinocytes increases a dayafter UV exposure, reaches a maximum 2 days later,and maintains this level for �1 wk. It then declines, andthe skin regains its original thickness after 1–2 monthsif there has been no subsequent exposure. After UV-R,the epidermal melanin unit responds with increasedlevels of TYR activity, increased synthesis of melano-somes, and higher rates of melanosome transfer tokeratinocytes to meet the new demand for melano-somes created by the proliferation of keratinocytes(64).

UV is part of the electromagnetic spectrum and it liesbetween the visible and X-ray regions. Only 5–10% ofthe total radiant energy received at the surface of theEarth from the sun is UV, and the rest is dividedbetween the visible (40%) and the infrared (50%) (65).According to wavelengths, UV radiation is divided inUV-A (320–400 nm), UV-B (280–320 nm), and UV-C(200–280 nm); the latter is normally screened by theozone layer and does not reach the Earth’s surface, likemost wavelengths �280 nm.

UV-A passes through most glass in automobiles,

offices, and windows whereas UV-B is blocked by win-dow glass. UV-A also penetrates deep into the dermis; itis estimated that �19–50% of the solar UV-A can reachthe depth of melanocytes, whereas only �9–14% ofsolar UV-B reaches these cells. Therefore, UV-A stimu-lates melanin pigmentation, but the resultant tan ap-pears to be transient and less protective against UV-induced injury than tans generated after UV-Bexposure. Although the amount of UV-A reaching theEarth’s surface is several orders of magnitude greaterthan the amount of UV-B, UV-A has 1000-fold lesserythema-producing effects than UV-B (65). However,Garland et al. were the first to hypothesize that UV-Acould cause melanoma in humans. The potential car-cinogenic effect of UV-A has also been demonstrated incultured human melanocytes (66). The authors sug-gested that endogenous pigments and/or melanin-related molecules seem to enhance DNA breakage afterUV-A irradiation. UV-A must first react with endoge-nous photosensitizers (flavins, porphorins, melanins),which in turn generate reactive oxygen species (ROS),which finally causes single-strand breaks or photoad-ducts.

UV-B is responsible for causing the sunburn reactionwithin the skin and is absorbed mainly by the epidermisand upper dermis. Like UV-A, UV-B stimulates theproduction of melanin, which constitutes the basis fortanning. UV-B has great potential to induce erythema,and therefore its influence on the skin has beenthoroughly investigated in vitro and in vivo (64). TheUV-B portion of the spectrum can promote skin cancer,especially if the exposure has been repeated and pro-longed. However, recent studies have shown that theuse of narrow-band UV-B (NB-UVB �311 nm) is abetter choice for phototherapy than frequently usedpsoralen and UV-A therapy (PUVA), which can causeundesirable side effects, including cutaneous cancers(67, 68–70).

The only known beneficial effect of UV-B is thestimulation of vitamin D synthesis in the epidermis.Vitamin D promotes the absorption of calcium fromthe intestine and ensures the proper mineralization ofbones. However, exposure of just a small area of thebody to a small amount of UV-B (5% of that requiredfor erythema) is all that is needed for adequate synthe-sis of the vitamin D in the skin (64).

One role of melanin in the skin is to neutralize theROS generated by a variety of factors, including UV-B(23), therefore functioning like a natural sunscreen.Until recently it was thought that the higher themelanin content, the less chance of DNA damageresulting from UV-R exposure. In a recent study, theeffects of melanin on UV responses in different racial/ethnic groups were investigated for the first time.Despite the general public assumption that dark skintypes are UV resistant and therefore not adverselyaffected by UV, this study showed that even the darkestUV-resistant skin types accumulated significant DNAdamage at levels �1 minimal erythema dose (MED)(71). The authors demonstrated that even very low UV


exposures cause measurable damage to DNA in all skintypes, although it was obvious that the most severe DNAdamage was in lightly pigmented skin.

The influence of UV on human pigmentation will bedetailed further from the perspective of tanning as wellas photoaging as a perfect example of factors sharingintracellular pathways with slightly different end resultson the skin.

Tanning response to UV-R

In humans, an increase of skin pigmentation over thebasal constitutive level is called tanning, and this isphysiologically stimulated by UV-R. UV-induced skindarkening involves an increase in the number of mela-nocytes as well as stimulation of melanin synthesis andmelanocyte dendricity, a crucial morphological featurerequired for melanin transfer to keratinocytes.

The tanning response has been shown to have twodistinct phases, termed immediate pigment darkeningand delayed tanning. Both have strong genetic deter-minants and are generally more pronounced in indi-viduals with dark baseline (constitutive) pigmentation(72).

Immediate tanning is a quick but transient brownishtan that follows the exposure of skin to UV-A or visiblelight. It begins immediately after exposure, reaches amaximum within 1–2 h, then fades between 3 and 24 hafter exposure (72). Certain ultrastructural changeshave been observed in melanocytes during immediatetanning: the appearance of thick filaments and micro-tubules and the translocation of melanosomes from theperinuclear area to the dendritic processes but noactual increase in the size or number of melanosomes.It thus seems possible that the immediate tanningreaction is based on the photoxidation of preexistingmelanin, melanin precursors, or even of other epider-mal constituents and/or their redistribution in theepidermis.

Delayed tanning gives rise to a durable tan inducedby repeated exposure mainly to UV-B, but also to UV-Aor to visible light. It is a gradual process in which theskin starts darkening 48–72 h after irradiation, reachesa maximum �3 wk after exposure, and the skin doesnot return to its original melanin content until �8–10months later (72). Delayed tanning is dependent onboth qualitative and quantitative changes within mela-nocytes, which enlarge in size, increase their dendricity,and develop a diffuse distribution of thick filaments intheir cell bodies. Ribosomes, ER, and Golgi apparatusare more prominent, reflecting an increase in thesynthesis of TYR and melanosomes in all developmen-tal stages, in their melanization, and in the number thatare transferred to keratinocytes. Therefore, delayedtanning is due to an increase in melanocyte numbersand melanogenesis.

Like all photobiologic responses, tanning requiresdirect interaction of UV photons with molecular targets(chromophores) in the skin. The major cellular chro-mophores that absorb in the UV-B range are nucleic

acids (mainly the pyrimidine and purine bases) andproteins (mostly tryptophan and tyrosine); cytotoxicity,mutagenesis, and new protein synthesis are all initiatedby photoreactions involving DNA (73). However, someresponses, such as activation of membrane enzymesand induction of early response genes, involve non-nuclear chromophores. Other biomolecules that ab-sorb UV-B include the reduced form of NAD, quinones,flavins, and other heterocyclic cofactors such as tetra-hydrobiopterin. In skin, UV-A- and UV-B-absorbingmolecules are present in addition to the above-men-tioned cellular chromophores and include 7-dehydro-cholesterol, urocanic acid, and melanin (73).

The products formed in DNA after absorbing UV-Bradiation have been studied extensively because of theirmajor role in the mechanisms underlying UV carcino-genesis. Two major types of bipyrimidine photoprod-ucts are created following UV-R: cyclobutylpyrimidinedimers (CPDs) and (6–4) photoproducts (Fig. 5; ref.73). Cells are equipped with a complicated and highlyregulated apparatus for repairing the DNA damageproduced in this manner; however, if that does notoccur with high efficiency and fidelity, the cells accu-mulate mutations that can eventually lead to skincancer.

UV-R also causes peroxidation of lipids in cellularmembranes, leading to generation of ROS, which maystimulate melanocytes to produce excess melanin (74).Usually, only lipids containing two or more conjugateddouble bonds in their structure absorb UV-B and thusliberate arachidonic acid, which is subsequently metab-olized to various species of PGs and leukotrienes,generates previtamin D3 from 7-dehydrocholesterolwith subsequent processing to various photoproductsand the biologically active 1�,25-dihydroxy-vitamin D3,and releases diacylglycerol (DAG), which in turn acti-vates PKC, among other possible roles in signal trans-duction (75; Fig. 5). It was observed that addition ofDAG to cultured human melanocytes increases theirmelanin content severalfold within 24 h (76), andsubsequent work demonstrated that UV-R acts synergis-tically with DAG to enhance melanogenesis (77). Thesedata suggest that DAG may be a physiological mediatorof the tanning response in UV-irradiated skin.

Direct melanogenic effects of UV on melanocytesmight also involve the production of NO, which isconsidered a major intra- and intercellular messengermolecule. NO elicits its effects through the activation ofa soluble guanylate cyclase, leading to an increase inintracellular cGMP content and the activation of cGMP-dependent protein kinase. Furthermore, it has beenshown that UV-R increases both NO and cGMP produc-tion, suggesting they are both required for UV-B-induced melanogenesis (Fig. 5).

Melanin plays a major photoprotective role inhuman skin by absorbing, scattering, photo-oxidiz-ing, and scavenging free radicals and acting as apseudo-dismutase to minimize the toxic effects ofROS and to prevent damage to DNA, proteins, andcell membrane lipids (78). It is known that UV-A


produces harmful oxygen species such as O.2-, .OH,

and 1O2 and that melanin interacts with them, thusprotecting the skin against the damage that couldoccur (78, 79).

