A mechanism of melanogenesis mediated by E-cadherin downregulation and its involvement in solar lentigines.

OBJECTIVE: Intensive studies have revealed that pleiotropic melanocytic factors are associated with age-spot formation. Dysfunctional keratinocyte differentiation is thought to be an upstream cause of age-spot formation. Although it has been shown that keratinocyte differentiation is mediated by the cell-cell contact factor E-cadherin, its involvement in age-spot formation remains unknown. Thus, to determine the origin of age-spots and an integrated solution for the same, we focused on E-cadherin expression in the present study. METHODS: First, we assessed the solar lentigines in cutaneous and cultured cells by means of immunofluorescence staining. Following that, keratinocytes treated with siRNAs against E-cadherin were co-cultured with melanocytes, and the secreted factors were identified by means of proteomic analysis of the culture supernatants. We also performed quantitative PCR to assess melanogenesis activity and screen ingredients. For behavioural analysis of melanocytes, we performed time-lapse imaging using confocal laser scanning microscopy. RESULTS: E-cadherin expression was downregulated in the epidermis of the solar lentigines, suggesting its involvement in age-spot formation. E-cadherin knocked down keratinocytes not only promoted the secretion of melanocytic/inflammatory factors but also increased melanogenesis by upregulating the expression of melanogenesis factors. Furthermore, live-imaging showed that the downregulation of E-cadherin inhibited melanocyte dynamics and accelerated melanin uptake. Finally, we identified Rosa multiflora fruit extract as a solution that can upregulate E-cadherin expression in keratinocytes. CONCLUSION: Our findings showed that E-cadherin downregulation triggers various downstream melanocytic processes, such as the secretion of melanocytic factors and melanogenesis. Additionally, we showed that the Rosa multiflora fruit extract upregulated E-cadherin expression in keratinocytes.

OBJECTIF: Des études intensives ont révélé que les facteurs mélanocytaires pléiotropiques sont associés à la formation de taches de vieillesse. On pense que la différenciation des kératinocytes dysfonctionnels est une cause en amont de la formation des taches de vieillesse. Bien qu'il ait été démontré que la différenciation des kératinocytes est médiée par le facteur de contact cellule-cellule E-cadhérine, son implication dans la formation des taches de vieillesse reste inconnue. Ainsi, pour déterminer l'origine des taches de vieillesse et une solution intégrée pour celles-ci, nous nous sommes concentrés sur l'expression de la E-cadhérine dans la présente étude. MÉTHODES: Tout d'abord, nous avons évalué les lentigines solaires dans les cellules cutanées et cultivées au moyen d'une coloration par immunofluorescence. Par la suite, les kératinocytes traités avec des siRNA contre l'E-cadhérine ont été co-cultivés avec des mélanocytes, et les facteurs sécrétés ont été identifiés au moyen d'une analyse protéomique des surnageants de culture. Nous avons également effectué une PCR quantitative pour évaluer l'activité de la mélanogénèse et dépister les ingrédients. Pour l'analyse comportementale des mélanocytes, nous avons réalisé une imagerie accélérée à l'aide de la microscopie confocale à balayage laser. RÉSULTATS: L'expression de l'E-cadhérine a été régulée à la baisse dans l'épiderme des lentigines solaires, suggérant son implication dans la formation des taches de vieillesse. Les kératinocytes dans lesquels l'E-cadhérine a été réduite non seulement ont favorisé la sécrétion de facteurs mélanocytaires/inflammatoires, mais ont également accru la mélanogenèse en régulant à la hausse l'expression de facteurs de mélanogenèse. De plus, l'imagerie en direct a montré que la régulation négative de l'E-cadhérine inhibait la dynamique des mélanocytes et accélérait l'absorption de la mélanine. Enfin, nous avons identifié l'extrait de fruit de Rosa multiflora comme une solution capable de réguler positivement l'expression de l'E-cadhérine dans les kératinocytes. CONCLUSION: Nos résultats ont montré que la régulation négative de la E-cadhérine déclenche divers processus mélanocytaires en aval, tels que la sécrétion de facteurs mélanocytaires et la mélanogénèse. De plus, nous avons montré que l'extrait de fruit de Rosa multiflora régulait à la hausse l'expression de l'E-cadhérine dans les kératinocytes.

Prevalence of nevi, atypical nevi, and lentigines in relation to tobacco smoking.

