Molecular Responses to Photogenotoxic Stress Induced by the Antibiotic Lomefloxacin in Human Skin Cells: From DNA Damage to Apoptosis

Photo-unstable chemicals sometimes behave as phototoxins in skin, inducing untoward clinical side-effects when exposed to sunlight. Some drugs, such as psoralens or fluoroquinolones, can damage genomic DNA, thus increasing the risk of photocarcinogenesis. Here, lomefloxacin, an antibiotic from the fluoroquinolone family known to be involved in skin tumor development in photoexposed mice, was studied using normal human skin cells in culture: fibroblasts, keratinocytes, and Caucasian melanocytes. When treated cells were exposed to simulated solar ultraviolet A (320–400 nm), lomefloxacin induced damage such as strand breaks and pyrimidine dimers in genomic DNA. Lomefloxacin also triggered various stress responses: heme-oxygenase-1 expression in fibroblasts, changes in p53 status as shown by the accumulation of p53 and p21 proteins or the induction of MDM2 and GADD45 genes, and stimulation of melanogenesis by increasing the tyrosinase activity in melanocytes. Lomefloxacin could also lead to apoptosis in keratinocytes exposed to ultraviolet A: caspase-3 was activated and FAS-L gene was induced. Moreover, keratinocytes were shown to be the most sensitive cell type to lomefloxacin phototoxic effects, in spite of the well-established effectiveness of their antioxidant equipment. These data show that the phototoxicity of a given drug can be driven by different mechanisms and that its biologic impact varies according to cell type

Viral sequence variation in chronic carriers of hepatitis C virus has a low impact on liver steatosis.: HCV variability and steatosis

International audienceMost clinical studies suggest that the prevalence and severity of liver steatosis are higher in patients infected with hepatitis C virus (HCV) genotype 3 than in patients infected with other genotypes. This may reflect the diversity and specific intrinsic properties of genotype 3 virus proteins. We analyzed the possible association of particular residues of the HCV core and NS5A proteins known to dysregulate lipid metabolism with steatosis severity in the livers of patients chronically infected with HCV. We used transmission electron microscopy to quantify liver steatosis precisely in a group of 27 patients, 12 of whom were infected with a genotype 3 virus, the other 15 being infected with viruses of other genotypes. We determined the area covered by lipid droplets in liver tissues and analyzed the diversity of the core and NS5A regions encoded by the viral variants circulating in these patients. The area covered by lipid droplets did not differ significantly between patients infected with genotype 3 viruses and those infected with other genotypes. The core and NS5A protein sequences of the viral variants circulating in patients with mild or severe steatosis were evenly distributed throughout the phylogenic trees established from all the collected sequences. Thus, individual host factors seem to play a much greater role than viral factors in the development of severe steatosis in patients chronically infected with HCV, including those infected with genotype 3 viruses

Hardware/software support for adaptive work-stealing in on-chip multiprocessor

International audienceDuring the past few years, embedded digital systems have been requested to provide a huge amount of processing power and functionality. A very likely foreseeable step to pursue this computational and flexibility trend is the generalization of on-chip multiprocessor platforms (MPSoC). In that context, choosing a programming model and providing optimized hardware support to it on these platforms is a challenging task. To deal in a portable way with MPSoCs having a different number of processors running possibly at different frequencies, work-stealing (WS) based parallelization is a current research trend. The contribution of this paper is to evaluate the impact of some simple MPSoCs' architecture characteristics on the performance of WS in the MPSoC context. The previous evaluations of WS, either theoretical or experimental, were done on fixed multicores architectures. This work extends these studies by exploring the use of WS for the codesign of embedded applications on MPSoC platforms with different hardware capabilities, thanks to cycle-accurate measures. We firstly study the architectural choices suited to WS algorithms and measure the benefit of these architectural modifications. To assert whether WS is suited to the MPSoC context, we experimentally measure its intrinsic implementation overhead on the most efficient architectural designs. Finally, we validate the performances of the approach on two real applications: a regular multimedia application (temporal noise reduction) and an irregular computation intensive application (frames of the Mandelbrot set). Our results show that enhancing MPSoC platforms having up to 16 processors with widespread hardware support mechanisms can lead to important performance improvements at acceptable hardware cost for the considered applications

Genotoxicity assays with Episkin ® , a reconstructed skin model: Towards new tools for in vitro risk assessment of dermally applied compounds?