Overall, UV-R acts on human skin by increasingtranscription of the TYR gene and the function ofMC1-R on melanocytes; it also increases expression ofPOMC and its derivative peptides by keratinocytes andcells within the dermis and the release of DAG from theplasma membrane, which in turn activates PKC. UV-Ralso activates the NO/cGMP pathway and the produc-tion of growth factors and ET-1 by keratinocytes, andinduces a SOS response to UV-induced DNA damage,which sets into action the cellular DNA repair mecha-nisms.

Photoaging: the UV-R contribution to the appearanceof solar lentigines (age spots)

Aging of the skin is a complex process induced by bothchronologic and environmental factors (mainly UV-R).Changes in the skin associated with chronologic aginginclude dryness, increased fragility, decreased epider-mal and dermal thickness (both influenced by circulat-ing levels of estrogens and potentially by androgens),fragmentation of elastic fibers (most likely influencedby circulating levels of estrogens), decreased sebumproduction, and the number and function of apocrineglands (probably influenced by levels of circulatingandrogens). Changes in the skin due to photoaginginclude wrinkles and furrows, solar lentigines (SL),mottled pigmentation, actinic keratoses, basal cell andsquamous cell carcinomas, subsets of melanomas, etc.(80).

Chronologic aging leads to a decrease in the turn-over rate of the epidermis as well as the flattening of

rete ridge patterns, which results in a decreased surfacearea of the basement membrane. This is clinicallydescribed as an increased fragility of the skin. Chrono-logic aging of the dermis is reflected by decreases incollagen content and thickness. Additional age-relatedchanges include decreases in the vascularity of thedermis as well as in the number of functional fibro-blasts. Clinically, this results in increased times requiredfor wound healing. Dermal elastic fibers are also af-fected by chronologic aging and undergo fragmenta-tion and granular disintegration by age 70 (mainlythose in the upper dermis) (80, 81).

In photoaging (sun-induced skin aging), the de-gree of damage to the epidermis depends on thecumulative dose of sun exposure as well as on theamount of protection provided by its pigmentation.In the dermis, the primary effects of photoaginginvolve degeneration of collagen and the depositionof abnormal elastotic material. The clinical manifes-tations of these changes include wrinkles and furrowsof the face, as mentioned above; this is in distinctionto the laxity (sagging) and fine wrinkling of non-sun-exposed areas that are due to chronological aging ofthe dermis (81).

From the viewpoint of pigmentation, aging results ina decline in functional melanocytes in both the skinand hair. Various studies have indicated that the num-ber of functioning DOPA-positive melanocytes in non-exposed human skin decreases with age by 8–20% ofthe surviving population each decade. However, inUV-irradiated skin there are approximately twice asmany melanocytes as in unexposed areas, but there isstill a comparable decrease in melanocytes with age. Itis surprising that, unlike hair color, there is no loss ofskin pigmentation with age. In fact, the chronicallysun-exposed skin of an older person is usually more

Figure 5. Mechanisms involved in the (hyper)-pigmentation induced by UV-R. The tanningresponse is determined by a complex set ofregulatory processes involving direct effects ofUV-R on melanocytes and indirect effectsthrough the release of keratinocyte-derived fac-tors.


pigmented than that of a younger subject of similarcomplexion despite the lower melanocyte density in theformer. This paradox has been explained by the greaterfunctional activity in older melanocytes after manyyears of cumulative sun exposure (82). An interestingfinding recently reported (83) is that DCT is notexpressed by melanocytes of human hair comparedwith human skin. This could potentially contribute tothe premature loss of melanin production by functionalmelanocytes in human hair with age due to addedcytotoxic stress of melanogenesis in the absence ofDCT.

Given the theme of this review, we outline thecurrent understanding of mechanisms responsible forthe appearance of SL, also known as senile lentigo,sun-, liver-, or age spots. Solar lentigines are circum-scribed, pigmented macules, which are usually lightbrown, but vary in degree of color to jet black. SL aretypically found on UV-exposed areas of the body (theface, dorsum of the hand, extensor forearm, and upperback) (Fig. 6). They can range anywhere in size from�1 mm up to a few centimeters in diameter and, inareas of severely sun-damaged skin, may coalesce intoeven larger lesions (84) (Table 1).

The molecular mechanisms responsible for skinhyperpigmentation in SL have recently been eluci-dated by the group of Imokawa. They found a 2-foldincrease of TYR-positive cells per length of thedermal/epidermal interface in SL lesions comparedwith unaffected skin (85). That group also showedthat there is a molecular regulatory network betweenmelanocytes/keratinocytes and melanocytes/dermalfibroblasts in which ET-1 and SCF are key regu-lators in the development of hyperpigmentation inSL (86, 87).

Exposure to UV-R induces an increase in the produc-tion of ET-1 by keratinocytes, and its secretion there-fore stimulates melanocytes to produce melanin. Asdescribed above, the actions of endothelins on melano-

cytes are initiated by the binding of ET-1 to its receptor(ETBR), followed by sequential signaling processesinvolving PKC and MAPK. On the other hand, SCF(also produced by keratinocytes) binds to the c-KITreceptor on melanocytes, thus mediating dimerization,activation of its intrinsic tyrosine kinase activity, andautophosphorylation (88). The activated c-KIT recep-tor then phosphorylates various substrates and associ-ates with various signaling molecules including phos-phatidylinositol 3-kinase, the Shc and Grb2 adaptorproteins, and the guanine nucleotide exchange factor,SOS, all of which lead to activation of the Ras-MAPKpathway (89) (Fig. 7).

The potential of keratinocytes located in SL lesionalepidermis to produce ET-1 is significantly higher thanin perilesional normal controls (46), and there is anaccentuated expression of ETBR transcripts as well(85). The increased production and localization ofET-1 was paralleled by increased amounts of TYR inmelanocytes. These findings suggest that stimulation ofthe epidermal ET cascade, especially with respect to theexpression of ET-1 and ETBR, plays an important rolein the mechanism involved in the hyperpigmentationof LS. The ET-1-inducible cytokine, tumor necrosisfactor �, is consistently up-regulated in the SL lesionalepidermis, suggesting its involvement in this hyperpig-mentation disorder through the induction of ET-1(85).

Imokawa et al. (90) also showed that SL lesionalepidermis expresses increased levels of SCF mRNAtranscripts and protein compared with nonlesionalcontrols. In epidermal keratinocytes, SCF is expressedas a membrane-bound form (mSCF), not in a secretoryor soluble cytokine form such as ET-1 (54), even afterUV-B exposure (54). In contrast, dermal fibroblasts cansecrete soluble SCF (sSCF), probably because of theaction of proteolytic enzymes capable of cleaving mSCFto release sSCF (91). Expression of sSCF could not bedetected in SL lesional epidermis, suggesting that the

Figure 6. Pictures of skin at different agesshowing differences in pigmentation pattern.


increased production of mSCF plays an essential role instimulating the proliferation and melanogenesis ofmelanocytes, leading to epidermal hyperpigmentationin SL (Fig. 7).

Therefore, the mechanism currently proposed forthe appearance of SL involves the stimulation of twoepidermal cascades, consisting of ET-1/ETBR andSCF/c-kit, and the cross-talk between those two afterthe UV exposure. However, other cascades in the skinmay also contribute to the hyperpigmentation seen inSL; these may be discovered as work in this fieldprogresses.

The only data regarding the analysis of SL in thedermal compartment of the skin became available justrecently (92). That study showed that the number ofmelanophages is increased in SL compared with unaf-fected skin in the same subject. These melanophageswere identified as FXIIIa� dermal dendrocytes; theyseem to be the main cell type that uptakes melanin(most likely from the epidermis, where it is increased in

SL) by a mechanism that remains to be determined. Itwas also reported recently that dickkopf (DKK1), se-creted by fibroblasts in the dermis, plays an importantrole in regulating melanocytes above them (93); work isneeded to resolve the role of that factor, if any, inhyperpigmentation of the skin.

The action of drugs, chemicals, etc., on humanskin pigmentation

Numerous common drugs can stimulate human skinhyperpigmentation such as certain antibiotics (sulfon-amides and tetracyclines), diuretics, nonsteroidal anti-inflammatory drugs, pain relievers, and some psycho-active medications.

The use of oral contraceptives has been associatedwith the development of discoloration of the cheeks,forehead, and nose (94) similar to chloasma (furtherdetailed below). Microscopic examination of the epi-

TABLE 1. Summary of hyperpigmentation conditions described, their causative factors, clinical features, and cellular characteristics

Hyperpigmentationdisorder Causative factor(s) Clinical features

Cellularcharacteristics Molecular markers modified

Solar lentigines (SL) Induced by UV-R Circumscribed, brown toblack macules

Range from �1 mm toseveral cm

Occur in epidermis

Found on UV-exposedareas of the body suchas the face, dorsum ofthe hand, extensorforearm, upper back

Increased melaninproduction

Slight increase innumber ofmelanocytes

Increased TYR-positive cellsper length of thedermal/epidermalinterface compared withunaffected skin

Keratinocytes’ potential toproduce ET-1 issignificantly highercompared withunaffected skin

TNF� is up-regulated inthe SL lesional epidermis

Melasma Exacerbated by sunexposure,pregnancy, oralcontraceptives,certain anti-epileptics, etc.