BACKGROUND: Melanocytic nevi have a complex evolution influenced by several endogenous and exogenous factors and are known risk factors for malignant melanoma. Interestingly, tobacco use seems to be inversely associated with melanoma risk. However, the association between tobacco use and nevi and lentigines has not yet been evaluated. METHODS: We investigated the prevalence of nevi, atypical nevi, and lentigines in relation to tobacco smoking in a cohort of 59 smokers and 60 age- and sex-matched nonsmokers, using a questionnaire and performing a total body skin examination by experts. RESULTS: No significant differences were detected between smokers and nonsmokers in the numbers of nevi, atypical nevi, and lentigines in sun-exposed areas (p = 0.966, 0.326, and 0.241, respectively) and in non-sun-exposed areas (p = 0.095, 0.351, and 0.546, respectively). CONCLUSION: Our results revealed no significant differences in the prevalence of nevi, atypical nevi, and lentigines between smokers and nonsmokers in sun-exposed and non-sun-exposed areas.

Autophagy Declines with Premature Skin Aging resulting in Dynamic Alterations in Skin Pigmentation and Epidermal Differentiation.

Autophagy is a membrane traffic system that provides sustainable degradation of cellular components for homeostasis, and is thus considered to promote health and longevity, though its activity declines with aging. The present findings show deterioration of autophagy in association with premature skin aging. Autophagy flux was successfully determined in skin tissues, which demonstrated significantly decreased autophagy in hyperpigmented skin such as that seen in senile lentigo. Furthermore, an exacerbated decline in autophagy was confirmed in xerotic hyperpigmentation areas, accompanied by severe dehydration and a barrier defect, which showed correlations with skin physiological conditions. The enhancement of autophagy in skin ex vivo ameliorated skin integrity, including pigmentation and epidermal differentiation. The present results indicate that the restoration of autophagy can contribute to improving premature skin aging by various intrinsic and extrinsic factors via the normalization of protein homeostasis.

Melanocyte Activation Mechanisms and Rational Therapeutic Treatments of Solar Lentigos.

To characterize the pathobiology of solar lentigos (SLs), analyses by semiquantitative RT-PCR, Western blotting, and immunohistochemistry revealed the upregulated expression of endothelin (EDN)-1/endothelin B receptors (EDNBRs), stem cell factor (SCF)/c-KIT, and tumor necrosis factor (TNF)α in the lesional epidermis, which contrasted with the downregulated expression of interleukin (IL) 1α. These findings strongly support the hypothesis that previous repeated UVB exposure triggers keratinocytes to continuously produce TNFα. TNFα then stimulates the secretion of EDNs and the production of SCF in an autocrine fashion, leading to the continuous melanogenic activation of neighboring melanocytes, which causes SLs. A clinical study of 36 patients with SLs for six months treated with an M. Chamomilla extract with a potent ability to abrogate the EDN1-induced increase in DNA synthesis and melanization of human melanocytes in culture revealed a significant improvement in pigment scores and color differences expressed as L values. Another clinical study using a tyrosinase inhibitor L-ascorbate-2-phosphate 3 Na (ASP) demonstrated that L values of test lotion (6% APS)-treated skin significantly increased in SLs and in non-lesional skin with a significantly higher ΔL value in SLs when compared with non-lesional skin. The sum of these findings strongly suggests that combined topical treatment with EDN signaling blockers and tyrosinase inhibitors is a desirable therapeutic choice for SLs.

Whitening effect of L-ascorbate-2-phosphate trisodium salt on solar lentigos.

Little is known about the anti-pigmenting effects of whitening agents on solar lentigos (SLs), which comprise ~ 60% of hyperpigmented facial lesions of Asian subjects. Lotions with or without 6% L-ascorbate-2-phosphate trisodium salt (APS) [test lotion (TL) and placebo lotion (PL), respectively] were applied twice daily for 24 weeks in a double-blind half-face study of 27 Japanese females with SLs on both sides of their faces. Pigmentation scores were evaluated using a photo-scale and the skin colors were assessed using a color difference meter and a mexameter for SLs and the non-lesional surrounding skin (NLS). Although the pigmentation scores were not significantly different between the TL and PL-treated SLs after 24 weeks, the L values of TL-treated SLs and NLS increased significantly with a significantly higher △L value in SLs than in NLS. In contrast, the L values of PL-treated SLs and NLS remained unchanged after the treatment. The number of subjects with > 2.0 △L was 7 of 27 (TL) and 0 of 27 (PL) in SLs and 3 of 27 (TL) and 0 of 27 (PS) in NLS. In contrast, the melanin index in TL-treated SLs and NLS significantly decreased with a significantly higher △melanin index in SLs than in NLS. Similarly, the melanin index of PL-treated SLs and NLS were significantly decreased with a significantly higher △melanin index in SLs than in NLS. These findings strongly indicate that APS has a weak but significant anti-pigmenting effect on SLs and a significant whitening effect even on normally pigmented healthy skin.