Abstract In vitro reconstructed human skin such as Episkin® is a widely used model in safety or efficacy pre-screening tests. It is of growing interest for regulatory purposes as an alternative to animal testing. The reduction and eventually the replacement of In vivo toxicity testing require the development of In vitro models to predict the genotoxic or other endpoint risk. This can be achieved if these new assays take into account the exposure conditions in a more relevant way. To that end, new approaches are proposed using human reconstructed skin model for in vitro toxicology assessment. Reconstructed skin models have not been used for In vitro genotoxicity testing so far, though they present clear advantages over mouse skin for human risk assessment. This work highlights the development of a specific protocol for performing genotoxicity assays. The skin is indeed a biologically active barrier driving the exposure to compounds and their possible metabolites. A specific co-culture system using Episkin® and target cells to perform a regular micronucleus assay is presented. This first step in using human reconstructed skin for genotoxicity testing has aimed at improving the relevance of exposure conditions in In vitro genotoxicity assays for topically applied compounds

Increased melanogenesis is a risk factor for oxidative DNA damage--study on cultured melanocytes and atypical nevus cells

Melanin synthesis is an oxygen-dependent process that acts as a potential source of reactive oxygen species (ROS) inside pigment-forming cells. The synthesis of the lighter variant of melanin, pheomelanin, consumes cysteine and this may limit the capacity of the cellular antioxidative defense. We show that tyrosine-induced melanogenesis in cultured normal human melanocytes (NHM) is accompanied by increased production of ROS and decreased concentration of intracellular glutathione. Clinical atypical (dysplastic) nevi (DN) regularly contain more melanin than do normal melanocytes (MC). We also show that in these cultured DN cells three out of four exhibit elevated synthesis of pheomelanin and this is accompanied by their early senescence. By using various redox-sensitive molecular probes, we demonstrate that cultured DN cells produce significantly more ROS than do normal MC from the same donor. Our experiments employing single-cell gel electrophoresis (comet assay) usually reveal higher fragmentation of DNA in DN cells than in normal MC. Even if in some cases the normal alkaline comet assay shows no differences in DNA fragmentation between DN cells and normal MC, the use of the comet assay with formamidopyrimidine DNA glycosylase can disclose that the DNA of the cultured DN cells harbor more oxidative damage than the DNA of normal MC from the same perso

Transcriptional upregulation of Nrf2-dependent phase II detoxification genes in the involved epidermis of vitiligo vulgaris

Oxidative stress is widely believed to be a contributing factor in vitiligo pathogenesis. To explore mechanisms by which epidermis responds to mounting oxidative stress, we investigated the involvement of phase II detoxification genes in vitiligo. Phase II detoxification pathways have recently been identified as being important in the regulation of epidermal skin homeostasis. In this study we show that the key transcription factor nuclear factor E2-related factor 2 (Nrf2) and the downstream genes NAD(P)H:quinone oxidase-1 (NQO-1), γ-glutamyl cystine ligase catalytic subunit (GCLC), and γ-glutamyl cystine ligase modifying subunit (GCLM) are upregulated in the lesional epidermal skin of subjects with vitiligo vulgaris. The differences between lesional and nonlesional skin were further investigated by studying the induced expression of Nrf2-dependent transcripts in skin punch biopsies using curcumin and santalol. Surprisingly, nonlesional skin showed induction of all transcripts while a similar effect was not observed for the skin punches from the lesional skin. The use of curcumin and santalol on epidermal cells showed that keratinocytes were more susceptible to apoptosis, whereas melanocytes induced phase II genes under the same concentrations with negligible apoptosis. Our studies provide new insights into the role of phase II detoxification pathway in maintaining skin homeostasis and sustaining redox balance in vitiligo patients

TEM observation of representative liver biopsy specimens from three patients chronically infected with HCV.

<p>The liver in A (patient P16) shows a small number of small LD (arrows) and was found to have a lipid droplet area <1000 µm² per 50,000 µm² of tissue. The liver in B (patient P20) has extremely large LD and was found to have a LD area >10,000 µm² per 50,000 µm² of tissue. The TEM photograph in C shows a high magnification of an individual hepatocyte with several large LD, taken from a liver biopsy specimen (patient P03) considered intermediate to the other two in terms of steatosis severity. Scale bars: 20 µm in A and B; 2 µm in C.</p