Air pollution-induced tanning of human skin.

BACKGROUND: Melanism is more frequent in animals living in polluted areas on urban-industrial sites. Given that an increasing number of people are exposed to elevated air pollution levels, it is possible that environmental pollutants affect melanogenesis in human skin. Epidemiological studies have shown that exposure to traffic-related air pollutants such as diesel exhaust particles (DEP) is associated with more clinical signs of hyperpigmentation. However, mechanistic evidence linking DEP exposure to pigmentation has been elusive. OBJECTIVES: To develop an ex␣vivo skin model to allow for repetitive topical application of relevant ambient DEP, and to provide proof of concept in humans. METHODS: We measured skin pigmentation, melanin and pigmentation-associated gene expression, and evaluated oxidative stress. RESULTS: Repetitive exposure of ex␣vivo skin to DEP at nontoxic concentrations increased skin pigmentation. This increase was visible to the naked eye, time dependent, and associated with an increase in melanin content and the transcription of genes involved in de novo melanin synthesis. Similarly, in healthy participants (n = 76), repetitive topical application of DEP at nontoxic concentrations increased skin pigmentation. DEP-induced pigmentation was mediated by an oxidative stress response. After the application of DEP, epidermal antioxidants were depleted, lipid peroxidation and oxidative DNA damage were enhanced, and in a vehicle-controlled, double-blind clinical study DEP-induced pigmentation was prevented by the topical application of an antioxidant mixture. CONCLUSIONS: Similar to solar radiation, air pollutants cause skin tanning. As eumelanin is an antioxidant, it is proposed that this response serves to protect human skin against air pollution-induced oxidative stress.

Quantitative ion determination in eccrine sweat gland cells correlates to sweat reduction of antiperspirant actives.

OBJECTIVE: Axillary wetness represents an unwanted effect of the physiologically vital sweating mechanism, especially when it becomes excessive. Cosmetic products reducing sweat secretion rely on aluminium salts as the active ingredient acting by physically blocking the sweat gland. Driven by the interest to better understand the sweat mechanism and to develop alternative technologies against excessive sweating a search for an effective testing approach started as up to now, cost- and time-consuming in vivo studies represent the standard procedure for testing and identifying these alternatives. MATERIAL AND METHODS: The herein described in vitro test system is based on the measurement of intracellular changes of the ion equilibrium in cultured eccrine sweat gland cells. Subsequently, in vivo studies on the back of volunteers were conducted to verify the sweat-reducing effect of in vitro newly discovered substance. RESULTS: In this study, we describe an effective cell-based in vitro method as a potent tool for a more targeted screening of alternatives to aluminium salts. Testing the commonly used aluminium chlorohydrate as one example of an aluminium-based active in this screening procedure, we discovered a distinct influence on the ion equilibrium: Intracellular levels of sodium ions were decreased while those of chloride increased. Screening of various substances revealed a polyethyleneimine, adjusted to pH 3.5 with hydrochloric acid, to evoke the same alterations in the ion equilibrium as aluminium chlorohydrate. Subsequent in vivo studies showed its substantial antiperspirant action and confirmed the high efficiency of the polyethyleneimine solution in vivo. Further, specific investigations connecting the chloride content of the tested substances with the resulting sweat reduction pointed towards a substantial impact of the chloride ions on sweating. CONCLUSION: The newly described in vitro cell-based screening method represents an effective means for identifying new antiperspirant actives and suggests an additional biological mechanism of action of sweat-reducing ingredients which is directed towards unbalancing of the ion equilibrium inside eccrine sweat gland cells.