Symmetric facialhyperpigmentation

May involve epidermis,dermis, or both


Normal numberof melanocytes

Melanocytes arelarger, moredendritic

High levels ofprogesterone, estrogen,and MSH

Increased transcription ofgenes encoding DCT,TYR

Post-inflammatoryhyperpigmentation

Develops afterresolution ofacne, contactdermatitis

Discrete hyperpigmentedmacules with hazymargins

May involve epidermis,dermis or both


Normal numberof melanocytes

PGE2 and PGF2� synthesisis up-regulated; they actas paracrine factorswhich stimulatemelanocyte dendricity

Leukotrienes andthromboxanes may beresponsible for theinduction of post-inflammatoryhyperpigmentation


dermis revealed increased melanogenesis and the pres-ence of enlarged melanocytes.

Certain antiepileptic agents (mainly hydantoins) mayalso cause skin hyperpigmentation (95). Their long-term use induces a brownish coloration of the face andneck, similar to chloasma of pregnancy. It has beenshown that melanin concentration is particularly in-creased in females; Caucasians especially seem to bemore affected (96).

It is already known that chloroquine has an affinityfor melanin and causes skin hyperpigmentation. Differ-ent studies have detected melanin in the dermis ofpatients undergoing chloroquine treatment (97).

Levodopa, often used to treat Parkinson’s disease,also induces hyperpigmentation of the skin (64).DOPA is normally transformed into melanin withinmelanosomes; therefore, DOPA therapy (applied aslevodopa treatment) may possibly enhance melaninbiosynthesis, perhaps even by extracellular oxidation,although the literature lacks strong evidence to supportthis hypothesis.

Heavy metals can also elicit hyperpigmentation,which can arise after the extensive use of drugs con-taining arsenic, bismuth, gold, or silver (98). Themetals are believed to act by binding, and therebyinactivating, sulfydryl compounds in the skin that nor-mally inhibit TYR activity. Removal of this inhibitionstimulates melanogenesis. Mercury products inactivateTYR probably by replacing the essential copper in theenzymatic site of that protein.

Some chemotherapy agents also can cause hyperpig-mentation, the most common ones being cyclophos-phamide, 5-fluorouracil, doxorubicin, daunorubicin,and bleomycin. Their mechanisms of action are cur-

rently unknown but may involve direct toxicity, stimu-lation of melanocytes, and/or inflammation.

HYPERPIGMENTATION INDUCED BYINTERNAL FACTORS

Hormonal influence on human skin pigmentation

Hyperpigmentation is sometimes seen during preg-nancy and this condition is called melasma, chloasma,or mask of pregnancy; it occurs mainly on the cheeks,upper lip, chin, and forehead. It is characterized by asymmetrical hypermelanosis with an irregular colora-tion, ranging from light brown to gray and dark brown.Although melasma is usually associated with pregnancy,multiple other factors can contribute to its develop-ment including UV exposure, hormone therapy, estro-gen-containing oral contraceptives, genetic influences,certain cosmetics, endocrine or hepatic dysfunction,and selected antiepileptic drugs (Table 1). Of theenvironmental sources, UV-R is obviously the mostinfluential (84, 99).

Recent studies have shown that the areas of hyper-pigmentation seen in melasma exhibit increased depo-sition of melanin in the epidermis and dermis (100,101). No increase in the number of melanocytes inthose areas was noted, but the melanocytes were larger,more dendritic, and showed increased melanogenesis,producing especially eumelanin (101). That study con-firmed an increased number of melanosomes in kera-tinocytes, melanocytes, and dendrites in lesional skincompared with nonlesional skin. During pregnancy(especially in the third trimester), elevated levels ofestrogen, progesterone, and MSH have often been

Figure 7. Potential mechanisms involvedin the appearance of senile lentigines.The dotted arrows suggest there are in-termediates involved in those pathways(see sSCF, mSCF, and HGF signaling);solid arrows describe the fully detailedpathway (see ET-1 signaling).


found in association with melasma (102, 103). TYRactivity increases and cellular proliferation is reducedafter treatment of melanocytes in culture with �-estra-diol (104). Sex steroids increase transcription of genesencoding melanogenic enzymes in normal human me-lanocytes, especially those for DCT and TYR (105).These results are consistent with the significant in-creases in melanin synthesis and TYR activity reportedfor normal human melanocytes under similar condi-tions in culture (106).

Since melanocytes contain both cytosolic and nu-clear estrogen receptors (107), melanocytes in patientswith melasma may be inherently more sensitive to thestimulatory effects of estrogens and possibly to othersex steroid hormones. Recent studies suggest that es-trogens exert their effect in skin through the samemolecular pathways used in other nonreproductivetissues (Fig. 8) (108). There is evidence that 17�-estradiol can use both signaling pathways (eithergenomic or nongenomic) in epidermal keratinocytes(109).

It is known that estrogens improve skin moisture andalso increase its thickness and collagen content. There-fore, estrogen plays a key role in skin aging homeostasisgiven the fact that skin appearance declines quickly inthe postmenopausal years. Despite the knowledge thatestrogens have such important effects on skin, theircellular and molecular mechanisms of action are stillpoorly understood and their influence on pigmenta-tion is still far from clear.

Estrogens mediate their activity by interaction andactivation of specific intracellular receptor proteins, theestrogen receptors (ERs) � and �, which often coexist

as homo- or heterodimers. ER� and ER� are distinctproteins encoded by separate genes located on differ-ent chromosomes (110); they share �60% homology,bind 17�-estradiol with nearly equal affinity, and ex-hibit a similar binding profile for a large number ofnatural and synthetic ligands (111). ER� is expressed inboth male and female reproductive tissues, bone, thecardiovascular system, and regions of the brain (112).ER� is also expressed in both male and female repro-ductive tissues and in many nonreproductive tissuessuch as the lung, bladder, thymus, pituitary, hypothal-amus, heart, kidney, adrenals, and skin (113–115).Estrogens have significant effects on different cell typesimportant in skin physiology, including keratinocytes,fibroblasts, and melanocytes. It has been shown thatkeratinocyte mitotic activity increases in the epidermisof women in response to estrogens (116). Furthermore,the stimulation of proliferation and DNA synthesis ofhuman epidermal keratinocytes by estrogens has beendemonstrated in vitro (117–119), and it has been shownthat human keratinocytes in culture have high affinityestrogen binding sites (119). In addition to increasingthe proliferation of keratinocytes, estradiol acceleratesthe secretion of GM-CSF by �3-fold in cultured humankeratinocytes (120). GM-CSF is secreted by keratino-cytes at the wound edge to promote the migration ofendothelial cells and keratinocytes in order to advanceneovascularization and re-epithelialisation (121).Treatment with estrogen accelerates cutaneous woundhealing in both sexes by decreasing wound size and byincreasing collagen and fibronectin levels (122).

Dermal fibroblasts have an important modulatoryrole in remodeling the ECM during wound repair.

Figure 8. Mechanism(s) of estrogen action.Estrogens can activate both genomic intracellu-lar receptors (ER� and ER�) (classical,genomic pathway) and nongenomic cell sur-face receptors (nonclassical, nongenomic path-way). In the classical mode, the hormone entersthe cell by passive diffusion and interacts withits intracellular receptors. Upon ligation, thereceptor forms homo- or heterodimers; afterthat, the steroid receptor complex tightly asso-ciates with specific consensus DNA sequencescalled estrogen response elements (ERE),which consist of 15 base pair inverted palin-dromes located within the regulatory region ofthe target gene. The nonclassical pathwayworks faster and depends on the ability ofestrogen to interact with either membrane es-trogen receptors or nonsteroid hormone recep-tors (e.g., G-protein-coupled receptor, GPR30).The nonclassical pathway acts via different con-ventional second messengers like cAMP andactivates mitogen-activated protein kinases,which regulate transcription of specific genes.Based on and modified from M. J. Thornton(2005) Estrogen functions in skin and skinappendages. Expert Opin. Ther. Targets, vol. 9,pp. 617–629, and S. Verdier-Sevrain, F. Bonte,B. Gilchrest, (2006) Biology of estrogens inskin: implications for skin aging. Exp. Dermatol.,vol. 15, pp. 83–94.


Primary cultures of human dermal fibroblasts fromfemale skin have recently been shown to express bothER� and ER� (123).

Studies using double immunofluorescence staininghave shown that epidermal melanocytes in human scalpexpress both ER� and ER� in situ (124); ligand bindingstudies confirmed that cultured human normal epider-mal melanocytes contain estrogen receptors (107).Recently, Im et al. reported the presence of ER� innormal human melanocytes using immunocytochemis-try and RT-polymerase chain reaction (RT-PCR) (125).Examination of the effects of estrogen treatment onTYR activity has revealed a stimulation of this melano-genic enzyme (104, 105). It was recently demonstratedthat androgens modulate TYR activity via regulation ofcAMP, a key regulator of skin pigmentation (126). Thesum of these studies emphasizes the importance ofboth sex hormones in regulating skin pigmentation.

Details of the mechanisms by which sex hormonesinfluence melanogenesis remain to be elucidated, but thisis a good example of the concurrent involvement ofdifferent cellular factors (secondary messengers, growthfactors, etc.) in multiple molecular pathways within theskin eliciting hyperpigmentation as a final result.

