Endothelial Cells Promote Pigmentation through Endothelin Receptor B Activation.

Findings of increased vascularization in melasma lesions and hyperpigmentation in acquired bilateral telangiectatic macules suggested a link between pigmentation and vascularization. Using high-magnification digital epiluminescence dermatoscopy, laser confocal microscopy, and histological examination, we showed that benign vascular lesions of the skin have restricted but significant hyperpigmentation compared with the surrounding skin. We then studied the role of microvascular endothelial cells in regulating skin pigmentation using an in vitro co-culture model using endothelial cells and melanocytes. These experiments showed that endothelin 1 released by microvascular endothelial cells induces increased melanogenesis signaling, characterized by microphthalmia-associated transcription factor phosphorylation, and increased tyrosinase and dopachrome tautomerase levels. Immunostaining for endothelin 1 in vascular lesions confirmed the increased expression on the basal layer of the epidermis above small vessels compared with perilesional skin. Endothelin acts through the activation of endothelin receptor B and the mitogen-activated protein kinase, extracellular signal-regulated kinase (ERK)1/2, and p38, to induce melanogenesis. Finally, culturing of reconstructed skin with microvascular endothelial cells led to increased skin pigmentation that could be prevented by inhibiting EDNRB. Taken together these results demonstrated the role of underlying microvascularization in skin pigmentation, a finding that could open new fields of research for regulating physiological pigmentation and for treating pigmentation disorders such as melasma.

Identification of Genes Expressed in Hyperpigmented Skin Using Meta-Analysis of Microarray Data Sets.

More than 375 genes have been identified that are involved in regulating skin pigmentation and these act during development, survival, differentiation, and/or responses of melanocytes to the environment. Many of these genes have been cloned, and disruptions of their functions are associated with various pigmentary diseases; however, many remain to be identified. We have performed a series of microarray analyses of hyperpigmented compared with less pigmented skin to identify genes responsible for these differences. The rationale and goal for this study was to perform a meta-analysis on these microarray databases to identify genes that may be significantly involved in regulating skin phenotype either directly or indirectly that might not have been identified due to subtle differences by any of these individual studies alone. The meta-analysis demonstrates that 1,271 probes representing 921 genes are differentially expressed at significant levels in the 5 microarray data sets compared, providing new insights into the variety of genes involved in determining skin phenotype. Immunohistochemistry was used to validate two of these markers at the protein level (TRIM63 and QPCT), and we discuss the possible functions of these genes in regulating skin physiology.

UV exposure modulates hemidesmosome plasticity, contributing to long-term pigmentation in human skin.

Human skin colour, ie pigmentation, differs widely among individuals, as do their responses to various types of ultraviolet radiation (UV) and their risks of skin cancer. In some individuals, UV-induced pigmentation persists for months to years in a phenomenon termed long-lasting pigmentation (LLP). It is unclear whether LLP is an indicator of potential risk for skin cancer. LLP seems to have similar features to other forms of hyperpigmentation, eg solar lentigines or age spots, which are clinical markers of photodamage and risk factors for precancerous lesions. To investigate what UV-induced molecular changes may persist in individuals with LLP, clinical specimens from non-sunburn-inducing repeated UV exposures (UVA, UVB or UVA + UVB) at 4 months post-exposure (short-term LLP) were evaluated by microarray analysis and dataset mining. Validated targets were further evaluated in clinical specimens from six healthy individuals (three LLP+ and three LLP-) followed for more than 9 months (long-term LLP) who initially received a single sunburn-inducing UVA + UVB exposure. The results support a UV-induced hyperpigmentation model in which basal keratinocytes have an impaired ability to remove melanin that leads to a compensatory mechanism by neighbouring keratinocytes with increased proliferative capacity to maintain skin homeostasis. The attenuated expression of SOX7 and other hemidesmosomal components (integrin α6ß4 and plectin) leads to increased melanosome uptake by keratinocytes and points to a spatial regulation within the epidermis. The reduced density of hemidesmosomes provides supporting evidence for plasticity at the epidermal-dermal junction. Altered hemidesmosome plasticity, and the sustained nature of LLP, may be mediated by the role of SOX7 in basal keratinocytes. The long-term sustained subtle changes detected are modest, but sufficient to create dramatic visual differences in skin colour. These results suggest that the hyperpigmentation phenomenon leading to increased interdigitation develops in order to maintain normal skin homeostasis in individuals with LLP.

Photobiological implications of melanin photoprotection after UVB-induced tanning of human skin but not UVA-induced tanning.

Repetitive suberythemal UVA and/or UVB exposures were used to generate comparable UV-induced tans in human skin over the course of 2 weeks. To evaluate the potential photoprotective values of those UVA- and/or UVB- induced tans and to avoid the confounding issue of residual UV-induced DNA damage, we waited 1 week before challenging those areas with a 1.5 MED of UVA+UVB after which we measure DNA damage. The results show that the type of UV used to induce skin pigmentation affects the redistribution of melanin in the skin and/or de novo melanin synthesis. The UVA-induced tans failed to even provide a minimal SPF of 1.5, which suggests that producing a tan with UVA-rich sunlamps prior to a holiday or vacation is completely counterproductive.

Epidermal gene expression and ethnic pigmentation variations among individuals of Asian, European and African ancestry.