Relationship between tumor-associated immune infiltrate and p16 staining over clinicopathological features in acral lentiginous melanoma.

PURPOSE: This study aims to evaluate the association between composition of tumor-infiltrating lymphocytes (TIL) and expression of p16 in acral lentiginous melanoma (ALM), and their impact on prognosis. MATERIALS AND METHODS: A cohort of 148 surgical pathology specimens of ALM was studied. TIL were evaluated by immunohistochemical detection of CD3 and CD8, along with CD20, CD4, CD68, and CD163 in a subset of 43 cases. p16 protein expression was also investigated in all the cases. RESULTS: The median age was 66 years, median Breslow thickness was 6.0 mm, grade III TIL was found in 28.4% and lymph nodes were involved in 54.2%. Breslow thickness (p < 0.001), stage I-II (p < 0.001), negative lymph nodes (p < 0.001) and < 10% p16 (p = 0.01) were associated with longer survival. Grade III of TIL was associated with thinner Breslow thickness (p = 0.008) and lower mitosis (p = 0.047). A higher density of CD3 TIL was associated with male gender (p = 0.008), thinner Breslow thickness (p = 0.047), negative lymph node (p = 0.031), early stage (p = 0.046), and p16 nuclear expression of > 10% (p = 0.045). Higher CD8 TIL was associated with > p16 (p = 0.03). Survival analysis found that longer survival had a trend to be associated with high TIL (p = 0.090). Levels of CD3+ and CD8+ cells were correlated with those of CD4+, CD20+, CD68+ and CD163+ immune cells. CONCLUSIONS: Higher levels of TIL tend to be associated with better overall survival in ALM. Loss of expression of p16 is associated with lower levels of CD3+ and CD8+ TIL, indicating a probable relationship between p16 and TIL immune response in ALM .

Molecular and histological characterization of age spots.

Age spots, also called solar lentigines and lentigo senilis, are light brown to black pigmented lesions of various sizes that typically develop in chronically sun-exposed skin. It is well known that age spots are strongly related to chronic sun exposure and are associated with photodamage and an increased risk for skin cancer; however, the mechanisms underlying their development remain poorly understood. We used immunohistochemical analysis and microarray analysis to investigate the processes involved in their formation, focusing on specific markers associated with the functions and proliferation of melanocytes and keratinocytes. A total of 193 genes were differentially expressed in age spots, but melanocyte pigment genes were not among them. The increased expression of keratins 5 and 10, markers of basal and suprabasal keratinocytes, respectively, in age spots suggests that the increased proliferation of basal keratinocytes combined with the decreased turnover of suprabasal keratinocytes leads to the exaggerated formation of rete ridges in lesional epidermis which in turn disrupts the normal processing of melanin upwards from the basal layer. Based on our results, we propose a model for the development of age spots that explains the accumulation of melanin and the development of extensive rete ridges in those hyperpigmented lesions.

Eruptive Facial Postinflammatory Lentigo: Clinical and Dermatoscopic Features.

The face has not been considered a common site of fixed drug eruption, and the authors lack dermatoscopic studies of this condition on the subject. The authors sought to characterize clinical and dermatoscopic features of 8 cases of an eruptive facial postinflammatory lentigo. The authors conducted a retrospective review of 8 cases with similar clinical and dermatoscopic findings seen from 2 medical centers in 2 countries during 2010-2014. A total of 8 patients (2 males and 6 females) with ages that ranged from 34 to 62 years (mean: 48) presented an abrupt onset of a single facial brown-pink macule, generally asymmetrical, with an average size of 1.9 cm. after ingestion of a nonsteroidal antiinflammatory drugs that lasted for several months. Dermatoscopy mainly showed a pseudonetwork or uniform areas of brown pigmentation, brown or blue-gray dots, red dots and/or telangiectatic vessels. In the epidermis, histopathology showed a mild hydropic degeneration and focal melanin hyperpigmentation. Melanin can be found freely in the dermis or laden in macrophages along with a mild perivascular mononuclear infiltrate. The authors describe eruptive facial postinflammatory lentigo as a new variant of a fixed drug eruption on the face.

