The repair of photo-induced DNA lesions through nucleotide excision repair machinery is still the source of important questions. It has been observed that the repair rate of the different cyclobutane pyrimidine dimers, i.e. the photoproducts induced by dimerization of two π-stacked pyrimidines (T < > T, T < > C, C < > T, C < > C), depends on the nucleobases involved in the lesion. TT derivatives (T < > T) are removed more slowly than those containing cytosine, especially in 5'. Using all-atom molecular dynamics simulations, we demonstrate that the variation of the repair rate observed in human skin and in cultured cutaneous cell may be associated to the recognition of the four lesions by the DDB2 protein moiety, and more specifically by the differential structural deformation induced on the complementary strand and the major groove. These effects may then hamper differentially the downstream recruitment of the repair complexes. The observed DNA deformation correlates with the experimental repair rate and suggests a structural rationale for the different repair rates of CPD by nucleotide excision repair machinery.

Environmental pollution by micro- and nanoplastics (MNPs) raises concerns about their toxicity to humans, particularly through ingestion. While the impact of native MNPs is increasingly documented, environmentally degraded MNPs remain poorly studied. We assessed the intestinal impact of biodegradable and non-biodegradable nanoplastics (NPLs), polyethylene terephthalate (PET) and poly-lactic acid (PLA), both pristine and weathered in environmental conditions. The response of an in vitro intestinal model representative of healthy populations was compared with that of a model of genetic susceptibility to inflammatory bowel disease (IBD), using HT29-MTX cells cocultured with Caco-2 cells expressing either wild-type or mutated nucleotide-binding oligomerization domain 2 (NOD2). Non-differentiated and differentiated cells were exposed for 24 h to these NPLs. Cellular uptake, cytotoxicity, genotoxicity, oxidative stress, inflammation, and epithelial barrier integrity were evaluated. Although PET and PLA accumulated in cells, they showed no significant toxicity. Therefore, as demonstrated for polystyrene NPLs, PLA and PET particles do not cause major toxic impact to intestinal cells upon acute exposure in vitro and, in this exposure scenario, weathering in environmental conditions does not increase their toxicity.

Bisphenol A diglycidyl ether (BADGE) is a diepoxide product used in the synthesis of epoxy resins. Its genotoxic properties were strongly suggested by its ability to induce mutations and micronuclei, but the exact nature of the underlying DNA damage remains to be established. For this purpose, we first applied cellular tests (the Comet assay and 53BP1 immunostaining) to show that direct induction of strand breaks was not the favored genotoxic pathway for BADGE. Consequently, we investigated the formation of DNA adducts using UHPLC coupled with tandem mass spectrometry (UHPLC-MS/MS) analysis following enzymatic hydrolysis. Analyses using single-stage mass spectrometry, product ion scan, and neutral loss monitoring showed that BADGE readily formed monoadducts with DNA bases. The structural assignments were confirmed by the results of accurate thermal degradation studies, which showed that BADGE reacted with DNA like other aliphatic epoxides. We also obtained unambiguous evidence that the reaction of BADGE with DNA led to the formation of biadducts between nonadjacent bases. This was explained by the transient formation of monoadducts bearing a BADGE moiety with an unreacted epoxide group. A sensitive quantitative UHPLC-MS/MS assay was then developed for the detection of BADGE adducts in cellular DNA. All the adducts identified in isolated DNA were also detected in cellular DNA. The proportions between the monoadducts were similar, but the relative yield of biadducts was lower in cellular than in isolated DNA. A time-course study of the level of adducts in DNA also showed that BADGE adducts were substrates for the cellular repair machinery. The present results definitively show that BADGE is a DNA-damaging chemical. The detected adducts represent novel biomarkers of the genotoxicity of BADGE.

