UV-induced DNA damage, repair, mutations and oncogenic pathways in skin cancer.

Repair of UV induced DNA damage is of key importance to UV-induced skin carcinogenesis. Specific signal transduction pathways that regulate cell cycling, differentiation and apoptosis are found to be corrupted in skin cancers, e.g., the epidermal growth-stimulating Hedgehog pathway in basal cell carcinomas (BCCs). Mutations in genes coding for proteins in these pathways lead to persistent disturbances that are passed along to daughter cells, e.g., mutations in the gene for the Patched (PTCH) protein in the Hedgehog pathway. Thus far only the point mutations in the P53 gene from squamous cell carcinomas and BCCs, and in PTCH gene from BCC of xeroderma pigmentosum (XP) patients appear to be unambiguously attributable to solar UV radiation. Solar UVB radiation is most effective in causing these point mutations. Other forms of UV-induced genetic changes (e.g., deletions) may, however, contribute to skin carcinogenesis with different wavelength dependencies.     

Protection by food-derived antioxidants from UV-A1-induced photodamage, measured using living skin equivalents

In a study of biomarkers of ultraviolet-A1 radiation (UV-A1)-induced skin damage, living skin equivalent cultures (LSE) were treated with the antioxidants hesperetin and quercetin-3-glucoside and irradiated with 25 or 50 J/cm2 UV-A1. Changes in the following biomarkers were measured; Interleukin 1-alpha (IL-1alpha), Heme Oxygenase-1 (HO-1), TdT-mediated dUTP nick end labeling (TUNEL) and 8-hydroxy-2'-deoxyguanosine (8-OHdG). IL-1alpha and HO-1 were analyzed by real-time PCR, Western blot, enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry. TUNEL and 8-OHdG were determined by (immuno)histochemical techniques. Sections were stained with hematoxylin and eosin (H&E). UV-A1 induced keratinocyte and fibroblast vacuolation and nuclear pyknosis, intense TUNEL staining of fibroblasts and increased staining of cells and nuclei for 8-OHdG. Lesser or marginal increases in intensity followed staining for HO-1 and IL-1alpha. The IL-1alpha increase was confirmed by ELISA assays of the medium supernatants. Hesperetin and quercetin-3-glucoside reduced changes in H&E, 8-OHdG, TUNEL and IL-1alpha. Quercetin-3-glucoside reduced the amount of IL-1alpha in LSE media. These observations support the use of the selected biomarkers to monitor UV-A1 damage and provide evidence that dietary ingredients could reduce ultraviolet-A radiation-induced damage.     

UV mutagenic photoproducts in Escherichia coli and human cells: a molecular genetics perspective on human skin cancer

Abstract— The relevance of photoproducts produced by 254 nm irradiation to human skin cancer is first critically evaluated. Experiments identifying the mutagenic photoproducts at 254 nm are then described. Mutations are primarily due to the(6–4) photoproduct and the cyclobutane pyrimidine dimer, both in E. coli and in human cells. The(6–4) photoproduct may be more important in E. coli and the cyclobutane dimer more important in mammalian cells. In human cells, mutations occur at the C of a TC, CT, or CC cyclobutane dimer, but not at TT cyclobutane dimers, and also appear to occur, less frequently, at the C of TC and CC(6–4) photoproducts. The local structure of DNA is more important in determining the frequency of mutation at a site than is the photoproduct frequency at that site. The effect of DNA structure appears to be due to site-specific lethality.     

Electrophoretic fingerprint metallothionein analysis as a potential prostate cancer biomarker

Prostate-specific antigen (PSA) is a routinely used marker of prostate cancer; however, the cut-off values for unambiguous positive/negative prostate cancer diagnoses are not defined. Therefore, despite the best effort, certain percentage of misdiagnosed cases is being recorded every year. For this reason, search for more specific diagnostic markers is of great interest. In this study, systematic comparison of PSA and metallothionein (MT) levels in blood serum of 46 prostate cancer-diagnosed patients is presented. It is clearly demonstrated that PSA levels vary significantly and despite normal total PSA values in the range of 0 - 4?ng/mL were obtained in over 36.9% of cases, positive prostate cancer was diagnosed by biopsy. In contrary, MT levels were considerably elevated in all tested samples and no significant variations were observed. These results are indicating the potential of MT as an additional prostate cancer marker reducing, in combination with PSA, the probability of false positive/negative diagnosis. To increase the throughput of the screening, chip-based capillary electrophoresis was suggested as a rapid and effective method for the fingerprinting analysis of prostate cancer from diseased blood sera.     