The Effect of MCP-1/CCR2 on the Proliferation and Senescence of Epidermal Constituent Cells in Solar Lentigo.

Solar lentigo (SL) is a representative photoaging skin disorder. Alteration of the main epidermal constituent cells-keratinocytes and melanocytes-in relation to the photoaged dermal environment or chemokine/cytokine network is suggested as its pathogenesis. Among these, we focused on monocyte chemoattractant protein-1 (MCP-1), as it is known to be associated with tissue aging. For the first time, we report that the MCP-1 receptor, CCR2, is expressed in normal human melanocytes. In SL tissue, there was an increase of CCR2+Melan A+ melanocytes with positivity to Rb protein compared to peri-lesional normal skin. MCP-1 induced the proliferation of normal human melanocytes without a significant change in the melanin content. MCP-1 treatment in normal human keratinocytes showed an increase in senescence-associated ß-galactosidase staining and p53 and p21 protein expressions. In summary, MCP-1 may participate in the development of SL by affecting epidermal constituent cells, for example, by inducing melanocyte proliferation and keratinocyte senescence.

Developmental SHP2 dysfunction underlies cardiac hypertrophy in Noonan syndrome with multiple lentigines.

Hypertrophic cardiomyopathy is a common cause of mortality in congenital heart disease (CHD). Many gene abnormalities are associated with cardiac hypertrophy, but their function in cardiac development is not well understood. Loss-of-function mutations in PTPN11, which encodes the protein tyrosine phosphatase (PTP) SHP2, are implicated in CHD and cause Noonan syndrome with multiple lentigines (NSML), a condition that often presents with cardiac hypertrophic defects. Here, we found that NSML-associated hypertrophy stems from aberrant signaling mechanisms originating in developing endocardium. Trabeculation and valvular hyperplasia were diminished in hearts of embryonic mice expressing a human NSML-associated variant of SHP2, and these defects were recapitulated in mice expressing NSML-associated SHP2 specifically in endothelial, but not myocardial or neural crest, cells. In contrast, mice with myocardial- but not endothelial-specific NSML SHP2 expression developed ventricular septal defects, suggesting that NSML-associated mutations have both cell-autonomous and nonautonomous functions in cardiac development. However, only endothelial-specific expression of NSML-associated SHP2 induced adult-onset cardiac hypertrophy. Further, embryos expressing the NSML-associated SHP2 mutation exhibited aberrant AKT activity and decreased downstream forkhead box P1 (FOXP1)/FGF and NOTCH1/EPHB2 signaling, indicating that SHP2 is required for regulating reciprocal crosstalk between developing endocardium and myocardium. Together, our data provide functional and disease-based evidence that aberrant SHP2 signaling during cardiac development leads to CHD and adult-onset heart hypertrophy.

A double-blind, placebo-controlled randomized trial of Serratulae quinquefoliae folium, a new source of ß-arbutin, in selected skin hyperpigmentations.

BACKGROUND: Arbutin is one of the most effective lightening substances. Serratula quinquefolia is a new source of its ß-anomer. The HPLC method showed that the solid content of this compound in the dried plant raw material accounts for 6.86%. The leaves of Serratula quinquefolia do not contain hydroquinone. AIMS: To assess the efficacy of the aqueous extract from' leaf of five-leaf serratula as a skin-lightening agent. PATIENTS/METHODS: We did a randomized, placebo-controlled, double-blind trial. The study involved 102 women aged 26-55, with two kinds of hyperpigmentary diseases: melasma and lentigo solaris. Patients were randomly assigned to one of the treatment groups: a study group (N = 54) or a control group (N = 48). The study group applied the cream with the aqueous extract from leaf of five-leaf serratula containing 2.51% of arbutin. The cream was applied twice a day on the discolored side for 8 weeks. RESULTS: The experimental data showed that the cream with the extract causes decreased level of melanin in the skin pigmentation spot. Clinical effect in the form of lightening and evening skin tone on the discolored side was observed in 75.86% of the female patients with melasma and 56.00 % of the female patients with lentigo solaris. CONCLUSIONS: The cream with the aqueous extract from leaf of five-leaf serratula proved to be an effective and safe preparation for lightening skin discolorations (66.67 % of the female patients in the study group).

