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Hermansky-Pudlak syndrome (HPS) is an autosomal recessive disorder characterized by the triad of tyrosinase-positive oculocutaneous albinism, bleeding diathesis due to storage-pool deficiency of platelets, and a lysosomal ceroid storage disease. The disorder is particularly frequent in Puerto Rico and in an isolated village in the Swiss Alps. We have used a linkage disequilibrium mapping approach to localize the HPS gene in both of these groups to a 0.6 centiMorgan interval in chromosome segment 10q23.1-q23.3. These results indicate that the Puerto Rican and Swiss forms of HPS are either allelic or that they result from mutations in very closely linked genes in this region. This region of distal chromosome 10q is syntenic to the region of mouse chromosome 19 that includes 'pale ear' (ep) and 'ruby-eye' (ru), which must be considered as potential murine homologues to human HPS.

As ferritin has been identified as an important factor in antioxidant defense in cultured human skin cells we evaluated the presence of ferritin in human skin in vivo and the modifications following irradiation with UVA I, UVA I + II, and solar simulating light by immunohistochemical analysis. We report that the putative protective protein ferritin is regularly present in the basal layer of unirradiated epidermis in vivo and that the induction of ferritin was dependent on wavelength and cell type. Following UVA I radiation, ferritin increased both in epidermal and in dermal tissue. The same response occurred, although to a lesser extent, with UVA I + II but did not occur following solar simulating radiation. Quantitative analysis for ferritin in cultured keratinocytes and fibroblasts from seven individuals following each UV spectra were also assessed by enzyme-linked immunosorbent assay. The induction of ferritin by UV was highly dependent on the waveband and cell type. UVA I and UVA I + II radiations induced ferritin expression in dermal fibroblasts up to 260% and 200% over basal levels, respectively. Solar simulating radiation produced only a small induction of ≈130% over basal ferritin levels in dermal fibroblasts. Ferritin increased in cultured fibroblasts as early as 3 h post-UVA with a peak at 6 h that remained until 48 h; there was no observable qualitative or quantitative increase seen in the undifferentiated cultured epidermal keratinocytes. Our findings indicate that the putative defense system of ferritin exists in human skin in vivo and its induction is dependent on UV spectra and cell type. The increased concentrations of this antioxidant in human skin following acute UV radiation could afford increased protection against subsequent oxidative stress. Key word: oxidative stress. J Invest Dermatol 111:159–163 1998

DNA damage by UV radiation plays an essential role in skin cancer induction. We report that even sub-erythemal doses of solar simulating radiation, are capable of inducing substantial nuclear damage, namely pyrimidine dimers and p53 induction in human skin in situ . The quantity and distribution of p53 induced in human skin by UV radiation depended highly on the waveband and dose of UV used. Solar simulating radiation induced very high levels of p53 throughout all layers in epidermal keratinocytes 24 hr following an erythemal dose (230 ± 15.9/1000 cells), and the induction followed a dose response. Following UVA I + II and UVA I radiations, p53 expression was approximately half of that seen with equivalent biological doses of solar simulating radiation (63.5 ± 28.5 and 103 ± 15.9, respectively). Expression of p53 was seen in basal cell keratinocytes at lower doses of UVA, but all layers of the epidermis were affected at higher doses. Pyrimidine dimer induction, however, was seen to be the same for equivalent biological doses of UVA I, UVA I + II and solar simulating radiations, which coincides with previous findings that pyrimidine dimers initiate the erythemal response and are implicated in skin carcinogenesis. When equivalent biological doses of pure UVA are used with no UVB contamination, significant nuclear alterations occur in human skin in situ, which can approach those seen with UVB radiation. Our results suggest that DNA damage assessed in vivo by immunohistochemistry could provide a very sensitive endpoint for determining the efficacy of protective measures, such as sunscreens or protective clothing, against both UVB- and UVA-induced damage in human skin. Int. J. Cancer 76:201–206, 1998.© 1998 Wiley-Liss, Inc.