Postinflammatory hyperpigmentation of the skin

Postinflammatory hyperpigmentation is manifested bydiscrete, hyperpigmented macules with hazy, featheredmargins, which may involve the epidermis and/ordermis. This usually develops after resolution of inflam-matory skin eruptions like acne, contact dermatitis, oratopic dermatitis. Postinflammatory hyperpigmenta-tion is more common in patients with darker skin and,at the cellular level, is characterized by a normalnumber of melanocytes that have increased melaninproduction (Table 1).

Arachidonate-derived chemical mediators, especiallyleukotrienes such as LTC4 and LTD4, and thrombox-anes such as TXB2 may be responsible for the inductionof postinflammatory hyperpigmentation of the skinbecause they can stimulate normal human melanocytesin vitro. These cells become swollen and more dendriticwith increased amounts of immunoreactive TYR whencultured for 2 days with LTC4, LTD4, or TXB2. Suchmorphological changes are thought to be required forthe transfer of melanosomes to surrounding keratino-cytes. Those effects were stronger than that elicited byPGE2, which, together with PGE1 and PGD2, are knownto be important endogenous regulators of inflamma-tory diseases in the skin and to stimulate mammalianpigment cells in vitro (127) and in vivo (128). Despite thecommon frequency of skin hyperpigmentation follow-ing inflammation, the mechanisms responsible for mel-anin synthesis have not yet been completely clarified,but some data have became available recently, as follows.

PGs are synthesized from arachidonic acid by cyclo-oxygenase and represent a group of potent lipid hor-mones that activate multiple signaling pathways, whichin turn regulate cellular growth, differentiation, and

apoptosis. In the skin, PGs (especially PGE2, PGF2�,and small quantities of prostacyclin) are produced(129) and rapidly released by keratinocytes after UV-R(130, 131). They are chronically present in inflamma-tory skin lesions and are involved in wound healing(132).

It has been shown that melanocytes express severaltypes of receptors for PGs; EP1-EP4 are receptors forPGE2 (EP3 and EP4 are high affinity whereas EP1 andEP2 are low affinity) (133); only EP1 and EP3 areexpressed by human melanocytes. The FP receptormediates the effects of PGF2� (134, 135) and is aheterotrimeric G-coupled protein receptor that signalsthrough Gq (136, 137). The FP receptor couples to Gqand activates phospholipase C-induced phosphoinosi-tide turnover, intracellular Ca2� mobilization, andMAPK/PKC activation (137). Scott et al. showed thatUV-R stimulates production of PGF2� by melanocytes,which in turn stimulates the activity and expression ofTYR, suggesting that PGF2� could act as an autocrinefactor for melanocyte differentiation (138).

On the other hand, PAR-2 is an important factorregulating skin pigmentation because its activation inkeratinocytes stimulates their uptake of melanosomesthrough phagocytosis. It has been reported that activa-tion of PAR-2 in keratinocytes stimulates the release ofPGE2 and PGF2�, which act as paracrine factors thatstimulate melanocyte dendricity (52). Melanocyte den-drite formation has been linked to the cAMP-depen-dent activation of Rac and the inhibition of Rho(139–141). However, recent studies demonstrated thatneither PGE2 nor PGF2� stimulates cAMP in melano-cytes, thus demonstrating that these PGs stimulatedendrite formation in a cAMP-independent manner(52). These data suggest that PAR-2 mediates cutane-ous pigmentation through regulation of melanosomeuptake and production of PGs, which act as paracrinefactors to stimulate melanocyte dendricity.

Secretory phospholipases comprise a large family ofCa2�-dependent enzymes that liberate arachidonicacid, which is also a precursor of lysophospholipids.The predominant secretory phospholipase (sPL) ex-pressed by keratinocytes is group X secretory phospho-lipase A2 (sPLA2-X), which liberates large amounts ofarachidonic acid and lysophospholipid lysophosphati-dylcholine (LPC) from membranes. They are releasedduring inflammation and their expression in the skin isincreased by UV-R (142, 143). Recent studies haveshown that low levels of sPLA2-X stimulate both TYRactivity and the dendricity of human melanocytes inculture. These cells have been shown to express thePLA2 receptor (PLA2R) and two G-protein-coupledreceptors for LPC (G2A and GPR119) (144). G2A andGPR119 couple to several G-proteins (including Gs)that regulate TYR activity and dendricity (141, 145).

In conclusion, recent studies help explain the mech-anisms involved in hyperpigmentation of the skin afterinflammation. All factors and pathways described aboveinteract within the skin; the final result is an increase ofTYR activity and melanocyte dendricity, which pro-


motes the production of melanin and its distribution tokeratinocytes.

Different factors responsible for increasing humanskin pigmentation do so via various intracellular path-ways, some of which are common and some distinct, asdetailed in this review. Table 2 summarizes some of theinternal or external stresses and the secondary messen-gers and effectors that are involved.

Given the complexity of skin and the pathways in-volved in regulating melanogenesis, one can assumethat stimulating or inhibiting more than one pathwayaffected by stress would lead to synergistic effects inincreasing or decreasing pigmentation. Shedding lighton the molecular mechanisms underlying hyperpig-mentation induced by internal or external factors couldbe applied to various ends (e.g., finding new technolo-gies/compounds) that could decrease pigmentation(in the case of melasma, senile lentigines, etc.), preventphotoaging (cosmetic industry), and/or designing tan-ning products with the potential to reduce the risk ofskin cancer (pharmaceutical industry).

CONCLUSIONS

There is no doubt that visual impressions of body formand color are important in interactions within andbetween human communities. Variation in human skincolor is clearly a multifactorial trait with a number ofmajor genetic determinants, several modifier genes,and environmental influences such as exposure toUV-R and gender effects. The pathological role ofpigmentation originates mainly from the fact that mel-anin pigments serve not only as the major determinantof skin color but also as the major source of skinprotection against UV-R, preventing sun-induced skindamage as well as skin cancer development. It is wellknown that repeated exposures to UV-R during anindividual’s lifetime are responsible not only for skinaging, but also for the appearance of skin cancer.Various types of skin cancers are responsible today for50% of all new cancer cases in the U.S. Melanoma,which accounts for � 85% of all skin cancer deathswithin the U.S., is the sixth most common type ofcancer in men and the seventh in women; over the past20 years the incidence of melanomas has more thandoubled.

Melanogenesis is a highly regulated process that is

modified by transcriptional, translational, and post-translational mechanisms. Melanin production resultsfrom strong cellular and molecular connections be-tween all cell populations in the skin, the key playersbeing fibroblasts, keratinocytes, and melanocytes. Mi-nor changes in the cellular physiology of the skin candramatically affect pigment production in positive ornegative manners.

The purpose of this review was to provide an overviewof the mechanisms by which external or internal factorsup-regulate melanin production in either transitory(such as in pregnancy) or permanent (such as aging)fashion. The action of UV-R has been extensivelyinvestigated, and there is a great amount of literatureavailable to explain its mechanism of action on the skinand its involvement in hyperpigmentation. Those stud-ies are the basis for research focused on understandingthe contribution of sun exposure to the appearance ofsolar lentigines and other age-related factors in thehyperpigmentation so often seen in older individuals.

The mechanisms of actions of other factors on theskin are less completely understood, such as thoseinvolving drugs or certain compounds in increasingmelanin production, but studies are ongoing in thisdirection, trying to provide a scientific rationale forsuch effects seen in different clinical treatments.

Melanocytes play a key role in tanning, responding toUV-R, and also to other factors secreted by keratino-cytes as a consequence of UV exposure. The identifica-tion of factors that modulate melanocyte functions couldresult in strategies to enhance photoprotection and con-sequently to decrease photoaging and photocarcinogen-esis. Therefore, understanding the mechanisms bywhich different factors and compounds induce mela-nogenesis is of great interest pharmaceutically (as ther-apy for pigmentary diseases) and cosmeceutically (e.g.,to design tanning products with the potential to reduceskin cancer risk).

This research was supported in part by the IntramuralResearch Program of the National Institutes of Health, Na-tional Cancer Institute. We thank Dr. Hong Hu (AvonProducts, Inc.) for assisting with photography and prepara-tion, and all persons who agreed in writing to have theirphotos taken and used in this review (among them, AbenaaBrew, Sheri Lenc, and Cielo Rivera, Avon Products, Inc.). Wethank Sheri Lenc (Avon Products, Inc.) for assisting inmanuscript preparation. We appreciate the help of Drs. DavidBrown, Uma Santhanam, and Bhavani Balasubramanian

TABLE 2. Summary of different external and internal stresses increasing human skin pigmentation and their intracellular secondarymessengers and effectors

Stress Secondary messenger Secondary effector Figure

UV induces the production of:NO cGMP PKG 5ET-1 DAG PKC 7�-MSH, ACTH, PGE2 cAMP PKA 5

Hormones (nonclassical pathway) cAMP MAPK 8Inflammation Inositol 1,4,5 triphosphate MAPK/PKC –


(Avon Products, Inc.) and we also thank Sergio Coelho(NIH/NCI) for providing clinical pictures. We thank Drs.Janice Teal, Xiaochun Luo, and Sherrie Tafuri (Avon Prod-ucts, Inc.) for critically reading the manuscript. We also thankDrs. Gopinathan Menon (Avon Products, Inc.) and StancaBirlea (University of Colorado Health Sciences Center) forhelpful discussions and advice.