Triple combination as adjuvant to cryotherapy in the treatment of solar lentigines: investigator-blinded, randomized clinical trial.

BACKGROUND: Post-inflammatory hyperpigmentation is a frequent concern when treating solar lentigines. OBJECTIVES: To assess the safety and efficacy of a triple combination cream with fluocinolone acetonide 0.01%, hydroquinone 4% and tretinoin 0.05% as adjuvant to cryotherapy in the treatment of solar lentigines in hands dorsum, and in the prevention of post-inflammatory hyperpigmentation after cryotherapy. METHODS: This prospective, randomized, controlled, investigator-blinded, single-centre study enrolled 50 patients. Twenty-five patients received a 2-week daily triple combination cream plus sunscreen pre-treatment and 25 received sunscreen alone. After that, cryotherapy was performed in all patients followed by a 3-week recovery period. After this period, patients received the same initial treatment and were followed up for 8 weeks. Melanin and erythema levels of a target and a control lentigo were objectively measured using a narrowband reflectance spectrophotometer. Lentigines count, colour homogeneity and global improvement were also assessed. RESULTS: The number of solar lentigines reduced in the first 2 weeks only in patients who used the triple combination 25 ± 7 vs. 22 ± 8 (P < 0.0001), and reduced at the end of the study for both groups (P < 0.0001). The melanin levels also reduced in the first 2 weeks only in patients who used the triple combination 297 ± 69 vs. 273 ± 66 (P < 0.0001) and reduced at the end of the study for both groups (P < 0.0001). Erythema and residual blisters from cryotherapy were the reported adverse reactions. CONCLUSION: Triple combination cream can be used to enhance the resolution of solar lentigines, and to significantly reduce melanin levels and lentigines count, improving treatment results. It was well-tolerated and did not increase the occurrence of neither erythema nor other side-effects after the cryotherapy.

Accelerated differentiation of melanocyte stem cells contributes to the formation of hyperpigmented maculae.

It has been reported that the abnormal regulation of melanocyte stem cells (McSCs) causes hair greying; however, little is known about the role of McSCs in skin hyperpigmentation such as solar lentigines (SLs). To investigate the involvement of McSCs in SLs, the canonical Wnt signalling pathway that triggers the differentiation of McSCs was analysed in UVB-induced delayed hyperpigmented maculae in mice and human SL lesions. After inducing hyperpigmented maculae on dorsal skin of F1 mice of HR-1× HR/De, which was formed long after repeated UVB irradiation, the epidermal Wnt1 expression and the number of nuclear ß-catenin-positive McSCs were increased as compared to non-irradiated control mice. Furthermore, the expression of dopachrome tautomerase (Dct), a downstream target of ß-catenin, was significantly upregulated in McSCs of UVB-irradiated mice. The Wnt1 expression and the number of nuclear ß-catenin-positive McSCs were also higher in human SL lesions than in normal skin. Recombinant Wnt1 protein induced melanocyte-related genes including Dct in early-passage normal human melanocytes (NHEMs), an in vitro McSC model. These results demonstrate that the canonical Wnt signalling pathway is activated in SL lesions and strongly suggest that the accelerated differentiation of McSCs is involved in SL pathogenesis.

Protective effect of red orange extract supplementation against UV-induced skin damages: photoaging and solar lentigines.