Ultraviolet radiation, and in particular UVA (320-400 nm), induces significant oxidative stress to human skin. Ferritin and glutathione have been shown to be among the more important molecules within human skin cells providing protection against this damage, the presence of lower levels of these anti-oxidants giving rise to increased cellular sensitivity to stress. We compared endogenous levels of ferritin and glutathione in human melanoma cells with normal human skin fibroblasts and keratinocytes, also the response of melanoma cells to oxidative stress with fibroblasts and keratinocytes. Ferritin levels were heterogenous in the untreated melanoma cell lines tested and remained the same following oxidative stress (UVA radiation) or hemin treatment. Epidermal keratinocytes were unaffected, as were the melanoma cell lines, but skin fibroblasts showed dose-dependent ferritin depletion. Similar results were seen for glutathione alterations resulting from UVA radiation : melanoma cell lines and epidermal skin keratinocytes remained unchanged following UVA radiation, while skin fibroblasts showed dose-dependent depletion. Our results show that human melanoma cells have low ferritin and glutathione levels, yet are resistant to oxidative stress.

Background: Retinoic acid (RA) cream treatment alters epidermal proliferation and differentiation in photoaged skin. Objective: To study the changes in the expression of markers of epidermal differentiation in photoaged skin following RA cream treatment. Methods: Ten subjects with photoaged skin were examined before treatment and at regular intervals during 12 months of once daily application of 0.05% tretinoin cream over the left forearm. Paraffin-embedded biopsy sections from the forearm were stained with loricrin, filaggrin, involucrin and cornifin antisera in addition to hematoxylin and eosin. The various layers were measured using a calibrated optical micrometer. Results: The thickness of the epidermis increased rapidly, reaching a substantial increase after 1 month of retinoid cream application and retaining it during the following 12 months. The stratum granulosum achieved a transient but substantial increase after 1 and 3 months. Except involucrin, the ratio between the layers expressing the various markers of epidermal differentiation and the epidermis significantly increased following tretinoin cream treatment. Conclusion: RA cream treatment not only increases the thickness of the epidermis but also the programming of late terminal epidermal differentiation, as expressed by the markers studied. Thus, RA appears to affect both proliferation and differentiation of keratinocytes in vivo.

Crude extracts of Panamian plants (153 representing 28 species from 21 families) have been screened for antifungal activity against Candida albicans and Cladosporium cucumerinum. Activity was asses...

A simple bioautographic agar overlay assay using Candida albicans as the indicator organism for the detection and activity-guided fractionation of antifungal compounds by thin layer chromatography has been developed. Inhibition of fungal growth was assessed by the detection of dehydrogenase activity with thiazolyl blue (methylthiazolyltetrazolium chloride; MTT). A series of clinically used antimycotic agents were tested in order to determine the sensitivity of the assay. The compatibility of the agar overlay technique with chemically modified silica gel (Diol and RP-18) plates and with various organic solvents was evaluated. The methodology is also applicable to the search for antibacterial compounds, as shown with Bacillus subtilis as a test organism.

Nucleolar organizer regions (NORs) are loops of DNA which are present within the nucleoli of cells that possess ribosomal RNA (rRNA) genes. NORs are associated with proteins which can be visualized by a simple silver staining technique. As the number of NORs appears to reflect cell and nuclear activity its determination in benign, intermediate and malignant conditions could be of help in their differential diagnosis. In this study, we investigated 35 fibrohistiocytic tumors of the skin of benign, intermediate and malignant potential from 32 patients: 15 fibrous histiocytomas (FH), 5 FH with atypia (AFH), 7 atypical fibroxanthomas (AFX), 5 dermatofibrosarcoma protuberans (DFSP) and 3 malignant FH (MFH). A one-step silver technique was used on formalin or Bouin fixed specimens. Ag-NOR counts from benign conditions (FH, AFH) significantly differed from that of their intermediate-risk (AFX, DFSP) and malignant (MFH) counterparts. Furthermore, in 2 cases (one AFX; one MFH) which twice recurred, Ag-NOR counts steadily increased with time. However, a distinction could not be achieved between AFX and MFH. Although Ag-NOR is a simple and valuable technique, it does not allow a definite distinction between malignant and intermediate processes, at least as far as fibrohistiocytic tumors of the skin are concerned.

Neonatal ichthyosis-sclerosing cholangitis (NISCH) syndrome, a rare autosomal recessive ichthyosis syndrome characterized by scalp hypotrichosis, scarring alopecia, ichthyosis, and sclerosing cholangitis, was described for the first time in 2002. It is caused by a mutation in the gene coding for the tight junction protein claudin-1. Only four patients carrying the same mutation of the CLDN1 gene have been described until now. We report a patient presenting with the clinical characteristics of NISCH syndrome and carrying a novel mutation in the CLDN1 gene.