OBJECTIF: l'humidité axillaire représente un effet indésirable du mécanisme physiologiquement vital de la sudation, en particulier lorsqu'elle devient excessive. Les produits cosmétiques réduisant la sécrétion de sueur reposent sur les sels d'aluminium comme principe actif agissant en bloquant physiquement la glande sudoripare. Motivée par l'intérêt de mieux comprendre le mécanisme de la sudation et de développer des technologies alternatives contre l'hypersudation, une recherche pour une approche de test efficace a commencé car, jusqu'à présent, les études in vivo coûteuses et chronophages représentent la procédure standard pour tester et identifier ces alternatives. MATÉRIELS ET MÉTHODES: le système de test in vitro décrit ici est basé sur la mesure des changements intracellulaires de l'équilibre ionique dans les cellules des glandes sudoripares exocrines cultivées. Par la suite, des études in vivo sur le dos de volontaires ont été menées pour vérifier l'effet réducteur de la sudation d'une substance nouvellement découverte in vitro. RÉSULTATS: dans cette étude, nous décrivons une méthode cellulaire efficace in vitro en tant qu'outil puissant pour un dépistage plus ciblé des alternatives aux sels d'aluminium. En testant le chlorohydrate d'aluminium couramment utilisé comme exemple d'un principe actif à base d'aluminium dans cette procédure de dépistage, nous avons découvert une influence distincte sur l'équilibre ionique : les taux intracellulaires d'ions sodium ont diminué tandis que ceux du chlorure ont augmenté. La recherche de diverses substances a révélé une polyéthylèneimine, ajustée au pH 3,5 avec de l'acide chlorhydrique, pour évoquer les mêmes altérations de l'équilibre ionique que le chlorohydrate d'aluminium. Des études in vivo ultérieures ont montré son action anti-transpirante substantielle et ont confirmé la haute efficacité de la solution de polyéthylèneimine in vivo. De plus, des études spécifiques établissant un lien entre la teneur en chlorure des substances testées et la réduction de la sudation qui en résulte ont indiqué que les ions chlorure ont un impact substantiel sur l'hypersudation. CONCLUSION: la nouvelle méthode de dépistage cellulaire in vitro décrite représente un moyen efficace d'identifier de nouveaux agents anti-transpirants actifs et suggère un mécanisme d'action biologique supplémentaire des ingrédients réducteurs de la sudation, dirigé vers le déséquilibre de l'équilibre ionique à l'intérieur des cellules des glandes sudoripares exocrines.

Theophylline exerts complex anti-ageing and anti-cytotoxicity effects in human skin ex vivo.

OBJECTIVE: Theophylline is a phosphodiesterase inhibitor that is being used clinically for asthma therapy. In addition, it is recognized as a cosmetic agent with possible anti-ageing and anti-oxidative properties. Nevertheless, how it affects human skin is still poorly examined. METHODS: Theophylline (10 or 100 µM) was administered to the culture medium of full-thickness human skin ex vivo for 24 or 72 h. RESULTS: Theophylline stimulated protein expression of the anti-oxidant metallothionein-1 and mRNA levels of collagen I and III. Assessment of fibrillin-1 immunohistology revealed enhanced structural stability of dermal microfibrils. Theophylline also exerted extracellular matrix-protective effects by decreasing MMP-2 and MMP-9 mRNA levels, partially antagonizing the effects of menadione, the potent, toxic ROS donor. In addition, it decreased menadione-stimulated epidermal keratinocytes apoptosis. Interestingly, theophylline also increased the level of intracutaneously produced melatonin, that is the most potent ROS-protective and DNA damage repair neuromediator, and tendentially increased protein expression of MT1, the melatonin receptor. Theophylline also increased the expression of keratin 15, the stem cell marker, in the epidermal basal layer but did not change mitochondrial activity or epidermal pigmentation. CONCLUSION: This ex vivo pilot study in human skin shows that theophylline possesses several interesting complex skin-protective properties. It encourages further examination of theophylline as a topical candidate for anti-ageing treatment.

OBJECTIF: la théophylline est un inhibiteur de la phosphodiestérase actuellement utilisée en clinique pour le traitement de l'asthme. En outre, elle est reconnue comme étant un agent cosmétique ayant des propriétés potentiellement anti-âge et antioxydantes. Cependant, la manière dont elle affecte la peau chez l'homme est encore très peu étudiée. MÉTHODES: de la théophylline (10 ou 100 µM) a été ajoutée dans le milieu de culture d'un échantillon de peau humaine d'épaisseur totale ex vivo pendant 24 ou 72 h. RÉSULTATS: la théophylline a stimulé l'expression de la métallothionéine-1, une protéine antioxydante, et les taux d'ARNm du collagène I et III. L'évaluation immunohistologique de la fibrilline-1 a révélé une meilleure stabilité structurale des microfibrilles du derme. La théophylline a également exercé des effets protecteurs sur la matrice extracellulaire en diminuant les taux d'ARNm des métalloprotéinases matricielles MMP-2 et MMP-9, neutralisant en partie les effets de la ménadione, puissant donneur d'espèces réactives de l'oxygène (ROS) toxiques. En outre, elle a diminué l'apoptose des kératinocytes épidermiques stimulés par la ménadione. Fait intéressant, la théophylline a également augmenté le taux de mélatonine produite de manière intra-cutanée, la mélatonine étant le plus puissant neuromédiateur protecteur contre les ROS et réparateur des lésions de l'ADN. Elle a augmenté de façon tendancielle l'expression de la protéine MT1, récepteur de la mélatonine. La théophylline a également augmenté l'expression de la kératine 15, marqueur de cellules souches, dans la couche basale épidermique, mais n'a pas modifié l'activité mitochondriale ou la pigmentation épidermique. CONCLUSION: cette étude pilote ex vivo réalisée sur de la peau humaine montre que la théophylline a plusieurs propriétés protectrices de la peau complexes et intéressantes. Ces résultats encouragent à poursuivre l'étude de la théophylline en tant que candidat à un traitement local anti-âge.