BACKGROUND: Exposure of the skin to solar ultraviolet (UV) radiations causes important oxidative damages that result in clinical and hystopathological changes, contributing to premature skin aging. Hyperpigmented lesions, also known as age spots, are one of the most visible alterations in skin photoaging. Skin is naturally equipped with antioxidant systems against UV-induced ROS generation; however, these antioxidant defenses are not completely efficient during exposure to sunlight. Oral antioxidants are able to counteract the harmful effects of UV radiation and to strengthen the physiological skin antioxidant defenses. AIMS: The present study was performed to evaluate the in vivo skin photo-protecting and anti-aging effects of a red orange (Citrus sinensis varieties Moro, Tarocco and Sanguinello) extract supplementation. Previous studies showed that red orange extracts possess strong in vitro free radical scavenging/antioxidant activity and photo-protective effects on human skin. MATERIALS/METHODS: The photo-protective effects of red orange extract intake against UV-induced skin erythema and melanin production in solar lentigo was evaluated on healthy volunteers by an objective instrumental method (reflectance spectrophotometry). RESULTS: Data obtained from in vivo studies showed that supplementation of red orange extract (100 mg/daily) for 15 days brought a significant reduction in the UV-induced skin erythema degree. Moreover, skin age spots pigmentation (melanin content) decreased from 27% to 7% when subjects were exposed to solar lamp during red orange extract supplementation. CONCLUSIONS: Red orange extract intake can strengthen physiological antioxidant skin defenses, protecting skin from the damaging processes involved in photo-aging and leading to an improvement in skin appearance and pigmentation.

Novel morphological study of solar lentigines by immunohistochemical and electron microscopic evaluation.

Solar lentigines (SL) are hyperpigmented lesions generally seen in elderly people. Their pathogenesis has not been completely elucidated. We examined 75 cases of SL using routine histopathology and immunohistochemistry. In addition, seven cases were evaluated by electron microscopy. Histopathologically, we observed vacuolar changes in the dermoepidermal junction in 85% of the cases. Dermal melanophages were seen in 77% of the cases. The immunohistochemical expression rates in the epidermis for cytokeratin (CK)15, CK14, CK10, p63 and nestin were 76%, 100%, 100%, 100% and 17%, respectively. In 58 cases showing dermal melanophages, expression rates of CD163 and factor XIIIa on melanophages were 79% and 83%, respectively. Double positivity for both proteins was identified in 44 cases (75%). Ultrastructurally, vacuolar structures were seen in the cytoplasm of basal cells and upper dermis in all cases examined. We observed elimination processes of damaged basal keratinocytes, which were probably produced by ultraviolet (UV) irradiation, into the papillary dermis. The segregated damaged cell bodies containing melanin granules seemed to be phagocytosed by poorly immunostimulatory macrophages labeled immunohistochemically by CD163 and factor X IIIa, contributing to prolonged pigmentation of SL. In addition, repeated basal keratinocyte damages may be in association with altered CK and p63 expression patterns in the constituent cells of SL.

Abrogating effect of N-linked carbohydrate modifiers on the stem cell factor and endothelin-1-stimulated epidermal pigmentation in human epidermal equivalents.

BACKGROUND: We previously demonstrated that the hyperpigmentation that occurs in UVB-melanosis as well as in solar lentigos is associated with the increased production of melanogenic cytokines, such as endothelin (EDN)-1 and stem cell factor (SCF), by keratinocytes in those areas of the skin. OBJECTIVE: We developed a model for these hyperpigmentary disorders in EDN1+SCF stimulated human epidermal equivalents (HEEs) and characterized the effects of the N-linked carbohydrate core synthesis inhibitor glucosamine or N-linked carbohydrate processing inhibitors deoxynojirimycin or monensin on the stimulated HEE pigmentation. METHODS: Those effects were assessed by melanin analysis, real-time RT-PCR and Western blotting. RESULTS: The addition of these N-linked carbohydrate modifiers (NCMs) markedly abolished the EDN1+SCF-elicited increase in HEE pigmentation over 14 days. Real-time RT-PCR and Western blotting of these NCM-treated HEEs unexpectedly revealed that the EDN1+SCF-stimulated steady-state levels of tyrosinase (TYR), TYR-related protein-1, dopachrome tautomerase and PMEL17 as well as microphthalmia-associated transcription factor (MITF) were significantly attenuated at the transcriptional and translational levels without any cytotoxic effects on keratinocytes and melanocytes in the HEEs. Pre-treatment of cultured normal human melanocytes with the NCMs interrupted the EDN1+SCF-induced stimulation of steady-state levels of MITF at the transcriptional and translational levels and TYR activity without any direct inhibitory effect on the catalytic activity of TYR in vitro. CONCLUSION: This study provides evidence that NCMs have a potential to attenuate the EDN1+SCF-stimulated pigmentation of HEEs by abrogating the increased steady-state levels of MITF mRNA, which results in the attenuation of the increased steady-state levels of these melanocyte-specific proteins.

Investigation by in vivo reflectance confocal microscopy: melanocytes at the edges of solar lentigines.