UV radiation and skin cancer in Norway

A distinct increase in skin cancer incidences is observed since the registration started in Norway in the 1950s. As UV radiation is assumed to be the main risk factor for skin cancer, hourly values of the UV irradiance were reconstructed for the period 1957–2005 for 17 of the Norwegian counties (58–70°N). For reconstruction, a radiation transfer model is run with total ozone amount and cloud information as meteorological input. Reconstructed hourly erythemally weighted UV irradiances for about 5 years are compared to measurements at four stations, two stations representing the north–south extension of Norway, and two stations at about 60°N representing the eastern inland – Western coastal contrasts. The agreement between reconstructed and measured UV varies between 0% for the northernmost site to 10–15% overestimation for the other locations. For clear sky, a reasonable agreement between reconstructed and measured data was found for all stations, while for overcast, an overestimation of 10–20% was found for all but the northernmost station. Both the cancer incidences and the reconstructed UV values have a distinct north–south increase. The UV increase towards south is mostly due to increasing solar elevation. The west to east increase is much smaller, and differences in UV are due to differences in both cloud optical thickness and total cloud amount. One additional outcome from this work is that long-term UV-data are reconstructed for Norway, data that can be used in further biological and medical studies related to UV effects.     

Effect of UV-A irradiance on lipid accumulation in Nannochloropsis oculata

Lipids produced by microalgae can be grouped into two categories, storage lipids and structural lipids. Storage lipids are mainly triglycerides (TGs) made up of saturated fatty acids; TGs can be transesterified to produce biodiesel. Structural lipids are made of polyunsaturated fatty acids (PUFAs), which are essential nutrients for aquatic animals and humans. The objectives of this study were: (1) to determine the effect of UV-A at different levels of exposure on total lipid accumulation in Nannochloropsis oculata and check for reciprocity and (2) to study the interactive effect of UV-A and nutrient concentration on lipid accumulation in N. oculata. Objective 1 was accomplished by testing the effects of a range of UV-A irradiance (I), duration of exposure (T) and UV-A doses (I × T) on lipid production by N. oculata. If the same doses have a similar effect, irrespective of I and T, reciprocity holds. UV-A treatments significantly increased the chlorophyll-specific lipid concentration of N. oculata cells, and we were unable to falsify that reciprocity holds. Objective 2 was addressed by a factorial bioassay experiment with manipulated nutrient and UV-A levels. UV-A and decreased nutrients had a synergistic effect on chlorophyll-specific lipid concentration of N. oculata, resulting in higher lipid:chl ratios.     

UV-A induced oxidative stress is more prominent in naturally pigmented aged human RPE cells compared to non-pigmented human RPE cells independent of zinc treatment

To investigate the effects of zinc supplementation on human amelanotic (ARPE-19) and native pigmented retinal pigment epithelial cells (hRPE) under normal light conditions and after ultraviolet A light exposure. hRPE cells, containing both melanin and lipofuscin granules, were prepared from human donor eyes of 60-70 year old patients. Cells of the amelanotic ARPE-19 cell line and pigmented hRPE cells were treated with zinc chloride and subjected to oxidative stress by UV-A irradiation. Intracellular H(2)O(2) formation was measured using a fluorescence oxidation assay. Additionally, apoptosis and viability assays were performed. Control cells were treated identically except for irradiation and zinc supplementation. Under normal light conditions, zinc treated hRPE cells produced less H(2)O(2) than unsupplemented hRPE cells. Viability and apoptosis events did not change. After UV-A irradiation, ARPE and hRPE cells were greatly impaired in all tests performed compared to the non-irradiated controls. No differences were found after zinc supplementation. hRPE cells showed a higher apoptosis and mortality rate than non-pigmented cells when stressed by UV-A light. ARPE cells never showed any zinc related effects. In contrast, without irradiation, zinc supplementation reduced H(2)O(2) production in pigmented hRPE cells slightly. We did not find any zinc effect in irradiated hRPE cells. After UV light exposure, pigmented cells showed a higher apoptosis and mortality than cells lacking any pigmentation. We conclude that cells with pigmentation consisting of melanin and lipofuscin granules have more prooxidative than antioxidative capacity when stressed by UV light exposure compared to cells lacking any pigmentation.     

Advances in fingerprint analysis

Fingerprints have been used in forensic investigations for the identification of individuals since the late 19th century. However, it is now clear that fingerprints can provide significantly more information about an individual. Here, we highlight the considerable advances in fingerprinting technology that can simultaneously provide chemical information regarding the drugs ingested and the explosives and drugs handled by a person as well as the identity of that individual.     

