Similar UV responses are seen in a skin organ culture as in human skin in vivo

Ultraviolet radiation (UVR) plays an important role in the development of non-melanoma skin cancer. Most tumors develop in chronically sun-exposed skin, most often in cosmetically sensitive locations, where in vivo experiments may be difficult to perform. In this study, we describe a skin organ culture model with preserved normal morphology and intact response to UVR. Skin explants from chronically sun-exposed and non-sun-exposed skin were irradiated with artificial UVA+UVB with and without topical sunscreen. UV-induced DNA damage, epidermal p53 response and repair kinetics were analyzed using immunohistochemistry. Four hours after UV-irradiation epidermal keratinocytes showed a strong immunoreactivity for thymine-dimers. Gradual repair during an incubation time resulted in few residual thymine-dimers after 48 h. Repair appeared to be more efficient in chronically sun-exposed skin compared with non-sun-exposed skin. There was also an accumulation of p53 protein in epidermal keratinocytes, peaking at 4-24 h after irradiation. Large interindividual differences with respect to formation and repair of thymine-dimers as well as induction and duration of the p53 response were observed. Skin explants treated with topical sunscreen prior to UV-irradiation showed a clear reduction of thymine-dimers and p53 expression. The epidermal UV-responses and repair kinetics in organ-cultured skin were similar to what was found in vivo. Our data suggest that organ-cultured skin provides a valuable tool for studies of UV-induced epidermal responses in chronically sun-exposed skin.     

Peak blistering point: influence on fluid levels of 5-MOP in human skin in vivo after systemic administration

Summary The concentration of 5-methoxypsoralen (5-MOP) in suction blister fluid (SBF) after oral intake was determined in relation to the peak blistering point. Interstitial fluid was obtained from nine healthy male volunteers by applying mild suction (300–350 mmHg) to the skin of the volar aspect of the forearm. Blisters were raised at three different times: 18 h prior to drug administration (group I); 2 h prior to drug administration (group II); and during drug ingestion (group III). SBF levels of 5-MOP were determined in each group of blisters 2 h after oral administration of 1.2 mg/kg micronized 5-MOP. The results showed a statistically significant difference in the concentrations of 5-MOP between each group of blisters. The highest concentration of 5-MOP in the SBF was found in group III and the lowest in group II blisters. These findings suggest that the point of peak blistering in relation to drug administration needs to be known in each pharmacological study using suction blisters.     

The physiological and phenotypic determinants of human tanning measured as change in skin colour following a single dose of ultraviolet B radiation

Please cite this paper as: The physiological and phenotypic determinants of human tanning measured as change in skin colour following a single dose of ultraviolet B radiation. Experimental Dermatology 2010; 19 : 667–673. Abstract: Experimental study of the in vivo kinetics of tanning in human skin has been limited by the difficulties in measuring changes in melanin pigmentation independent of the ultraviolet‐induced changes in erythema. The present study attempted to experimentally circumvent this issue. We have studied erythemal and tanning responses following a single exposure to a range of doses of ultraviolet B irradiation on the buttock and the lower back in 98 subjects. Erythema was assessed using reflectance techniques at 24 h and tanning measured as the L* spectrophotometric score at 7 days following noradrenaline iontophoresis. We show that dose (P < 0.0001), body site (P < 0.0001), skin colour (P < 0.0001), ancestry (P = 0.0074), phototype (P = 0.0019) and sex (P = 0.04) are all independent predictors of erythema. Quantitative estimates of the effects of these variables are reported, but the effects of ancestry and phototype do not appear solely explainable in terms of L* score. Dose (P < 0.0001), body site (P < 0.0001) and skin colour (P = 0.0365) or, as an alternative to skin colour, skin type (P = 0.0193) predict tanning, with those with lighter skin tanning slightly more to a defined UVB dose. If erythema is factored into the regression, then only dose and body site remain significant predictors of tanning: therefore neither phototype nor pigmentary factors, such as baseline skin colour, or eye or hair colour, predict change in skin colour to a unit erythemal response.     