In the framework of a Safe, Sustainable, and Recyclable by Design approach for the synthesis of novel plastics, we investigated the toxicological impact of epoxy vitrimers' synthesis. Emphasis was placed on the substitution of conventional hardeners and flame retardants with alternative ingredients. Data were also obtained on the leachates extracted from the final materials. Several toxicological endpoints (cytotoxicity, oxidative stress, epigenetics, genotoxicity, endocrine disruption) were assessed on human cell lines, and aquatic toxicity was evaluated in Daphnia and zebrafish embryo. The bulk of the results showed that use of alternative products did not increase, and in some cases decreased, the toxicological impact of the synthesis of the vitrimers. To facilitate the use of the numerous collected data, they were combined in scores. We first designed a toxicity score that compared each substance with the positive controls of the assays. This score was found to be valuable to report the results in a convenient and easily understandable way. We also developed a performance score that compared the properties of each alternative substance to the conventional reference. This score aimed to be used in a digital decision-making tool for the development of new industrial processes.

Pigments used in tattoos represent a unique case of voluntary, long-term exposure to particles trapped in the skin. Evidence is accumulating that these compounds, and in particular red pigments, are associated with skin disorders and possibly cutaneous cancers. In the present work, we tested the hypothesis of degradation of pigments in the dermis that would lead to the release of diffusible nanoparticles or soluble degradation products. These species could then reach the epidermis and trigger physiological responses by impacting keratinocytes. Two degradation pathways of tattoo pigments were investigated in the present work, which combined physicochemical characterization and toxicological assessment in the HaCaT human keratinocyte cell line. The first pathway was the photodegradation of pigments that we explored in a previous work on Pigment orange 13 (PO13) and extended here to Pigment red 254 (PR254) and Pigment red 122 (PR122). While PR254 was photostable, PR122 was found to release toxic photoproducts. The second studied pathway was the possible degradation in the phagolysosomes of macrophages where pigment particles are stored in the skin. We did not observe degradation upon incubation in reconstructed phagolysosomal medium but rather found that the added immunoglobulins completely inhibited the cytotoxicity of PO13, PR254, and PR122. The observation of a drastic decrease in ζ potential strongly suggested the creation of a protein corona, which led to a decrease in cellular toxicity.

The p53 tumor suppressor is an indispensable regulator of DNA damage responses that accelerates carcinogenesis when mutated. In this report, we uncover a new mechanism by which p53 maintains genomic integrity in the absence of canonical DNA damage response activation. Specifically, loss of p53 dramatically alters chromatin structure at the nuclear periphery, allowing increased transmission of an environmental carcinogen, ultraviolet (UV) radiation, into the nucleus. Genome-wide mapping of UV-induced DNA lesions in p53-deficient primary cells reveals elevated lesion abundance in regions corresponding to locations of high mutation burden in malignant melanomas. These findings uncover a novel role of p53 in the suppression of mutations that contribute to cancer and highlight the critical influence of nuclear architecture in regulating sensitivity to carcinogens.

Pigment particles used in tattooing may exert long terms effect by releasing diffusible degradation products. In the present work, aqueous suspensions of the organic orange diazo pigment PO13 were aged by exposure to simulated sunlight at 40 °C. The morphology and the surface charge of PO13 particles were barely modified upon aging, but primary particles were released by de-agglomeration. Soluble photoproducts were detected in the liquid fractions. One of this photoproduct (DCBP) was produced in large amount in suspension in isopropanol and purified. The toxicological profiles of aged suspensions, their soluble fractions and DCBP were then determined on the keratinocyte cell line HaCaT. Impact of suspensions of PO13 on viability was hardly affected by aging. In contrast, the soluble fractions were more toxic after photo-aging. Suspensions and filtrates induced neither release of reactive oxygen species nor formation of DNA strand breaks. The samples exhibited only limited effects on the proteome of HaCaT cells. Conversely, DCBP was cytotoxic and induced the production of ROS, but was not genotoxic. DCBP was found to activate CYP450 monooxygenases known to be involved in the metabolism of xenobiotics. Altogether, our results show that aging of PO13 leads to the release of toxic soluble compounds.