REFERENCES

1. Haake, A., and Holbrook, K. (1999) The structure and devel-opment of skin. In Fitzpatrick’s Dermatology in General Medicine(Freedberg, I. M., Eisen, A. Z., Wolff, K., Austen, K. F.,Goldsmith, L. A., Katz, S. I., and Fitzpatrick, T. B., eds) pp.70–114, McGraw-Hill, New York

2. Pathak, M. A. (1995) Functions of melanin and protection bymelanin. In Melanin: Its Role in Human Photoprotection (Zeise, L.,Chedekel, M. R., and Fitzpatrick, T. B., eds) pp. 125–134,Valdemar Publishing Company, Overland Park

3. Tobin, D. J. (2006) Biochemistry of the skin—our brain on theoutside. Chem. Soc. Rev. 35, 52–67

4. Elias, P. M. (2005) Stratum corneum defensive functions: anintegrated view. J. Invest. Dermatol. 125, 183–200

5. Cotsarelis, G., Sun, T. T., and Lavker, R. M. (1990) Label-retaining cells reside in the bulge area of pilosebaceous unit:implications for follicular stem cells, hair cycle, and skincarcinogenesis. Cell 61, 1329–1337

6. Ito, M., Liu, Y., Yang, Z., Nguyen, J., Liang, F., Morris, R. J., andCotsarelis, G. (2005) Stem cells in the hair follicle bulgecontribute to wound repair but not to homeostasis of theepidermis. Nat. Med. 11, 1351–1354

7. Hearing, V. J. (1997) The regulation of melanin production. InDrug Discovery Approaches for Developing Cosmeceuticals, AdvancedSkin Care and Cosmetic Products (Hori, W., ed) pp. 3.1.1–3.1.21,IBC Library Series, Southborough, Massachusetts

8. Fitzpatrick, T. B., and Breathnach, A. S. (1963) The epidermalmelanin unit system. Dermatol. Wochenschr. 147, 481–489

9. Boissy, R. E., and Nordlund, J. J. (1997) Molecular basis ofcongenital hypopigmentary disorders in humans: a review.Pigment Cell Res. 10, 12–24

10. Bolognia, J. (1999) Molecular advances in disorders of pigmen-tation. Adv. Dermatol. 15, 341–365

11. Seiji, M., Shimao, K., Birbeck, M. S., and Fitzpatrick, T. B.(1963) Subcellular localization of melanin biosynthesis. Ann.N. Y. Acad. Sci. 100, 497–533

12. Kushimoto, T., Basrur, V., Valencia, J., Matsunaga, J., Vieira,W. D., Ferrans, V. J., Muller, J., Appella, E., and Hearing, V. J.(2001) A model for melanosome biogenesis based on thepurification and analysis of early melanosomes. Proc. Natl.Acad. Sci. U. S. A. 98, 10698–10703

13. Berson, J. F., Harper, D. C., Tenza, D., Raposo, G., and Marks,M. S. (2001) Pmel17 initiates premelanosome morphogenesiswithin multivesicular bodies. Mol. Biol. Cell 12, 3451–3464

14. Basrur, V., Yang, F., Kushimoto, T., Higashimoto, Y., Yasumoto,K., Valencia, J., Muller, J., Vieira, W. D., Watabe, H., Shabanow-itz, J., et al. (2003) Proteomic analysis of early melanosomes:identification of novel melanosomal proteins. J. Proteome. Res.2, 69–79

15. Hearing, V. J., and Tsukamoto, K. (1991) Enzymatic control ofpigmentation in mammals. FASEB J. 5, 2902–2909

16. Halaban, R., Cheng, E., Zhang, Y., Moellmann, G., Hanlon, D.,Michalak, M., Setaluri, V., and Hebert, D. N. (1997) Aberrantretention of tyrosinase in the endoplasmic reticulum mediatesaccelerated degradation of the enzyme and contributes to thededifferentiated phenotype of amelanotic melanoma cells.Proc. Natl. Acad. Sci. U. S. A. 94, 6210–6215

17. Branza-Nichita, N., Petrescu, A. J., Dwek, R. A., Wormald,M. R., Platt, F. M., and Petrescu, S. M. (1999) Tyrosinasefolding and copper loading in vivo: a crucial role for calnexinand alpha-glucosidase II. Biochem. Biophys. Res. Commun. 261,720–725

18. Branza-Nichita, N., Negroiu, G., Petrescu, A. J., Garman, E. F.,Platt, F. M., Wormald, M. R., Dwek, R. A., and Petrescu, S. M.(2000) Mutations at critical N-glycosylation sites reduce tyrosi-

nase activity by altering folding and quality control. J. Biol.Chem. 275, 8169–8175

19. Toyofuku, K., Wada, I., Valencia, J. C., Kushimoto, T., Ferrans,V. J., and Hearing, V. J. (2001) Oculocutaneous albinism types1 and 3 are ER retention diseases: mutation of tyrosinase orTyrp1 can affect the processing of both mutant and wild-typeproteins. FASEB J. 15, 2149–2161

20. Urabe, K., Aroca, P., Tsukamoto, K., Mascagna, D., Palumbo,A., Prota, G., and Hearing, V. J. (1994) The inherent cytotox-icity of melanin precursors: a revision. Biochim. Biophys. Acta1221, 272–278

21. Prota, G. (1992) Melanins and Melanogenesis, pp. 1–290, Aca-demic, New York

22. Ito, S., Wakamatsu, K., and Ozeki, H. (2000) Chemical analysisof melanins and its application to the study of the regulation ofmelanogenesis. Pigment Cell Res. 13 Suppl. 8, 103–109

23. Nordlund, J. J. (1985) The pigmentary system. New interpre-tation of old data. J. Dermatol. 12, 105–116

24. Zecca, L., Shima, T., Stroppolo, A., Goj, C., Battiston, G. A.,Gerbasi, R., Sarna, T., and Swartz, H. M. (1996) Interaction ofneuromelanin and iron in substantia nigra and other areas ofhuman brain. Neuroscience 73, 407–415

25. Hearing, V. J. (2000) The melanosome: the perfect model forcellular response to the environment. Pigment Cell Res. 13Suppl. 8, 23–34

26. Fitzpatrick, T. B., and Szabo, G. (1959) The melanocyte:cytology and cytochemistry. J. Invest. Dermatol. 32, 197–209

27. Sturm, R. A., Box, N. F., and Ramsay, M. (1998) Humanpigmentation genetics: the difference is only skin deep. Bioes-says 20, 712–721

28. Halaban, R., Pomerantz, S. H., Marshall, S., Lambert, D. T.,and Lerner, A. B. (1983) Regulation of tyrosinase in humanmelanocytes grown in culture. J. Biol. Chem. 97, 480–488

29. Toda, K., Pathak, M. A., Parrish, J. A., Fitzpatrick, T. B., andQuevedo, W. C., Jr. (1972) Alterations of racial differences inmelanosome distribution in human epidermis after exposureto ultraviolet light. Nat. New. Biol. 236, 143–145

30. Jimbow, K., Quevedo, W. C., Jr., Fitzpatrick, T. B., and Szabo,G. (1976) Some aspects of melanin biology: 1950–1975. J. In-vest. Dermatol. 67, 72–89

31. Kobayashi, N., Nakagawa, A., Muramatsu, T., Yamashina, Y.,Shirai, T., Hashimoto, M. W., Yshigaki, Y., Ohnishi, T., andMori, T. (1998) Supranuclear melanin caps reduce ultravioletinduced DNA photoproducts in human epidermis. J. Invest.Dermatol. 110, 806–810

32. Chakraborty, A. K., Funasaka, Y., Slominski, A., Ermak, G.,Hwang, J., Pawelek, J. M., and Ichihashi, M. (1996) Productionand release of proopiomelanocortin (POMC) derived peptidesby human melanocytes and keratinocytes in culture: regulationby ultraviolet B. Biochim. Biophys. Acta 1313, 130–138

33. Funasaka, Y., Chakraborty, A. K., Hayashi, Y., Komoto, M.,Ohashi, A., Nagahama, M., Inoue, Y., Pawelek, J., and Ichi-hashi, M. (1998) Modulation of melanocyte-stimulating hor-mone receptor expression on normal human melanocytes:evidence for a regulatory role of ultraviolet B, interleukin-1alpha, interleukin-1 beta, endothelin-1 and tumor necrosisfactor-alpha. Br. J. Dermatol. 139, 216–224

34. Tada, A., Suzuki, I., Im, S., Davis, M. B., Cornelius, J., Babcock,G., Nordlund, J. J., and Abdel-Malek, Z. A. (1998) Endothelin-1is a paracrine growth factor that modulates melanogenesis onhuman melanocytes and participates in their responses toultraviolet radiation. Cell Growth Differ. 9, 575–584

35. Abdel-Malek, Z., Scott, M. C., Suzuki, I., Tada, A., Im, S.,Lamoreux, L., Ito, S., Barsh, G., and Hearing, V. J. (2000) Themelanocortin-1 receptor is a key regulator of human cutaneouspigmentation. Pigment Cell Res. 13 Suppl. 8, 156–162

36. Hirobe, T. (2005) Role of keratinocyte-derived factors involvedin regulating the proliferation and differentiation of mamma-lian epidermal melanocytes. Pigment Cell Res. 18, 2–12