Change in ultraviolet (UV) transmission following the application of vaseline to non-irradiated and UVB-exposed split skin

BACKGROUND: Topical preparations such as emollients used in combination with phototherapy can interfere with such treatment. OBJECTIVES: This study was performed to investigate the impact of vaseline on the ultraviolet (UV) transmission of non-irradiated split skin and on split skin previously exposed to UVB radiation. METHODS: Split-skin specimens were obtained from 20 patients. In each case, one sample was taken from an area of non-irradiated skin, while the second was taken from an area that had been previously exposed to UVB. The transmission was spectrophotometrically measured with split skin placed in specially designed quartz glass cuvettes before and after the application of two different amounts of vaseline (2.5 and 17.5 mg cm-2). RESULTS: Application of vaseline to skin previously exposed to UVB caused significant (P < 0.0001) changes in UV transmission in certain wavelength ranges. In the UVA range, a greater increase in transmission was achieved with 2.5 mg cm-2 vaseline, whereas in the UVB range, a greater increase was achieved with 17.5 mg cm-2 vaseline. The thicker the layer of vaseline applied, the lower was the difference in transmission between non-irradiated split skin and UVB-exposed split skin. CONCLUSIONS: Application of the correct amount of vaseline can enhance transmission in either the UVA or UVB range, and would enable dose reduction during a course of phototherapy.     

UVA‐1 exposure in vivo leads to an IL‐6 surge within the skin

UVA‐1 is a known promotor of skin ageing. Cytokines like IL‐1α, Il‐1β or TNF‐α, VEGF and IL‐6 orchestrate UV effects, and IL‐6 is furthermore an effector of UVA‐induced photoageing. We investigated how fractionated UVA‐1 doses influence the cytokine milieu and especially the IL‐6 levels in the skin in vivo. In a study with 35 participants, we exposed previously unirradiated human skin to three UVA‐1 irradiation regimes. Cytokine levels in interstitial skin fluid were measured up to 48 hours postexposure and compared to unirradiated control skin fluid. Our results show that IL‐6 levels increased significantly after UVA‐1 exposure at selected time points. The other candidates IL‐1α, Il‐1β or TNF‐α and VEGF show no significant response after UVA‐1 exposure in vivo. UVA‐1 thus raises selectively IL‐6 levels in vivo, a fact that underlines its role in photoageing and has potential implications for its modulatory effect on photoageing pathology.     

Photodamage to human skin by suberythemal exposure to solar ultraviolet radiation can be attenuated by sunscreens: a review

The effects of acute or repeated suberythemal solar ultraviolet radiation (UVR) exposure on human skin have been insufficiently investigated. Such exposure almost certainly has important long-term consequences that include skin ageing and skin cancer. This review summarizes the published data on the biological effects of suberythemal exposure using a wide range of clinical, cellular and molecular endpoints, some of which may be considered as biomarkers for skin cancer and photoageing. We also include some recent unpublished results from our laboratories. The effects of UVA (320-400 nm), UVB (290-320 nm) and total solar UVR (290-400 nm) are compared. We demonstrate that avoiding sunburn does not prevent many indicators of cutaneous biological damage and that use of low sun protection factor (SPF) sunscreen can inhibit much of the damages induced by suberythemal exposure to UVR. However, even when applied correctly, sunscreen use will result in suberythemal exposure. The degree and spectral quality of such exposure will depend on the SPF and absorption spectrum of the sunscreen, but nonetheless it may contribute to cumulative photodamage. This review may help to determine the level of photoprotection required in sunscreens and daily use products, as well as the ideal ratio of UVB/UVA protection, to improve long-term photoprotection outcomes.     

Effects of PUVA and UVB treatments on restoration of epidermal barrier function and vascular response after suction blister injury in human skin in vivo