Accumulation of micro- and nanoplastics in the environment is a major concern, and biodegradable polymers are receiving growing interest as an alternative to more stable material. It remains yet to identify the degradation products of biodegradable plastics. In the present work, we studied polycaprolactone (PCL), a polymer used in a growing number of packaging and medical applications. We gathered quantitative data on the release of soluble oligomers from submicrometric PCL particles (100-1000 mu m in diameter) throughout their degradation. This was made possible by the synthesis of well-characterized PCL oligomers, which were then used for the development of HPLC-mass spectrometry methods. The assay was first applied to degradation studies performed in aqueous media, in relationship to environmental degradation. We observed an influence of temperature and salinity, and obtained specific information on the mechanism of release of PCL hydrolysis products at the sub-micrometric scale. The same approach was then applied to study PCL degradation in cell culture media used in toxicity studies. Oligomers arising from the hydrolysis of PCL particles were released in both the presence and the absence of cells. The size distribution under these conditions was shifted to shorter oligomers than in water. This observation, supported by experiments on synthetic PCL oligomers, pointed to a significant contribution of fetal calf serum used as an additive in cell culture medium. The bulk of these results show that hydrolysis of submicrometric PCL particles and the subsequent production of oligomers is an efficient process that is strongly modulated by the degradation medium.

AbstractBackgroundStudies have demonstrated the potential for damage caused by exposure to radiation at the UVR/visible border region (380–410 nm) and beyond. This includes potentially mutagenic delayed DNA damage, increased gene expression related to photoageing and inflammation, pigmentation and the production of reaction oxygen species. Photoprotection in this region is limited, with a focus on shorter, more energetic UVR regions.ObjectivesTo assess the ability of two sunscreens for their ability to prevent photodamage in the UVA/visible region. Both sunscreens were labelled as SPF 15 and would meet requirements for labelling as UVA protective in the EU and USA. Their ingredients were identical apart from the addition of bis‐ (diethylaminohydroxybenzoyl benzoyl) piperazine (BDBP), a recently approved organic filter that absorbs between 350 and 425 nm.MethodsSunscreens were assessed in vitro in human cell lines and in vivo in healthy human volunteers (Fitzpatrick skin type I–II volunteers). Endpoints were assessed including oxidative stress, gene expression and DNA damage.ResultsThe formulation including the new filter provided significantly more protection than the conventional sunscreen for almost all endpoints tested. The conventional formulation provided some protection compared to unprotected skin or placebo control.ConclusionsThis study demonstrates the requirement for improved photoprotection at the UVR‐visible border region and the importance of assessing sunscreens across a broader range of wavelengths than currently approved protocols require.

Four dinucleotide analogs of thymidylyl(3 '-5 ')thymidine (TpT) have been designed and synthesized with a view to increase the selectivity, with respect to CPD, of efficient UV-induced (6-4) photoproduct formation. The deoxyribose residues of these analogs have been modified to increase north and south conformer populations at 5 '- and 3 '-ends, respectively. Dinucleotides whose 5 '-end north population exceeds ca. 60% and whose 3 '-end population is almost completely south display a three-fold selective enhancement in (6-4) adduct production when exposed to UV radiation, compared to TpT. These experimental results undoubtedly provide robust foundations for studying the singular ground-state proreactive species involved in the (6-4) photoproduct formation mechanism. Evidence is presented that (6-4) photoproduct formation between two thymine residues in dinucleotide analogues is significantly and specifically enhanced when the 5 ''- and 3 ''-end sugar puckering are mainly north and south, respectively.

Sulfur mustard (SM) is a highly potent alkylating vesicant agent and remains a relevant threat to both civilians and military personnel. The eyes are the most sensitive organ after airborne SM exposure, causing ocular injuries with no antidote or specific therapeutics available. In order to identify relevant biomarkers and to obtain a deeper understanding of the underlying biochemical events, we performed an untargeted metabolomics analysis using liquid chromatography coupled to high-resolution mass spectrometry of plasma samples from New Zealand white rabbits ocularly exposed to vapors of SM. Metabolic profiles (332 unique metabolites) from SM-exposed (n = 16) and unexposed rabbits (n = 8) were compared at different time intervals from 1 to 28 days. The observed time-dependent changes in metabolic profiles highlighted the profound dysregulation of the sulfur amino acids, the phenylalanine, the tyrosine and tryptophan pathway, and the polyamine and purine biosynthesis, which could reflect antioxidant and anti-inflammatory activities. Taurine and 3,4-dihydroxy-phenylalanine (Dopa) seem to be specifically related to SM exposure and correspond well with the different phases of ocular damage, while the dysregulation of adenosine, polyamines, and acylcarnitines might be related to ocular neovascularization. Additionally, neither cysteine, N-acetylcysteine, or guanine SM adducts were detected in the plasma of exposed rabbits at any time point. Overall, our study provides an unprecedented view of the plasma metabolic changes post-SM ocular exposure, which may open up the development of potential new treatment strategies.