BP180/collagen XVII is a hemidesmosomal transmembrane molecule serving as cell-surface receptor. Mutations in its gene cause junctional epidermolysis bullosa. Here, we report a patient with mutations in the gene for BP180/collagen XVII, COL17A1, but predominant phenotypic features of epidermolysis bullosa simplex. At birth, the proband presented with bullous lesions on the trunk, face, and hands. Ultrastructurally, hemidesmosomes were fairly normal, but the attachment of intermediate filaments with the hemidesmosomal plaques appeared to be impaired. Blister formation demonstrated both intraepidermal and junctional cleavage. Immunofluores cence staining with antibodies to keratins, several hemidesmosomal proteins, and the extracellular domain of BP180/collagen XVII showed normal staining patterns, whereas an antibody against the intracellular domain of BP180/collagen XVII yielded a negative immunofluorescence signal. Analysis of BP180/collagen XVII cDNA revealed a 1172 bp deletion corresponding to an in-frame deletion from Ile-18 to Asn-407 from the intracellular domain of the polypeptide. Mutation analysis of the COL17A1 gene disclosed a paternal nonsense mutation, R1226X, and a large maternal genomic deletion extending from intron 2 to intron 15, but no mutations in basal keratin genes. These findings underline the functional importance of the intracellular BP180/collagen XVII domain for the interaction of hemidesmosomes with keratin intermediate filaments and for the spatial stability of basal keratinocytes, and provide a functional explanation for the epidermolysis-bullosa- simplex-like phenotype. Further, the data demonstrate that defects in a given gene can cause unexpected phenotypes of epidermolysis bullosa categories, depending on the function of the affected protein domain.

Epidermolytic hyperkeratosis is a hereditary skin disorder characterized by blistering and a marked thickening of the stratum corneum. In one family, affected individuals exhibited a mutation in the highly conserved carboxyl terminal of the rod domain of keratin 1. In two other families, affected individuals had mutations in the highly conserved amino terminal of the rod domain of keratin 10. Structural analysis of these mutations predicts that heterodimer formation would be unaffected, although filament assembly and elongation would be severely compromised. These data imply that an intact keratin intermediate filament network is required for the maintenance of both cellular and tissue integrity.

We report the case of a 30-year-old Nigerian patient with mycetoma of the foot without bone involvement caused by Fusarium sp.. Long-term administration of ketoconazole produced a s

The modification of ferritin in human skin cells in vitro and in vivo following infrared-A irradiation by immunohistochemical analysis and ELISA were evaluated. In addition, we observed that IR-A is not capable of inducing frank damage to DNA (pyrimidine dimers, p53), induction of oxidative stress proteins (heme oxygenase, nitric oxide, superoxide dismutase, heat shock proteins) or proteases (collagenase, stromelysin, gelatinase) involved in carcinogenesis and photoaging of the skin. in vivo, basal levels of ferritin were heterogeneous for all individuals tested but all showed ferritin to stain precisely in the basal layer of unirradiated epidermis. Following IR-A radiation, the ferritin increase was localized to epidermal tissue and showed an increase from 120 to 220%. Parallel to the in vivo analysis, dermal fibroblasts were cultured from six individuals. Quantitative analysis for ferritin in cultured fibroblasts was assessed by ELISA and increases were seen to be dose-dependent and up to 130% of basal levels of ferritin following infrared-A. Our findings indicate that the putative defense system of ferritin that exists in human skin in vivo can be induced by infrared-A radiation and that these wavelengths may prove to be beneficial for human skin. Importantly, following the same doses of IR-A that induced ferritin levels, there was no alteration seen for nuclear DNA type damage, oxidative stress proteins or proteases involved in the degradation of skin. The increased concentrations of this antioxidant in human skin following acute UV radiation could afford increased protection against subsequent oxidative stress.