Photodimerization of 7,8-dihydroneopterin in aqueous solution under UV-A irradiation

7,8-Dihydroneopterin (H(2) Nep) is secreted during the oxidative burst of stimulated macrophages. The photochemistry of H(2) Nep was investigated in neutral aqueous solutions exposed to UV-A radiation (320-400nm) at room temperature. The kinetics were followed by UV/Vis spectrophotometry and HPLC, whereas the photoproducts were analyzed by electrospray ionization mass spectrometry. Excitation of H(2) Nep leads to the formation of isomeric dimers with molecular masses equal to exactly twice the molecular mass of the reactant. The corresponding quantum yield of H(2) Nep consumption (Φ(-R) =(3.8±0.5)×10(-2)) was independent of O(2) and reactant concentrations. Mechanistic implications are discussed.     

Tribological characteristics of medical gloves in contact with human skin and skin equivalents

The progress in new surface modification techniques towards the manufacture of tailored latex articles is faced with the requirement of new characterization techniques to determine the tribological properties of elastomer surfaces. Thus, the present study aims at the characterization of friction properties of commercially available medical gloves to assess their donning performance. The experimental design of the friction test method involves a linear movement of the glove sample across selected counterpart materials. In the first step, bio-tribological studies with a panel are carried out enabling an objective evaluation of the donning properties of gloves. These results are compared to the coefficient of friction of the gloves against various skin equivalents including polymer based materials, glass and biological tissue. In terms of skin friction, a high correlation between human skin and the biological skin equivalent is obtained. However, in contrast to bio-tribological studies, the friction experiments with biological tissue benefit from high reproducibility and low standard deviation. With the established test method the influence of state-of-the-art surface functionalization processes (chlorination and polymer-based coatings) and lubricants (cornstarch) on the friction properties of medical gloves is investigated in order to evaluate crucial process and surface parameters for low surface friction against human skin.     

Arachidonic acid metabolism in human skin fibroblast cultures

Conclusion Our ongoing study shows that HSF cultures can be used as an in vitro model for AA metabolism studies. The incubation medium however has to be standardised in order to obtain comparable results. The addition of albumin to the medium is necessary for dissolution of different agents like ionophore, indomethacin or others and does not change the AA metabolism. Differences between different cell lines of healthy donors are under investigation but seem to be of minor importance when the skin biopsie is taken from the same place, whereas greater differences were observed between HSF cultured from arm or foreskin biopsies. Finally it could be proved that the cyclooxygenase pathway is more important than that of the lipoxygenase and that 11-HETE derives from the cyclooxygenase and not from the lipoxygenase activity. The main lipoxygenase product is 15-HETE.

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Arachidonsäurestoffwechsel in menschlichen Hautfibroblasten-Kulturen

     

Microcalorimetric studies of human skin cells

A short but comprehensive review is given about direct calorimetric measurements on human skin cells in vitro.     

On-line detection of human skin vapors

Vapors released by the skin in the hand of one human subject are detected in real time by sampling them directly from the ambient gas surrounding the hand, ionizing them by secondary electrospray ionization (SESI, via contact with the charged cloud from an electrospray source), and analyzing them in a mass spectrometer with an atmospheric pressure source (API-MS). This gas-phase approach is complementary to alternative on-line surface ionization methods such as DESI and DART. A dominating peak of lactic acid and a complete series of saturated and singly unsaturated fatty acids (C₁₂ to C₁₈) are observed, in accordance with previous off-line studies by gas chromatography-mass spectrometry. Several other metabolites have been identified, including ketomonocarboxylic and hydroxymonocarboxylic acids.     

MALDI-MS imaging of lipids in ex vivo human skin

Lipidomics is a rapidly expanding area of scientific research and there are a number of analytical techniques that are employed to facilitate investigations. One such technique is matrix-assisted laser desorption ionisation (MALDI) mass spectrometry (MS). Previous MALDI-MS studies involving lipidomic investigation have included the analysis of a number of different ex vivo tissues, most of which were obtained from animal models, with only a few being of human origin. In this study, we describe the use of MALDI-MS, MS/MS and MS imaging methods for analysing lipids within cross-sections of ex vivo human skin. It has been possible to tentatively identify lipid species via accurate mass measurement MALDI-MS and also to confirm the identity of a number of these species via MALDI-MS/MS, in experiments carried out directly on tissue. The main lipid species detected include glycerophospholipids and sphingolipids. MALDI images have been generated at a spatial resolution of 150 and 30 μm, using a MALDI quadrupole time-of-flight Q-Star Pulsar-i ^TM (Applied Biosystems/MDS Sciex, Concord, ON, Canada) and a MALDI high-definition MS (HDMS) SYNAPT G2-HDMS^TM system (Waters, Manchester, UK), respectively. These images show the normal distribution of lipids within human skin, which will provide the basis for assessing alterations in lipid profiles linked to specific skin conditions e.g. sensitisation, in future investigations.     