PUVA and UVB phototherapies are used in the treatment of psoriasis and other inflammatory skin diseases. Ultraviolet radiation causes inflammation and modulates cell kinetics in the skin. PUVA also has an inhibitory effect on skin DNA synthesis. In this study, the effects of PUVA and UVB treatments on epidermal wound healing were examined using the suction blister wound model. The healing of the wound was studied indirectly by measuring water evaporation and blood flow in the wound area. On the fourth day, water evaporation was more abundant in PUVA-treated patients (42±5 g/m²h) than in UVB treated (36±4 g/m²h) or control patients (27±3 g/m²h) (analysis of variance, the least significant difference test at a level of 0.05). The P value for the difference of means between the PUVA and control groups was 0.014. Blood flow was also more abundant during the fourth (PUVA 162±11 arbitrary units, UVB 122±10, controls 115±15) and sixth (PUVA 108±18, UVB 73±17, controls 57±13) day in PUVA treated patients (analysis of variance, the least significant difference test at a level of 0.05). The results suggest that PUVA treatment decreases the restoration of the epidermal barrier function. The PUVA-treated patients also showed a more intense and prolonged vascular response that may be due to PUVA-related inflammation.     

Effect of photoreactivating light on UV radiation-induced alterations in human skin

BACKGROUND/AIMS: Photoreactivating light (PRL) after ultraviolet radiation (UVR) exposure causes photoreversal of cyclobutane pyrimidine dimers through the activation of photolyase. Although photoreversal has been demonstrated in the "three kingdoms of life," its existence in man remains controversial. We sought evidence for photoreversal in man. METHODS AND RESULTS: Seven subjects were spot-irradiated at two sites with 4 minimal erythema doses (MED) of solar-simulating UVR. Of the two sites, one was then immediately exposed to a PRL source. Epidermal biopsies were taken immediately after exposure. No significant difference in the quantity of pyrimidine dimers was detected comparing the "UVR only" site to the "UVR, PRL-exposed" site. Biopsies were repeated 24 h later and no significant difference in p53 protein expression or dendritic cell number was detected. However, the "UVR, PRL-exposed" site showed a greater reduction in pyrimidine dimer quantity. CONCLUSIONS: We found no evidence for a direct effect of PRL causing photoreversal of UVR-induced pyrimidine dimers in man. Our results do, however, suggest that some indirect effect of PRL may enhance pyrimidine dimer repair in the 24-h period following UVR exposure.     

Transmission of UVA radiation through epithelium of oral mucosa and skin in rat and man

BACKGROUND/METHODS: Transmission of 362 nm ultraviolet A radiation (UVA) through epithelium of oral mucosa and skin epidermis was compared in rat and man. The transmitted irradiance was measured by means of a radiometer. RESULTS: In man, significantly (P<0.01) greater transmittance (2.8-fold) was observed in skin compared to buccal mucosa. In rat, no significant differences were found. CONCLUSION: In different species and different types of epitheliae, separate studies are needed for the evaluation of UVA transmission.     

Novel TGM5 mutations in acral peeling skin syndrome

Acral peeling skin syndrome (APSS, MIM #609796) is a rare autosomal recessive disorder characterized by superficial exfoliation and blistering of the volar and dorsal aspects of hands and feet. The level of separation is at the junction of the stratum granulosum and stratum corneum. APSS is caused by mutations in the TGM5 gene encoding transglutaminase‐5, which is important for structural integrity of the outermost epidermal layers. The majority of patients originate from Europe and carry a p.(Gly113Cys) mutation in TGM5. In this study, we report both European and non‐European families carrying other mutations in the TGM5 gene. In 5 patients, we found 3 novel mutations: c.1001+2_1001+3del, c.1171G>A and c.1498C>T. To confirm their pathogenicity, we performed functional analyses with a transglutaminase activity assay, determined alternative splicing by reverse‐transcribed PCR analysis and used databases and in silico prediction tools.     

Acute exposure of human skin to ultraviolet or infrared radiation or heat stimuli increases mast cell numbers and tryptase expression in human skin in vivo