Hermansky–Pudlak syndrome (HPS) is an often–fatal autosomal recessive disease in which albinism, bleeding, and lysosomal storage result from defects of diverse cytoplasmic organelles: melanosomes, platelet dense bodies, and lysosomes. HPS is the most common single–gene disorder in Puerto Rico, with an incidence of 1 in 1,800. We have identified the HPS gene by positional cloning, and found homozygous frameshifts in this gene in Puerto Rican, Swiss, Irish and Japanese HPS patients. The HPS polypeptide is a novel transmembrane protein that is likely to be a component of multiple cytoplasmic organelles and that is apparently crucial for their normal development and function. The different clinical phenotypes associated with the different HPS frameshifts we observed suggests that differentially truncated HPS polypeptides may have somewhat different consequences for subcellular function.

Four compounds including a flavone, an acetylenic lactone, a prenylated coumarin, and a 3-methyl ether flavone were isolated from the dichloromethane leaf extract of Baccharis pedunculata (Mill.) Cabr. (Asteraceae). The latter three compounds were identified to be responsible for the antifungal activity against some human pathogenic and phytopathogenic fungi. The most active compound, lachnophyllum lactone, an acetylenic lactone, showed a very high toxicity (LD50 2 micrograms/ml) against human keratinocytes.

— Activation of expression of the heme oxygenase (HO) gene appears to be involved in a cellular defense system in mammalian cells. We now demonstrate that while HO-1 mRNA levels are strongly inducible in dermal fibroblasts they are barely inducible in human epidermal keratinocytes following oxidative stress (UVA radiation and hydrogen peroxide). Paralleling this result was the observation that HO-2 mRNA levels were low in dermal fibroblasts but were high in epidermal keratinocytes. In neither case was the HO-2 gene inducible. The expression of the two HO genes led to enzymatic activity in both types of skin cells with an approximately 2.5-fold higher level of enzymatic activity present in keratinocytes compared with fibroblasts derived from the same biopsy. In addition, ferritin levels, which have been found to be augmented via the HO-dependent release of iron from endogenous heme sources, were two- to three-fold higher in keratinocytes compared with matching fibroblasts. This higher ferritin pool would result in an enhancement of cellular iron sequestering capacity that may confer increased resistance to oxidative stress. Indeed, keratinocytes showed less UVA radiation-dependent cell membrane damage than fibroblasts. These results are consistent with the hypothesis that HO expression in human epidermis and dermis is related to cellular defense mechanisms that operate in human skin.

The detection limits in two bioautographic assays have been determined for a series of antifungal compounds, including clinically used antimycotics, fungicidal agrochemicals, and various classes of secondary plant metabolites. Target organisms were the filamentous fungus Cladosporium cucumerinum and the yeast Candida albicans. For clinical agents and agrochemicals, the detection limits in the two assays reflected to a certain extent their known spectrum of activity. Most of the plant-derived compounds tested showed a positive response in both assays, but with detection limits varying by a factor up to tenfold. For screening purposes, it is thus advisable to use both tests, as some activities would otherwise go undetected. The MIC values of these substances were determined in order to verify a possible correlation with the detection limit in the bioautographic assays.

To examine the longitudinal and cross-sectional patterns of yeast colonization in critically ill patients using genotypic characteristics defined by contour-clamped homogeneous electric field (CHEF) gel electrophoresis, 322 clinical isolates of Candida species were prospectively collected from 29 critically ill patients under routine surveillance over a 6-month period. All isolates, recovered from multiple anatomic sites and from the same sites on different days, were characterized by several identification methods (germ tube test), phenotyping (API system), and genotyping (electrophoretic karyotyping). Electrophoretic karyotype (EK) was determined using pulsed field electrophoresis with the CHEF technique. We used a karyotyping system for Candida albicans (EK code) that facilitated intraspecies delineation. C. albicans colonized 83% of the 29 patients. Candida sp. strains isolated from an individual patient had an identical EK pattern, even when isolated from different body sites, and remained the same over a prolonged period, up to 140 days. EK delineated not only the different Candida species, but also different strains of C. albicans. Strains of C. albicans isolated from different patients were distinguished using the EK pattern, but not API system. Minor variations in EK pattern could be demonstrated in a minority of strains recovered from four patients and were interpreted as chromosomal rearrangements between parent strains. Severe candidal infections, including eight episodes of fungemia, occurred in 11 of 29 patients (38%). All patients had been previously colonized with strains with identical EK patterns. Infection occurred a mean of 25 days after initial surveillance cultures grew yeast. No horizontal transmission could be demonstrated during the study period. In conclusion, EK is a reproducible, stable marker allowing inter-, as well as, intraspecies Candida strain delineation. EK strain delineation is a useful tool in candidal epidemiologic and pathogenic studies. Yeast colonization with the same strain preceded infection in critically ill patients.