Differences in visible skin pigmentation give rise to the wide variation of skin colours seen in racial/ethnic populations. Skin pigmentation is important not only from cosmetic and psychological points of view, but more importantly because of its implications for the risk of all types of skin cancers, on photoaging, etc. Despite differences in those parameters in Caucasian and Asian skin types, they are remarkably similar in their production and distribution of melanins, and the mechanism(s) underlying their different characteristics have remained obscure. In this study, we used microarray analysis of skin suction blisters to investigate molecular differences underlying the determination of pigmentation in various skin types, and we used immunohistochemistry to validate the expression patterns of several interesting targets that were identified. Intriguingly, Caucasian and Asian skins had highly similar gene expression patterns that differed significantly from the pattern of African skin. The results of this study suggest the dynamic interactions of different types of cells in human skin that regulate its pigmentation, reveal that the known pigmentation genes have a limited contribution and uncover a new array of genes, including NINL and S100A4, that might be involved in that regulation.

Extra-hepatic cancer represses hepatic drug metabolism via interleukin (IL)-6 signalling.

PURPOSE: In many cancer patients, the malignancy causes reduced hepatic drug clearance leading to potentially serious complications from the use of anticancer drugs. The mechanisms underlying this phenomenon are poorly understood. We aimed to identify tumor-associated inflammatory pathways that alter drug response and enhance chemotherapy-associated toxicity. METHODS: We studied inflammatory pathways involved in extra-hepatic tumor mediated repression of CYP3A, a major hepatic drug metabolizing cytochrome P450 subfamily, using a murine Engelbreth-Holm-Swarm sarcoma model. Studies in IL-6 knockout mice determined the source of elevated IL-6 in tumor-bearing animals and monoclonal antibodies against IL-6 were used to intervene in this inflammatory pathway. RESULTS: Our studies confirm elevated plasma IL-6 levels and reveal activation of Jak/Stat and Mapk signalling pathways and acute phase proteins in livers of tumor-bearing mice. Circulating IL-6 was predominantly produced by the tumor xenograft, rather than being host derived. Anti IL-6 antibody intervention partially reversed tumor-mediated inflammation and Cyp3a gene repression. CONCLUSIONS: IL-6 is an important player in cancer-related repression of CYP3A-mediated drug metabolism and activation of the acute phase response. Targeting IL-6 in cancer patients may prove an effective approach to alleviating cancer-related phenomena, such as adverse drug-related outcomes commonly associated with cancer chemotherapy.

Pregnane X receptor: promiscuous regulator of detoxification pathways.

The Pregnane X Receptor (PXR) is pivotal for the body's response to toxic xenobiotics and endogenous metabolites. By acting as a ligand-activated transcription factor, PXR regulates all stages of xenobiotic metabolism and transport and is responsible for important inductive drug interactions. Screening assays to assess the PXR activation potential of new and existing drugs are becoming integral components of drug discovery programs. PXR is also involved in lipid homeostasis providing opportunities for treatments based on PXR agonists for diseases involving aberrant cholesterol and bile acid levels. The expression of PXR in many other tissues besides liver and intestine suggest PXR may have additional protective functions in the body, which contribute to disease outcomes in diverse clinical situations with potential for novel therapeutic approaches.

Contribution of UVB and UVA to UV-dependent stimulation of cyclooxygenase-2 expression in artificial epidermis.

Both UVB (280-320 nm) and UVA (320-400 nm) radiation lead to an enhanced expression of cyclooxygenase-2 (COX-2) in epidermal cells in various in-vitro and in-vivo models. It is demonstrated here that the expression of COX-2 is induced in artificial human epidermis exposed to simulated solar light (>290 nm). Employing filters eliminating specified regions from the simulated solar spectrum, the UVB and UVA-2 (320-350 nm) regions are shown to fully account for induction of COX-2 mRNA and protein as well as the enhanced production of prostaglandin E(2) after irradiation. At the protein level, approximately 70% of the total induction by solar light is due to light in the UVA-2 region. UVA-1 (350-400 nm), visible light and IR radiation are practically ineffective. COX-2 induction by simulated solar light is attenuated in the presence of inhibitors of p38(MAPK) or of c-Jun-N-terminal kinases (JNK), whereas COX-2 induction by UVA is blocked only by inhibition of JNK. UV-induced COX-2 expression is not affected by inhibition of the MEK 1,2/ERK 1,2 pathways.

Irradiation of cells with ultraviolet-A (320-400 nm) in the presence of cell culture medium elicits biological effects due to extracellular generation of hydrogen peroxide.

Biological effects of ultraviolet A (UVA) irradiation have been ascribed to the photochemical generation of singlet oxygen. Not all effects described in the literature, however, are explicable solely by the generation of singlet oxygen, but rather resemble effects elicited by hydrogen peroxide (H2O2). Here, we show that when cells are kept in cell culture media during exposure to UVA, stress kinases, including ERK 1 and ERK 2 as well as Akt (protein kinase B), are activated, whereas there is no or only minor activation when cells are kept in phosphate-buffered saline during irradiation. Indeed, the exposure of cell culture media to UVA (30 J/cm2) results in the generation of significant amounts of H2O2, with concentrations of about 100 microM. H2O2 concentrations are at least three-fold higher in HEPES-buffered culture media after UVA irradiation. From experiments with solutions of riboflavin, tryptophan or HEPES, as well as combinations thereof, it is concluded that riboflavin mediates the photooxidation of either tryptophan or HEPES, resulting in the generation of H2O2. Thus, if signaling effects of UVA radiation are to be investigated in cell culture systems, riboflavin and HEPES/tryptophan should be avoided during irradiation because of artificial H2O2 generation. It should be taken into account, however, that in vivo tryptophan and riboflavin might play an important role in the generation of reactive oxygen species by UVA as both substances are abundant in living tissues.