Background  Mast cells are key effector cells in diverse immunological and pathological processes. It is still unclear why there are more mast cells at peripheral and sun-exposed skin sites than at sun-protected sites.
Objectives  To investigate changes in mast cell numbers associated with natural ageing and photoageing, and to observe the effects of ultraviolet (UV) and infrared (IR) radiation and heat on the prevalence of mast cells and tryptase expression in human skin in vivo .
Methods  Sun-exposed and sun-protected skin samples were taken from individuals in four different age groups. UV, IR or heat-treated buttock skin of young volunteers was also obtained. Mast cells were quantified by immunohistochemical staining of mast cell-specific tryptase and chymase. The expression of tryptase was determined by Western blotting.
Results  Both sun-exposed and sun-protected skin showed a gradual decrease in total mast cells (MC_Total) number with ageing. The number of mast cells in sun-exposed skin was significantly higher than that in sun-protected skin. After UV irradiation (2 minimal erythema doses), MC_Total and mast cells expressing tryptase and chymase were significantly increased at 24 and 48 h postirradiation. After IR irradiation (3 minimal heating doses) and heat treatment (43 °C for 90 min), MC_Total reached peak induction at 8 and 48 h after stimulation, respectively. Tryptase expression was also clearly upregulated by UV, IR and heat.
Conclusions  Our data demonstrate that mast cell numbers decreased with ageing in human skin. Also, mast cells may be activated and recruited by UV, IR and heat. These findings should further our understanding of the reason for the high prevalence of mast cells at peripheral sun-exposed skin sites.     

Immunolocalization of aquaporin-5 in normal human skin and hypohidrotic skin diseases

Aquaporin (AQP)‐5 has been shown to be expressed in the secretory parts of mouse, rat and horse sweat glands. However, the precise localization of AQP‐5 in normal and diseased human skin has not been fully determined. The aim of the present study was to further clarify the immunolocalization of AQP‐5 in normal human skin and hypohidrotic skin diseases. Normal human scalp skin and biopsies from skin affected by hypohidrotic diseases were analyzed for AQP‐5 and/or dermcidin expression by immunohistochemistry, immunofluorescence and/or immunoelectronmicroscopy. AQP‐5 was expressed on the apical and basolateral plasma membranes of the clear cells in eccrine sweat coils, but not in ductal components or apocrine glands. Numbers of AQP‐5‐positive coils in the secretory part of eccrine sweat glands were decreased in Sjögren’s syndrome, but not in skin affected by idiopathic segmental anhidrosis or idiopathic pure sudomotor failure. AQP‐5 was mostly localized to the plasma membranes of clear cells in the secretory coils of eccrine sweat glands, suggesting that it plays a role in producing the primary sweat fluid.     

Presence of immunoreactive beta-endorphin in human skin

The production and its induction by ultraviolet radiation (UVR) of proopiomelanocortin (POMC)-derived peptides by keratinocytes has been reported, albeit not consistently. Recently we demonstrated that only under specific culturing conditions human keratinocytes are capable of producing a beta-endorphin (betaE)-like peptide with the characteristics of beta-lipotropin (betaLPH). Here the presence and UV-induction of betaE-immunoreactivity (betaE-IR) in keratinocytes in human skin in vivo was investigated. betaE-IR was detectable by immunohistochemistry in keratinocytes of the follicular matrix and to some extent in cells of sweat ducts, but was absent from epidermal keratinocytes. Absence of betaE-IR was confirmed by radioimmunoassay of HPLC-fractionated extracts of normal epidermis. Repeated exposure to solar-simulated UVR had no effect. This investigation is the first to demonstrate the presence of betaE-immunoreactive material in the follicular matrix of corporal hairs and in duct cells of sweat glands. The possible meaning of these results is discussed.     

Photoprotection of human skin beyond ultraviolet radiation

Photoprotection of human skin by means of sunscreens or daily skin‐care products is traditionally centered around the prevention of acute (e.g. sunburn) and chronic (e.g. skin cancer and photoaging) skin damage that may result from exposure to ultraviolet rays (UVB and UVA). Within the last decade, however, it has been appreciated that wavelengths beyond the ultraviolet spectrum, in particular visible light and infrared radiation, contribute to skin damage in general and photoaging of human skin in particular. As a consequence, attempts have been made to develop skin care/sunscreen products that not only protect against UVB or UVA radiation but provide photoprotection against visible light and infrared radiation as well. In this article, we will briefly review the current knowledge about the mechanisms responsible for visible light/infrared radiation‐induced skin damage and then, based on this information, discuss strategies that have been successfully used or may be employed in the future to achieve photoprotection of human skin beyond ultraviolet radiation. In this regard we will particularly focus on the use of topical antioxidants and the challenges that result from the task of showing their efficacy.     