Aspergillus fumigatus secreted an inducible alkaline protease (AlPase) when cultivated in the presence of collagen (200 micrograms/ml) as sole nitrogen and carbon source. Proteolytic activity was maximum at pH 9.0 with azocollagen as substrate. The enzyme, which was the major protein found in the supernate of a liquid culture, was purified by ammonium sulphate precipitation and gel filtration. The Mr was determined to be 33 Kda by gel filtration and sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The isoelectric point was estimated to be pH 8.2. Divalent cations strongly inhibited enzyme activity, whereas non-ionic detergents and reducing agents had no effect. A. fumigatus AlPase was totally inhibited by phenylmethanesulphonyl fluoride, antipain, chymostatin and alpha-2-macroglobulin. A. fumigatus AlPase is closely related to the A. oryzae AlPase, a serine protease of the subtilisin family, as attested by the antigen pattern seen by immunoblotting. The high collagenic activity and the ability of A. fumigatus AlPase to digest elastin could play a role in the invasion of the tissues by the fungus.

The epidermis of superficial human skin samples could easily be separated from the dermis following incubation at +4 degrees C for 1 h in a solution containing 250-500 micrograms/ml thermolysin, a proteolytic enzyme hitherto mostly used for protein analysis. Light and electron microscopy revealed that the dermo-epidermal separation occurred at the basement membrane between the sites of bullous pemphigoid antigen and laminin and that the hemidesmosomes were selectively disrupted. The cohesion and morphology of the separated epidermis as well as the immunologic parameters investigated were not altered by this procedure. The clear cut dermo-epidermal separation produced by thermolysin treatment differed from the separation obtained with trypsin, which predominantly occurred between basal and suprabasal cells by disruption of desmosomes.

The Spitz nevus (large spindle and/or epithelioid cell nevus) is a benign acquired melanocytic tumour found predominantly in children and adolescents. Depending on the architecture (junctional, compound, dermal) and cytology (predominance of spindle or epithelioid cells, or presence of both cell types) several variants have been distinguished. A histological feature occasionally leading to diagnostic difficulties is the involvement of the suprabasal epidermis. The present study details the occurrence of intraepidermal involvement (pagetoid spread, nesting of cells, trans-epidermal elimination) according to the age of the patients. 89 cases of Spitz nevus were reviewed. There were 19 compound Spitz nevi with evidence of epidermal involvement, 56 cases of compound Spitz nevi without evidence of epidermal involvement and 14 cases of dermal Spitz nevi. The epidermal involvement mainly consisted of nesting of nevus cells above the basal cell layer, 9 cases clearly showing evidence of transepidermal elimination of cell nests. Upward spread of single melanocytes (pagetoid spread) was minimal and always associated with nesting or trans-epidermal elimination. The mean age of patients was significantly lower in the group of compound nevi with intraepidermal involvement (7.16 +/- 4.52 years), than in the group of compound nevi without epidermal involvement (13.18 +/- 8.88 years). The age of the latter group was significantly lower than the group of pure intradermal nevi (30.14 +/- 11.25 years). Thus, involvement of the suprabasal epidermis is not a feature of Spitz nevi in adults; in such a case the diagnosis of malignant melanoma should be considered.

The monoethyl ether of hydroquinone (MEH) is known to be a very potent depigmenting agent for skin and hair of black guinea pigs. Its action is due to selective destruction of melanocytes. In order to better understand its pharmacology, MEH was applied or injected into the skin of red and albino guinea pigs, grey DBA-1 mice, black C-57 mice and Syrian golden hamsters. Experimental melanomas S 91, B 16, Asc.M.Mel.1 and V 15 were also treated with this agent. MEH depigmented the brown epidermis of the red guinea pigs; the red hair, however, regrew without any color change. The melanocytes of the albino guinea pigs were not damaged by MEH. The hair of the 2 types of mice treated always became white and the fur of Syrian hamsters lost its black component. The survival time of animals with transplanted melanomas was not increased and gross and microscopic appearance of their melanomas was not altered by intratumoral injections of MEH.