Cloning and characterization of hurpin (protease inhibitor 13): A new skin-specific, UV-repressible serine proteinase inhibitor of the ovalbumin serpin family.

Epidermal keratinocytes are the primary target of the midrange ultraviolet part (UVB, 280-320 nm) of terrestrial sunlight. Analysis of the resulting UV response at the transcriptional level by differential display PCR identified a formerly unrecognized large group of repressed genes. Among those UV-repressible genes, a novel serine proteinase inhibitor (serpin) termed hurpin (HaCaT UV-repressible serpin) has been identified. The isolated full-length cDNAs harbour a 1176 bp open reading frame encoding a potential protein with 391 amino acid residues and a predicted molecular mass of approximately 44 kDa. The novel serpin has nearly 59 % amino acid identity with the squamous cell carcinoma antigen 1 (SCCA1) and squamous cell carcinoma antigen 2 (SCCA2). In addition, it displays all of the structural features unique to the ovalbumin family of serpins (ov-serpins). The amino acid sequence of the hinge region in the reactive site loop suggests that hurpin has the potential for protease inhibition. The putative reactive center P1-P1'residues were identified as Thr356-Ser357 by alignment with other ov-serpins. The physiological target protease is unknown and the in vitro translated hurpin does not form SDS-stable complexes with a variety of known serine proteases. Expression of hurpin is restricted to epidermal cells where two distinct transcripts of 3.0 and 3.4 kb are detectable. Furthermore, expression of hurpin appears to be related to the activation or proliferation state of keratinocytes, since hurpin transcripts are more abundant in immortalized keratinocytes (HaCaT) and in cultured normal human keratinocytes, compared to the expression in normal skin. Moreover, in psoriasis, a skin disease characterized by hyperproliferation of keratinocytes and responsive to therapeutic UV irradiation, overexpression of hurpin is noted in psoriatic skin lesions compared to non-lesional skin.

Sequence, organization, chromosomal localization, and alternative splicing of the human serine protease inhibitor gene hurpin (PI13) which is upregulated in psoriasis.

Hurpin (protease inhibitor 13; PI13) is the most recently identified member of the ovalbumin family of serine protease inhibitors (serpins). It is expressed in human epidermal keratinocytes and is downregulated by exposure to ultraviolet irradiation. A role for hurpin in the proliferation or differentiation of keratinocytes has been proposed because of its strong expression in proliferating cells and its deregulated expression in the lesional epidermis of psoriatic patients. Here, we report the cloning, chromosomal localization, and complete sequence of the human hurpin gene. By PCR-based screening of the GeneBridge 4 radiation hybrid panel, we mapped the gene to chromosome 18q21.3, close to a known cluster of ov-serpin genes. Using the full-length cDNA for hurpin, we identified two clones from an arrayed genomic P1 placental library that contain the entire hurpin gene. Sequencing revealed that the gene covers 12.253 kb and is comprised of eight exons and seven introns. The exon--intron boundaries are identical in position and phasing to those in other members of the 18q serpin gene cluster, and analysis of hurpin variants indicated that modified functional inhibitors, differing only in the CD interhelical loop, can be generated by differential splicing of exon 3. These data show that hurpin is a typical member of the 18q ovalbumin-serpins most closely related to the serpins squamous-cell carcinoma antigens 1 and 2.