Zinc analysis in human skin by laser induced-breakdown spectroscopy

The feasibility of using laser induced-breakdown spectroscopy (LIBS) as a quick and simple method to analyze trace elemental concentrations in the stratum corneum of human skin was investigated. A 60 mJ pulse(-1) Nd:YAG laser operating at 1064 nm was used to form the laser induced plasma. Zinc was chosen for this study. By using poly-(methyl methacrylate) (PMMA) solution to coat the slides, zinc standard solutions were dispersed on the slides with the help of a centrifuge. The percent of R.S.D. of the dispersed area size obtained from 17 measurements was 6%. The precision from intra-measurements (precision on one slide) and inter-measurements (precision between slides) was 4-8% and less than 1%, respectively. A calibration curve with a linear correlation coefficient of 0.998 and a detection limit of 0.3 ng cm(-2) were obtained. An aqueous zinc solution or zinc ointment was deposited on the biceps area of the forearms of several volunteers for a time period of 30 min and 2.75 h. Six skin layers of 2-3 mum each in thickness were then removed using cyanoacrylate glue without causing any pain for the volunteers. The results indicated that Zn was absorbed through the skin and the concentration decreased exponentially with depth into the skin. The results indicate that LIBS is a useful tool for trace elemental analysis in human skin.     

Confocal Raman microspectroscopy for skin characterization: a comparative study between human skin and pig skin

The present paper provides a spectral comparison between abdominal human skin (Transkin) and pig ear skin using confocal Raman microspectroscopy at 660 nm. Pig ear skin is usually utilized as a substitute for human skin for active ingredients assessment in dermatological and cosmetics fields. Herein, the comparison is made at the level of the stratum corneum (SC), the SC/epidermis junction and the viable epidermis. The 660 nm excitation source appears to be the most appropriate wavelength for such skin characterization. From Raman signatures of both skin types, a tentative assignment of vibrations was performed in the fingerprint and the high wavenumber spectral regions. Significant differences were highlighted for lipid content in in-depth spectra and for hyaluronic acid (HA) and carotenoid in SC spectra. Marked tissular variability was also revealed by certain Raman vibrations. These intrinsic molecular data probed by confocal Raman microspectroscopy have to be considered for further applications such as cutaneous drug permeation.     

On the assessment of melanin in human skin in vivo

Abstract— A method for estimating the amount of pigment in normal human skin in uivo is presented. This method is based on remittance spectroscopy. The spectrum of normal skin is compared to amelanotic skin and the logarithm of the ratio is fitted with a straight line in the range 620-720 nm. The parameters obtained are strongly correlated for all the volunteers in this study. therefore each spectrum determines one parameter for each individual. When similar analysis was performed on DOPA-melanin we obtained the same strong correlation among different concentrations. We are thus able to determine a relation between the coefficient obtained from the remittance spectrum from normal skin and an equivalent concentration of DOPA-melanin in aqueous solution. We can thus estimate. to a first approximation, the total melanin mass in human skin non-invasively, and can determine a parameter that is uniquely correlated to the amount of melanin pigment in the skin.     

Use of UV-A Energy for Photosynthesis in the Red Macroalga Gracilaria lemaneiformis

UV radiation is known to inhibit photosynthetically active radiation (PAR)-driven photosynthesis; however, moderate levels of UV-A have been shown to enhance photosynthesis and growth rates of some algae. Here, we have shown that UV-A alone could drive photosynthetic utilization of bicarbonate in the red alga Gracilaria lemaneiformis as evidenced in either O₂ evolution or carbon fixation as well as pH drift. Addition of UV-B inhibited the apparent photosynthetic efficiency, raised the photosynthetic compensation point and photosynthesis-saturating irradiance level, but did not significantly affect the maximal rate of photosynthetic O₂ evolution. The electron transport inhibitor, DCMU, inhibited the photosynthesis completely, reflecting that energy of UV-A was transferred in the same way as that of PAR. Inorganic carbon acquisition for photosynthesis under UV alone was inhibited by the inhibitors of carbonic anhydrase. The results provided the evidence that G. lemaneiformis can use UV-A efficiently to drive photosynthesis based on the utilization of bicarbonate, which could contribute significantly to the enhanced photosynthesis in the presence of UV-A observed under reduced levels of solar radiation.