The albinotic skin and hair of 2 patients with Hermansky-Pudlak syndrome were investigated by light and electron microscopy. Incubation of hairbulbs and epidermis in l-dopa revealed a weak tyrosinase activity. The epidermal melanocyte population was of normal density. The most striking feature was the presence of numerous giant melanosomes resembling those mainly reported in various hyperpigmented skin lesions. The association of this melanosomal disorder with the platelet dysfunction and ceroid storage typical of the autosomal recessive Hermansky-Pudlak syndrome might provide new insights into the mechanism leading to formation of giant melanosomes.

• Background.— Loricrin and involucrin are major precursor proteins to the cornified cell envelope expressed late in epidermal differentiation. Involucrin expression starts in the upper spinous layers in normal human epidermis and precedes loricrin expression, which is restricted to the granular layer. Subsequently, both proteins become cross-linked by the activity of transglutaminases TGK/E as major components of the cornified cell envelope by N e -(γ-glutamyl)lysine isopeptide bonds. In this study, three cases of lamellar ichthyosis were analyzed by immunohistologic study with antibodies to loricrin, involucrin, filaggrin, and transglutaminase TGK. Observations.— A high expression of loricrin and involucrin with a peculiar and abnormal cytoplasmic staining concurred with a diminished cytoplasmic staining of transglutaminase TGK as assessed by antibodies B.C.1 and K.D.3. This pattern was absent in a collodion baby at birth but present 2 weeks later before a phenotype of lamellar ichthyosis appeared clinically. Conclusions.— The results suggest that in our cases of lamellar ichthyosis, (1) disturbed membrane anchorage of transglutaminase TGK could alter loricrin and involucrin cross-linkage and the formation of the cornified cell envelope and that (2) immunohistologic study might serve as an early diagnostic and prognostic tool in the treatment of collodion babies. ( Arch Dermatol. 1993;129:618-624)

Epidermolytic hyperkeratosis is caused by mutations of the differentiation-specific keratins K1 and R10. These mutations produce a weakened cytoskeleton that is prone to collapse resulting in cell fragility and lysis. In this study we have analyzed cultured keratinocytes from EHK patients bearing 10R-to-H and 15L-to-S mutations within the 1A segment of the K10 rod domain. Keratinocytes were grown submerged in serum-free medium and induced to differentiate by growing to confluence and increasing the Ca++ concentration in the medium. Cultures were either harvested for mRNA sequence analysis or subjected to immunofluorescence microscopy. Differentiating keratinocytes from these patients were found to express these K10 mutations in their mRNA. Moreover, these cells could be distinguished from normal kerati- nocytes by their aberrrant morphology. EHK keratinocytes frequently exhibited a collapsed perinuclear network of K1/ K10 filaments and sometimes peripheral granules of K1 and K10 aggregates, reminiscent of the cells of the suprabasal layers in these patients. This report documents the expression of mutant keratin 10 in cultured EHK keratinocytes.

Lamellar ichthyosis is a severe congenital skin disorder characterized by generalized large scales and variable redness. Affected individuals in three families exhibited drastically reduced keratinocyte transglutaminase (TGK) activity. In two of these families, expression of TGK transcripts was diminished or abnormal and no TGK protein was detected. Homozygous or compound heterozygous mutations of the TGK gene were identified in all families. These data suggest that defects in TGK cause lamellar ichthyosis and that intact cross-linkage of cornified cell envelopes is required for epidermal tissue homeostasis.

Experiments with various topically applied compounds have demonstrated that 2-mercaptoethylamine HCI (MEA) and N(2-mercaptoethyl)-dimethylamine HCI (MEDA) are very potent agents that cause depigmentation in treated areas only. When applied to the skin of black guinea pigs, they inhibited melanogenesis, acted selectively on melanocytes, decreased melanocyte density significantly, and rendered the few remaining melanocytes degenerative. In epidermis from MEDA-treated areas, melanocytes could be detected only rarely by electron microscopy. Those that were found contained few melanized melanosomes, many vacuoles, and swollen mitochondria. Depigmentation by any exogenous agent can result from destruction or loss of melanocytes; interference with (1) the biosynthesis of premelanosomes and melanosomes, (2) the conversion of tyrosine to dopa to melanin, (3) the biosynthesis of tyrosinase or the active center of the enzyme, (4) the transfer of melanosomes to keratinocytes; and alteration of the melanin present in melanosomes. Depigmentation by MEDA appears to result from selective destruction or degeneration of melanocytes.