Alterations in human epidermal Langerhans cells by ultraviolet radiation: quantitative and morphological study

BACKGROUND: Ultraviolet (UV) exposure of human skin induces local and systemic immune suppression. This phenomenon has been well documented when UVB radiation (290-320 nm) is used. The mechanism is thought to involve Langerhans cells (LCs), the epidermal dendritic cells that play a crucial role in antigen presentation. A variety of studies have clearly demonstrated that UVB radiation decreases LC density and alters their morphology and immunological function, but little is known about the effects of the entire UV spectrum (ultraviolet solar simulated radiation, UV-SSR or UVB + UVA) or UVA (320-400 nm) radiation alone. OBJECTIVES: The purpose of this study was to analyse and compare the effects of a single exposure of human volunteers to UV-SSR, total UVA or UVA1 (340-400 nm) in the human epidermal LC density and morphology. METHODS: Immunohistochemistry on epidermal sheets with various antibodies and transmission electron microscopy (TEM) were used. RESULTS: Immunostaining for class II antigen revealed that a single UV-SSR exposure, corresponding to twice the minimal erythemal dose (MED), induced a significant reduction in LC density with only slight morphological alterations of remaining cells. After a single UVA exposure, LC density showed a dose-dependent reduction with a significant effect at 60 J cm(-2) (well above the MED). Moreover, the reduction of LC dendricity was also dose-dependent and significant for doses exceeding 30 J cm(-2). UVA1 radiation was as effective as total UVA for the later endpoint. As demonstrated by TEM, the location of Birbeck granules containing epidermal cells was modified in UVA-exposed areas. They were located in the spinous rather than in the suprabasal layer. In addition, the morphology of these cells was altered. We observed a rounding up of the cell body with a reduction of dendricity. Alterations of mitochondrial membrane and ridges were also seen. CONCLUSIONS: A single exposure of human skin in vivo to UV-SSR, UVA or UVA1 radiation results in different alterations of density and/or morphology of LCs. All these alterations may impair the antigen-presenting function of LCs leading to an alteration of immune response.     

Expression of extracellular matrix protein 1 (ECM1) in human skin is decreased by age and increased upon ultraviolet exposure

BACKGROUND: The extracellular matrix protein 1 (ECM1) is expressed in human skin and plays an important role in its normal structure and function. In the rare genetic skin disease lipoid proteinosis, which is characterized by a loss-of-function mutation in the ECM1 gene, skin areas habitually exposed to the sun may show a more severely scarred and photoaged appearance. However, no data are available on the possible involvement of ECM1 expression in intrinsic and extrinsic skin ageing. OBJECTIVES: We hypothesized that ECM1 expression in human skin is regulated by age- and ultraviolet (UV)-dependent mechanisms. METHODS: Skin biopsies from 12 patients with histologically confirmed solar elastosis, from non-UV-exposed sites of 12 age-matched controls and 12 young subjects were analysed. To evaluate the influence of acute UV exposure, buttock skin of 10 healthy subjects was irradiated repetitively on 10 days with a solar simulator and compared intraindividually with non-UV-treated contralateral sites. The expression of ECM1 was investigated by immunohistochemistry using an ECM1 antibody detecting ECM1a and ECM1c isoforms. Semiquantitative analysis of staining intensity was carried out by densitometric image analysis. RESULTS: In normal human skin ECM1a and ECM1c are expressed mainly in the basal cell layers of epidermal keratinocytes and in dermal vessels. For the first time, an expression in the outer root sheath of hair follicles, in sebaceous lobules and epithelium of sweat glands is described. Intrinsically (UV-protected) aged skin shows a significantly reduced expression in basal and upper epidermal cell layers compared with young skin. In photoaged skin, expression is significantly increased within the lower and upper epidermis compared with age-matched UV-protected sites. Importantly, after acute UV exposure in young healthy subjects expression of ECM1 is markedly increased in both lower and upper epidermal cell layers. CONCLUSIONS: This is the first study to demonstrate a regulation of ECM1 expression in human skin by age and UV exposure. These data suggest that ECM1 expression may represent a cutaneous stress response to acute and chronic UV irradiation.