37. Slominski, A., Szczesniewski, A., and Wortsman, J. (2000)Liquid chromatography-mass spectrometry detection of corti-cotropin-releasing hormone and proopiomelanocortin-de-rived peptides in human skin. J. Clin. Endocrinol. Metab. 85,3582–3588

38. Wakamatsu, K., Graham, A., Cook, D., and Thody, A. J. (1997)Characterization of ACTH peptides in human skin and their


activation of the melanocortin-1 receptor. Pigment Cell Res. 10,288–297

39. Cone, R. D., Lu, D., Koppula, S., Vage, D. I., Klungland, H.,Boston, B., Chen, W., Orth, D. N., Pouton, C., and Kesterson,R. A. (1996) The melanocortin receptors: agonists, antago-nists, and the hormonal control of pigmentation. Recent Prog.Horm. Res. 51, 287–317

40. Im, S., Moro, O., Peng, F., Medrano, E. E., Cornelius, J.,Babcock, G., Nordlund, J. J., and Abdel-Malek, Z. A. (1998)Activation of cyclic AMP pathway by alpha-melanotropin me-diates the response of human melanocytes to ultraviolet Bradiation. Cancer Res. 58, 47–54

41. Insel, P. A., Bourne, H. R., Coffino, P., and Tomkins, G. M.(1975) Cyclic AMP-dependent protein kinase: pivotal role inregulation of enzyme induction and growth. Science 190, 896–898

42. Busca, R., and Ballotti, R. (2000) Cyclic AMP a key messengerin the regulation of skin pigmentation. Pigment Cell Res. 13,60–69

43. Tachibana, M. (2000) MITF: a stream flowing for pigment cell.Pigment Cell Res. 13, 230–240

44. Yaar, M., Grossman, K., Eller, M., and Gilchrest, B. A. (1991)Evidence for nerve growth factor-mediated paracrine effects inhuman epidermis. J. Cell Biol. 115, 821–828

45. Peacocke, M., Yaar, M., Mansur, C. P., Chao, M. V., andGilchrest, B. A. (1988) Induction of nerve growth factorreceptors on cultured human melanocytes. Proc. Natl. Acad. Sci.U. S. A. 85, 5282–5286

46. Imokawa, G., Yada, Y., and Miyagishim, M. (1992) Endothelinssecreted from human keratinocytes are intrinsic mitogens forhuman melanocytes. J. Biol. Chem. 267, 24675–24680

47. Yohn, J. J., Morelli, J. G., Walchak, S. J., Rundell, K. B., Norris,D. A., and Zamora, M. R. (1993) Cultured human keratino-cytes synthesize and secrete endothelin-1. J. Invest. Dermatol.100, 23–26

48. Hara, M., Yaar, M., and Gilchrest, B. A. (1995) Endothelin-1 ofkeratinocyte origin is a mediator of melanocyte dendricity.J. Invest. Dermatol. 105, 744–748

49. Imokawa, G., Yada, Y., and Kimura, M. (1996) Signalingmechanisms of endothelin-induced mitogenesis and melano-genesis in human melanocytes. Biochem. J. 314, 305–312

50. Swope, V. B., Medrano, E. E., Smalara, D., and Abdel-Malek,Z. A. (1995) Long-term proliferation of human melanocytes issupported by the physiologic mitogens alpha-melanotropin,endothelin-1, and basic fibroblast growth factor. Exp. Cell Res.217, 453–459

51. Imokawa, G., Miyagishi, M., and Yada, Y. (1995) Endothelin-1as a new melanogen: coordinated expression of its gene andthe tyrosinase gene in UVB-exposed human epidermis. J. In-vest. Dermatol. 105, 32–37

52. Scott, G., Leopardi, S., Printup, S., Malhi, M., Seiberg, M., andLapoint, R. (2004) Proteinase-activated receptor-2 stimulatesprostaglandin production in keratinocytes: analysis of prosta-glandin receptors on human melanocytes and effects on PGE2and PGF2alpha on melanocyte dendricity. J. Invest. Dermatol.122, 1214–1224

53. Halaban, R., Langdon, R., Birchall, N., Cuono, C., Baird, A.,Scott, G., Moellmann, G., and McGuire, J. (1988) Basic fibro-blast growth factor from human keratinocytes is a naturalmitogen for melanocytes. J. Cell Biol. 107, 1611–1619

54. Hachiya, A., Kobayashi, A., Ohuchi, A., Takema, Y., andImokawa, G. (2001) The paracrine role of stem cell factor/c-kitsignaling in the activation of human melanocytes in ultraviolet-B-induced pigmentation. J. Invest. Dermatol. 116, 578–586

55. Schauer, E., Trautinger, F., Kock, A., Schwarz, A., Bhardwaj, R.,Simon, M., Ansel, J. C., Schwarz, T., and Luger, T. A. (1994)Proopiomelanocortin-derived peptides are synthesized andreleased by human keratinocytes. J. Clin. Invest. 193, 2258–2262

56. Imokawa, G., Yada, Y., Kimura, M., and Morisaki, N. (1996)Granulocyte/macrophage colony-stimulating factor is an in-trinsic keratinocyte-derived growth factor for human melano-cytes in UVA-induced melanosis. Biochem. J. 313, 625–631

57. Bottaro, D. P., Rubin, J. S., Faletto, D. L., Chan, A. M., Kmiecik,T. E., Vande Wounde, G. F., and Aaronson, S. A. (1991)Identification of the hepatocyte growth factor receptor as thec-met proto-oncogene product. Science 251, 802–804

58. Halaban, R., Tyrrell, L., Longley, J., Yarden, Y., and Rubin, J.(1993) Pigmentation and proliferation of human melanocytesand the effects of melanocyte-stimulating hormone and ultra-violet B light. Ann. N. Y. Acad. Sci. 680, 290–301

59. Matsumoto, K., Tajima, H., and Nakamura, T. (1991) Hepato-cyte growth factor is a potent stimulator of human melanocyteDNA synthesis and growth. Biochem. Biophys. Res. Commun. 176,45–51

60. Chiba, S., Shibuya, K., Miyazono, K., Tojo, A., Oka, Y., Miya-gawa, K., and Takaku, F. (1990) Affinity purification of humangranulocyte macrophage colony-stimulating factor receptoralpha-chain. J. Biol. Chem. 265, 19777–19781

61. Mui, A. L., Wakao, H., O’Farrell, A. M., Harada, N., andMiyajima, A. (1995) Interleukin-3, granulocyte-macrophagecolony stimulating factor and interleukin-5 transduce signalsthrough two STAT5 homologs. EMBO J. 14, 1166–1175

62. Wang, Y., Morrella, K. K., Ripperger, J., Lai, C. F., Gearing,D. P., Fey, G. H., Campos, S. P., and Baumann, H. (1995)Receptors for interleukin-3 (IL-3) and growth hormone medi-ate an IL-6-type transcriptional induction in the presence ofJAK2 or STAT3. Blood 86, 1671–1679

63. Okuda, K., Sanghera, J. S., Pelech, S. L., Kanakura, Y., Hallek,M., Griffin, J. D., and Druker, B. J. (1992) Granulocyte-macrophage colony-stimulating factor, interleukin-3, and steelfactor induce rapid tyrosine phosphorylation of p42 and p44MAP kinase. Blood 79, 2880–2887

64. Robins, A. H. (1991) Biological Perspectives on Human Pigmenta-tion, pp. 1–253, Cambridge Univ. Press, Cambridge, UK

65. Wang, S. Q., Setlow, R., Berwick, M., Polsky, D., Marghoob,A. A., Kopf, A. W., and Bart, R. S. (2001) Ultraviolet A andmelanoma: a review. J. Am. Acad. Dermatol. 44, 767–774

66. Garland, C. F., Garland, F. C., and Gorham, E. D. (1993) Risingtrends in melanoma: an hypothesis concerning sunscreeneffectiveness. Ann. Epidemiol. 3, 103–110

67. Stern, R. S., Nichols, K. T., and Vakeva, L. H. (1997) Malignantmelanoma in patients treated for psoriasis with methoxsalen(psoralen) and ultraviolet A radiation (PUVA). The PUVAFollow-Up Study. N. Engl. J. Med. 336, 1041–1045

68. Lindelof, B., Sigurgeirsson, B., Tegner, E., Larko, O., Johan-nesson, A., Berne, B., Ljunggren, B., Andersson, T., Molin, L.,Nylander-Lundqvist, E., and Emtestam, L. (1999) PUVA andcancer risk: the Swedish follow-up study. Br. J. Dermatol. 141,108–112

69. Stern, R. S. (2001) The risk of melanoma in association withlong-term exposure to PUVA. J. Am. Acad. Dermatol. 44, 755–761

70. Garland, C. F., Garland, F. C., and Gorham, E. D. (2003)Epidemiologic evidence for different roles of ultraviolet A andB radiation in melanoma mortality rates. Ann. Epidemiol. 13,395–404

71. Tadokoro, T., Kobayashi, N., Smudzka, B. Z., Ito, S., Waka-matsu, K., Yamaguchi, Y., Korossy, K. S., Miller, S. A., Beer, J. Z.,and Hearing, V. J. (2003) UV-induced DNA damage andmelanin content in human skin differing in racial/ethnicorigin. FASEB J. 17, 1177–1179