•CRISPR-Cas9 is a valuable tool for creating XPC-deficient primary melanocytes •XPC-deficient primary melanocytes (©-XPC-Mel) are hypersensitive to UV •©-XPC-Mel pave the way for studying the effects of NER on melanocyte biology •About 500 proteins are differentially expressed in ©-XPC-Mel compared to ©-Ctrl-Mel •The balance between pro- and anti-inflammatory signals is disturbed in ©-XPC-Mel

The low molecular weight degradation products of nanosize biodegradable plastics were identified by mass spectrometry. The only detected products for PLA, PCL and PET were found to a rise from hydrolysis.

A growing number of experimental evidence emphasizes that photobiological phenomena are not always the sum of the effect of individual wavelengths present in the emission spectrum of light sources. Unfortunately, tools are missing to identify such non-additive effects and predict effects of various exposure conditions. In the present work, we addressed these points for the formation of pyrimidine dimers in DNA upon co-exposure to UVC, UVB and UVA radiation. We first applied a combination index approach to determine whether mixtures of theses UV ranges exhibited additive, inhibitory or synergistic effects on the formation of cyclobutane pyrimidine dimers, (6-4) photoproducts and Dewar valence isomers. A predictive approach based on an experimental design strategy was then used to quantify the contribution of each wavelength range to the formation of DNA photoproducts. The obtained models allowed us to accurately predict the level of pyrimidine dimers in DNA irradiated under different conditions. The data were found to be more accurate than those obtained with the simple additive approach underlying the use of action spectra. Experimental design thus appears as an attractive concept that could be widely applied in photobiology even for cellular experiments.

In recent years, interest is growing in the biological cutaneous effects of high-energy visible light (400-450 nm). In the present study, we explored the impact of blue light (BL) on the repair of pyrimidine dimers, the major class of premutagenic DNA damage induced by exposure to sunlight. We unambiguously demonstrate that the exposure of in vitro reconstructed human epidermis to environmentally relevant doses of BL strongly decreases the rate of repair of cyclobutane pyrimidine dimers and pyrimidine (6-4) pyrimidone photoproducts induced by a subsequent UVB irradiation. Using the highly sensitive and specific liquid chromatography-tandem mass spectrometry assay, we did not observe induction of pyrimidine dimers by BL alone. Finally, we showed that application, during the BL exposure step, of a formula containing a new filter, named TriAsorB and affording BL photoprotection, prevented the decrease in DNA repair efficiency. These results emphasize the potential deleterious effects of BL on DNA repair and the interest in providing adequate skin protection against this wavelength range of sunlight. Exposure of reconstructed human epidermis with blue light decreases the rate of repair of pyrimidine dimers induced in DNA by a subsequent UVB irradiation. This deleterious effect is prevented by application of a sunscreen exhibiting blue light photoprotection.image

Post-transcriptional modifications of tRNA nucleotide are important determinants in folding, structure and function. We have successfully identified and characterized a new modified base named 2-methylthio-methylenethio-N-6-(cis-4-hydroxyisopentenyl)adenosine, which is present at position 37 in some tRNAs. We also showed that this new modified adenosine is derived from the known 2-methylthio-methylenethio-N-6-(isopentenyl)adenosine nucleoside by a catalytic cycle of the tRNA-diiron monooxygenase, MiaE, present in Salmonella typhimurium.