In vivo reflectance confocal microscopy (RCM) provides high-resolution, real-time optical sections of the skin in a non-invasive manner, allowing visualization of the skin in its native state. Highly reflective skin components including melanin, collagen and keratin appear bright (white) in RCM images. RCM examination of solar lentigines is known to show features that correlate well with histologic findings such as supranuclear melanin caps, but there are a limited number of reports on melanocyte dendrites. In this study, we utilized RCM to investigate the melanocyte dendricity and distribution within solar lentigines. Seventeen healthy Japanese females who had fairly large solar lentigines on their faces were recruited to join our clinical study, and we examined them by using RCM on their non-lesional areas, and the inside and the outer rim of the lesional areas. As a result, we discovered that dendritic melanocytes were rarely seen in the center of a solar lentigo (SL), but were seen at a very high frequency in the outer rim of a SL. The results suggest that the melanocytes are more active at the edge of a SL, produce more melanin, and often spread their dendrites widely in a horizontal direction. The findings in this report might shed light on the dynamic pathomechanisms of solar lentigines in vivo.

Quantitative comparison of MiTF, Melan-A, HMB-45 and Mel-5 in solar lentigines and melanoma in situ.

BACKGROUND: It is often challenging to reliably assess the number of lesional melanocytes in intraepidermal melanocytic proliferations involving sun-damaged skin. Therefore, dermatopathologists routinely use immunostains to help differentiate melanocytes from surrounding keratinocytes. METHODS: Forty-three cases of solar lentigo or melanoma in situ (of the lentigo maligna type) were retrospectively chosen (20 melanomas in situ and 23 solar lentigo). Microphthalmia transcription factor (MiTF), HMB-45, Melan-A and Mel-5 immunostains were performed with an Azure blue counterstain, and the mean melanocyte counts were calculated within a 1-mm segment of epidermis. RESULTS: In solar lentigines, the mean melanocyte counts were 27 (MiTF), 23 (HMB-45 and Mel-5) and 41 (Melan-A), as compared to hematoxylin and eosin (H&E) (25). In melanoma in situ, the mean melanocyte counts were 112 (MiTF), 149 (Melan-A), 111 (HMB-45) and 80 (Mel-5), as compared to H&E (109). CONCLUSIONS: These results show that Melan-A significantly overestimates the density of melanocytes within dermatoheliotic skin. Compared to other tested stains, nuclear staining MiTF allowed greater distinction of melanocytes from keratinocytes with melanized cytoplasm. These findings indicate that MiTF is a superior marker for quantification of melanocytes in the evaluation of subtle intraepidermal melanocytic proliferations and in the differential diagnosis of solar lentigo.

Role of fibroblast-derived growth factors in regulating hyperpigmentation of solar lentigo.

BACKGROUND: Cutaneous pigmentation is regulated by a complex melanogenic network in which both keratinocytes and fibroblasts synthesize growth factors and cytokines. Solar lentigo (SL) is characterized by hyperpigmented lesions occurring on photodamaged skin areas. Despite the association of SL to ultraviolet (UV) exposure, the mechanisms underlying the development of these spots are not completely defined. OBJECTIVES: To analyse the involvement of the fibroblast-derived growth factors, hepatocyte growth factor (HGF), keratinocyte growth factor (KGF) and stem cell factor (SCF) in SL hyperpigmentation; to evaluate whether the photoageing process occurring in fibroblasts could be responsible for the altered expression of these cytokines; and to investigate a new possible role of KGF in regulating pigmentation through the specific induction of melanogenic cytokines by keratinocytes. METHODS: We performed immunohistochemical analysis of HGF, KGF and SCF on SL biopsies. We analysed the mRNA expression of these cytokines using an in vitro model of photoageing induced on fibroblasts. Finally, we evaluated the effects of KGF on the expression of melanogenic cytokines at the mRNA and protein levels on keratinocytes. RESULTS: We found positive staining for HGF, KGF and SCF in the upper dermis of SL lesions and a significant induction of the three cytokines in photoaged fibroblasts. We also demonstrated the contribution of KGF to pigmentation, showing its ability specifically to modulate the expression of SCF in keratinocytes. CONCLUSIONS: Fibroblasts may be persistently activated by UV exposure to release melanogenic growth factors; this inducible cytokine network acts both directly and indirectly through keratinocytes and may contribute to the hyperpigmentation of SL.