72. Gilchrest, B. A., Park, H. Y., Eller, M. S., and Yaar, M. (1996)Mechanisms of ultraviolet light-induced pigmentation. Photo-chem. Photobiol. 63, 1–10

73. Kochevar, I. E. (1995) Molecular and cellular effects of UVradiation relevant to chronic photodamage. In Photodamage(Gilchrest, B. A., ed) pp. 51–67, Blackwell Science, Cambridge,Massachusetts

74. Sies, H., and Stahl, W. (2004) Nutritional protection againstskin damage from sunlight. Annu. Rev. Nutr. 24, 173–200

75. Nishizuka, Y. (1986) Studies and perspectives of protein kinaseC. Science 233, 305–312

76. Gordon, P. R., and Gilchrest, B. A. (1989) Human melanogen-esis is stimulated by diacylglycerol. J. Invest. Dermatol. 93,700–702

77. Friedmann, P. S., Wren, F. E., and Matthews, J. N. (1990)Ultraviolet stimulated melanogenesis by human melanocytes isaugmented by di-acyl glycerol but not TPA. J. Cell Physiol. 142,334–341

78. Pathak, M. A., and Fitzpatrick, T. B. (1993) Preventive treat-ment of sunburn, dermatohekiosis and skin cancer with sunprotective agents. In Dermatology in General Medicine (Fitz-


patrick, T. B., Eisen, A. Z., Wolff, K., Feedberg, I. M., Austin,and K. F., eds) pp. 1689–1717, McGraw-Hill, New York

79. Pathak, M. A., and Stratton, K. (1968) Free radicals in humanskin before and after exposure to light. Arch. Biochem. Biophys.123, 468–476

80. Bolognia, J. (1995) Aging skin. Am. J. Med. 98, 99S–103S81. Bolognia, J. L. (1993) Dermatologic and cosmetic concerns of

the older woman. Clin. Geriatr. Med. 9, 209–22982. Gilchrest, B. A., Frederick, B. B., and Szabo, G. (1979) Effects

of aging and chronic sun exposure on melanocytes in humanskin. J. Invest. Dermatol. 73, 141–143

83. Commo, S., Gaillard, O., Thibaut, S., and Bernard, B. A.(2004) Absence of TRP-2 in melanogenic melanocytes ofhuman hair. Pigment Cell Res. 17, 488–497

84. Ortonne, J. P., Bahadoran, P., Fitzpatrick, T. B., Mosher, D. B.,and Hory, Y. (2003) Hypomelanoses and hypermelanoses. InFitzpatrick’s Dermatology in General Medicine (Freedberg, I. M.,Eisen, A. Z., Wolff, K., Austen, K. F., Goldsmith, L. A., and Katz,S. I., eds) pp. 836–881, McGraw-Hill Medical PublishingDivision, New York

85. Kadono, S., Manaka, I., Kawashima, M., Kobayashi, T., andImokawa, G. (2001) The role of the epidermal endothelincascade in the hyperpigmentation mechanism of lentigo seni-lis. J. Invest. Dermatol. 116, 571–577

86. Imokawa, G., Kobayasi, T., and Miyagishi, M. (2000) Intracel-lular signaling mechanisms leading to synergistic effects ofendothelin-1 and stem cell factor on proliferation of culturedhuman melanocytes. Cross-talk via trans-activation of the ty-rosine kinase c-kit receptor. J. Biol. Chem. 275, 33321–33328

87. Hattori, H., Kawashima, M., Ichikawa, Y., and Imokawa, G.(2004) The epidermal stem cell factor is over-expressed inlentigo senilis: implication for the mechanism of hyperpigmen-tation. J. Invest. Dermatol. 122, 1256–1265

88. Blume-Jensen, P., Claesson-Welsh, L., Siegbahn, A., Zsebo,K. M., Westermark, B., and Heldin, C. H. (1991) Activation ofthe human c-kit product by ligand-induced dimerization me-diates circular actin reorganization and chemotaxis. EMBO J.10, 4121–4128

89. Lennartsson, J., Blume-Jensen, P., Hermanson, M., Ponten, E.,Carlberg, M., and Ronnstrand, L. (1999) Phosphorylation ofShc by Src family kinases is necessary for stem cell factorreceptor/c-kit mediated activation of the Ras/MAP kinasepathway and c-fos induction. Oncogene 18, 5546–5553

90. Imokawa, G. (2004) Autocrine and paracrine regulation ofmelanocytes in human skin and in pigmentary disorders.Pigment Cell Res. 17, 96–110

91. Imokawa, G., Yada, Y., Morisaki, N., and Kimura, M. (1998)Biological characterization of human fibroblast-derived mito-genic factors for human melanocytes. Biochem. J. 330, 1235–1239

92. Unver, N., Freyschmidt-Paul, P., Horster, S., Wenck, H.,Stab, F., Blatt, T., and Elsasser, H. P. (2006) Alterations inthe epidermal-dermal melanin axis and factor XIIIa mela-nophages in senile lentigo and ageing skin. Br. J. Dermatol.155, 119 –128

93. Yamaguchi, Y., Itami, S., Watabe, H., Yasumoto, K., Abdel-Malek, Z. A., Kubo, T., Rouzaud, F., Tanemura, A., Yoshikawa,K., and Hearing, V. J. (2004) Mesenchymal-epithelial interac-tions in the skin: increased expression of dickkopf1 by palmo-plantar fibroblasts inhibits melanocyte growth and differentia-tion. J. Cell Biol. 165, 275–285

94. Goh, C. L., and Dlova, C. N. (1999) A retrospective study onthe clinical presentation and treatment outcome of melasma ina tertiary dermatological referral centre in Singapore. SingaporeMed. J. 40, 455–458

95. Levantine, A., and Almeyda, J. (1973) Drug induced changesin pigmentation. Br. J. Dermatol. 89, 105–112

96. Robins, A. H. (1972) Skin melanin concentrations in schizo-phrenia. Br. J. Psychiatry 121, 613–617

97. Levy, H. (1982) Chloroquine-induced pigmentation. S. Afr.Med. J. 62, 735–737

98. Molokhia, M. M., and Portnoy, B. (1973) Trace elements andskin pigmentation. Br. J. Dermatol. 89, 207–209

99. Barankin, B., Silver, S. G., and Carruthers, A. (2002) The skinin pregnancy. J. Cutan. Med. Surg. 6, 236–240

100. Kang, W. H., Yoon, K. H., Lee, E. S., Kim, J., Lee, K. B., Yim,H., Sohn, S., and Im, S. (2002) Melasma: histopathological

characteristics in 56 Korean patients. Br. J. Dermatol. 146,228 –237

101. Grimes, P. E., Yamada, N., and Bhawan, L. (2005) Lightmicroscopic, immunohistochemical, and ultrastructural alter-ations in patients with melasma. Am. J. Dermatopathol. 27,96–101

102. Smith, A. G., Shuster, S., Thody, A. J., and Peberdy, M. (1977)Chloasma, oral contraceptives, and plasma immunoreactivebeta-melanocyte-stimulating hormone. J. Invest. Dermatol. 68,169–170

103. Parker, F. (1981) Skin and hormones. In Textbook of Endocrinol-ogy (Williams, R. H., ed) pp. 1088–1091, W. B. Saunders Co.,Philadelphia, Pennsylvania

104. Ranson, M., Posen, S., and Mason, R. S. (1988) Humanmelanocytes as a target tissue for hormones: in vitro studieswith 1 alpha-25, dihydroxyvitamin D3, alpha-melanocyte stim-ulating hormone, and beta-estradiol. J. Invest. Dermatol. 91,593–598

105. Kippenberger, S., Loitsch, S., Solano, F., Bernd, A., andKaufmann, R. (1998) Quantification of tyrosinase, TRP-1, andTRP-2 transcripts in human melanocytes by reverse tran-scriptase-competitive multiplex PCR-regulation by steroid hor-mones. J. Invest. Dermatol. 110, 364–367

106. McLeod, S. D., Ranson, M., and Mason, R. S. (1994) Effects ofestrogens in human melanocytes in vitro. J. Steroid Biochem. Mol.Biol. 49, 9–14

107. Jee, S. H., Lee, S. Y., Chiu, H. C., Chang, C. C., and Chen, T. J.(1994) Effects of estrogen and estrogen receptor in normalhuman melanocytes. Biochem. Biophys. Res. Commun. 199, 1407–1412

108. Verdier-Sevrain, S., Bonte, F., and Gilchrest, B. (2006) Biologyof estrogens in skin: implications for skin aging. Exp. Dermatol.15, 83–94

109. Thornton, M. J. (2005) Oestrogen functions in skin and skinappendages. Expert. Opin. Ther. Targets 9, 617–629

110. Enmark, E., Pelto-Huikko, M., Grandien, K., Lagercrantz, S.,Lagercrantz, J., Fried, G., Nordenskjold, M., and Gustafsson,J. A. (1997) Human estrogen receptor beta-gene structure,chromosomal localization, and expression pattern. J. Clin.Endocrinol. Metab. 82, 4258–4265

111. Kuiper, G. G., Carlsson, B., Grandien, K., Enmark, E., Hagg-blad, J., Nilsson, S., and Gustafsson, J. A. (1997) Comparison ofthe ligand binding specificity and transcript tissue distributionof estrogen receptors alpha and beta. Endocrinology 138, 863–870

112. Couse, J. F., Lindzey, J., Grabduebm, K., Gustafsson, J. A., andKorach, K. S. (1997) Tissue distribution and quantitativeanalysis of estrogen receptor-alpha (ERalpha) and estrogenreceptor-beta (ERbeta) messenger ribonucleic acid in thewild-type ERalpha-knockout mouse. Endocrinology 138, 4613–4621

113. Taylor, A. H., and Al-Azzawi, F. (2000) Immunolocalisation ofoestrogen receptor beta in human tissues. J. Mol. Endocrinol. 24,145–155

114. Thornton, M. J., Taylor, A. H., Mulligan, K., Al-Azzawi, F.,Lyon, C. C., O’Driscoll, J., and Messenger, A. G. (2003) Thedistribution of estrogen receptor beta is distinct to that ofestrogen receptor alpha and the androgen receptor in humanskin and the pilosebaceous unit. J. Investig. Dermatol. Symp. Proc.8, 100–103

115. Pelletier, G., and Ren, L. (2004) Localization of sex steroidreceptors in human skin. Histol. Histopathol. 19, 629–636

116. Punnonen, R. (1972) Effect of castration and peroral estrogentherapy on the skin. Acta Obstet. Gynecol. Scand. Suppl. 21, 3–44

117. Urano, R., Sakabe, K., Seiki, K., and Ohkido, M. (1995) Femalesex hormone stimulates cultured human keratinocyte prolifer-ation and its RNA- and protein-synthetic activities. J. Dermatol.Sci. 9, 176–184

118. Kanda, N., and Watanabe, S. (2004) 17beta-Estradiol stimu-lates the growth of human keratinocytes by inducing cyclin D2expression. J. Invest. Dermatol. 123, 319–328

119. Verdier-Sevrain, S., Yaar, M., Cantatore, J., Traish, A., andGilchrest, B. A. (2004) Estradiol induces proliferation ofkeratinocytes via a receptor mediated mechanism. FASEB J. 18,1252–1254

120. Kanda, N., and Watanabe, S. (2004) 17beta-Estradiol en-hances the production of granulocyte-macrophage colony-


stimulating factor in human keratinocytes. J. Invest. Dermatol.123, 329 –337

121. Mann, A., Breuhahn, K., Schirmacher, P., and Blessing, M.(2001) Keratinocyte-derived granulocyte macrophage colonystimulating factor accelerates wound healing: stimulation ofkeratinocyte proliferation, granulation tissue formation, andvascularization. J. Invest. Dermatol. 117, 1382–1390

122. Ashcroft, G. S., Greenwell-Wild, T., Horan, M. A., Wahl, S. M.,and Ferguson, M. W. (1999) Topical estrogen acceleratescutaneous wound healing in aged humans associated with analtered inflammatory response. Am. J. Pathol. 155, 1137–1146

123. Haczynski, J., Tarkowski, R., Jarzabek, K., Slomczynska, M.,Wolczynski, S., Magoffin, D. A., Jakowicki, J. A., and Jakimiuk,A. J. (2002) Human cultured skin fibroblasts express estrogenreceptor alpha and beta. Int. J. Mol. Med. 10, 149–153

124. Tobin, D. J., Taylor, A. H., Messenger, A. G., and Thornton,M. J. (2002) Melanocytes in human scalp epidermis and hairfollicles express the androgen receptor (AR) and both estro-gen receptors (ERalpha) and (ERbeta). 9th Annual EuropeanHair Research Society Conference (poster presentation)

125. Im, S., Lee, E. S., Kim, W., On, W., Kim, J., Lee, M., and Kang,W. H. (2002) Donor specific response of estrogen and proges-terone on cultured human melanocytes. J. Korean Med. Sci. 17,58–64

126. Tadokoro, T., Rouzaud, F., Itami, S., Hearing, V. J., andYoshikawa, K. (2003) The inhibitory effect of androgen andsex-hormone-binding globulin on the intracellular cAMP leveland tyrosinase activity of normal human melanocytes. PigmentCell Res. 16, 190–197

127. Tomita, Y., Iwamoto, M., Masuda, T., and Tagami, H. (1987)Stimulatory effect of prostaglandin E2 on the configuration ofnormal human melanocytes in vitro. J. Invest. Dermatol. 89,299–301

128. Nordlund, J. J., Collins, C. E., and Rheins, L. A. (1986)Prostaglandin E2 and D2 but not MSH stimulate the prolifer-ation of pigment cells in the pinnal epidermis of the DBA/2mouse. J. Invest. Dermatol. 86, 433–437

129. Pentland, A. P., and Mahoney, M. G. (1990) Keratinocyteprostaglandin synthesis is enhanced by IL-1. J. Invest. Dermatol.94, 43–46

130. Hanson, D., and DeLeo, V. (1990) Long-wave ultraviolet lightinduces phospholipase activation in cultured human epider-mal keratinocytes. J. Invest. Dermatol. 95, 158–163

131. Pentland, A. P., Mahoney, M., Jacobs, S. C., and Holtzman,M. J. (1990) Enhanced prostaglandin synthesis after ultravioletinjury is mediated by endogenous histamine stimulation. Amechanism for irradiation erythema. J. Clin. Invest. 86, 566–574

132. Pentland, A. P., George, J., Moran, C., and Needleman, P.(1987) Cellular confluence determines injury-induced prosta-glandin E2 synthesis by human keratinocyte cultures. Biochim.Biophys. Acta 919, 71–78

133. Coleman, R. A., Eglen, R. M., Jones, R. L., Narumiya, S.,Shimizu, T., Smith, W. L., Dahlen, S. E., Drazen, J. M.,Gardiner, P. J., Jackson, W. T., et al. (1995) Prostanoid andleukotriene receptors: a progress report from the IUPHARworking parties on classification and nomenclature. Adv. Pros-taglandin Thromboxane Leukot. Res. 23, 283–285

134. Abramovitz, M., Boie, Y., Nguyen, T., Rushmore, T. H., Bayne,M. A., Metters, K. M., Slipetz, D. M., and Grygorczyk, R. (1994)Cloning and expression of a cDNA for the human prostanoidFP receptor. J. Biol. Chem. 269, 2632–2636

135. Pierce, K. L., Fujino, H., Srinivasan, D., and Regan, J. W.(1999) Activation of FP prostanoid receptor isoforms leads toRho-mediated changes in cell morphology and in the cellcytoskeleton. J. Biol. Chem. 274, 35944–35949

136. Carrasco, M. P., Phaneuf, S., Asboth, G., and Lopez Bernal, A.(1996) Fluprostenol activates phospholipase C and Ca2� mo-bilization in human myometrial cells, J. Clin. Endocrinol. Metab.81, 2104–2110

137. Breyer, M. D., Jacobson, H. R., and Breyer, R. M. (1996)Functional and molecular aspects of renal prostaglandin re-ceptors. J. Am. Soc. Nephrol. 7, 8–17

138. Scott, G., Jacobs, S., Leopardi, S., Anthony, F. A., Learn, D.,Malaviya, R., and Pentland, A. (2005) Effects of PGF2alpha onhuman melanocytes and regulation of the FP receptor byultraviolet radiation. Exp. Cell Res. 304, 407–416

139. Busca, R., Bertolotto, C., Abbe, P., Englaro, W., Ishizaki, T.,Narumiya, S., Boquet, P., Ortonne, J. P., and Ballotti, R. (1998)Inhibition of Rho is required for cAMP induced melanoma celldifferentiation. Mol. Biol. Cell 9, 1367–1378

140. Scott, G. (2002) Rac and rho: the story behind melanocytedendrite formation. Pigment Cell Res. 15, 322–330

141. Scott, G., and Leopardi, S. (2003) The cAMP signaling pathwayhas opposing effects on Rac and Rho in B16F10 cells: implica-tions for dendrite formation in melanocytic cells. Pigment CellRes. 16, 139–148

142. Li-Stiles, B., Lo, H. H., and Fischer, S. M. (1998) Identificationand characterization of several forms of phospholipase A2 inmouse epidermal keratinocytes. J. Lipid Res. 39, 569–582

143. Schadow, A., Scholz-Pedretti, K., Lambeau, G., Gelb, M. H.,Furstenberger, G., Pfeilschifter, J., and Kaszkin, M. (2001)Characterization of group X phospholipase A(2) as the majorenzyme secreted by human keratinocytes and its regulation bythe phorbol ester TPA. J. Invest. Dermatol. 116, 31–39

144. Scott, G. A., Jacobs, S. E., and Pentland, A. P. (2006) sPLA2-Xstimulates cutaneous melanocyte dendricity and pigmentationthrough a lysophosphatidylcholine-dependent mechanism.J. Invest. Dermatol. 126, 855–861

145. Hirobe, T. (1992) Melanocyte stimulating hormone inducesthe differentiation of mouse epidermal melanocytes in serum-free culture. J. Cell Physiol. 152, 337–345

Received for publication August 30, 2006.Accepted for publication November 21, 2006.


Human skin pigmentation: melanocytes modulate skin color in … · 2017-09-23 · Human skin...

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