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Background: Although chlorpyrifos (CPS) has been banned in many developed countries, it still remains one of the best-selling pesticides in the world. Widespread environmental and occupational exposure to CPS pose a serious risk to human health. Another environmental factor that can adversely affect human health is ultraviolet radiation B (UVB, 280-315 nm wave length). Here we attempt determine if exposure to CPS can modify toxic effects of UVB. Such situation might be a common phenomenon in agriculture workers, where exposure to both factors takes place., Methods: Two skin cell lines; namely human immortalized keratinocytes HaCaT and BJ human fibroblasts were used in this study. Cytotoxicity was investigated using a cell membrane damage detection assay (LDH Cytotoxicity Assay), a DNA damage detection assay (Comet Assay), an apoptosis induction detection assay (Apo-ONE Homogeneous Caspase-3/7 Assay) and a cell reactive oxygen species detection assay (ROS-Glo H 2 O 2 assay). Cytokine IL-6 production was also measured in cells using an ELISA IL-6 Assay., Results: Pre-incubation of skin cells with CPS significantly increased UVB-induced toxicity at the highest UVB doses (15 and 20 mJ/cm 2 ). Also pre-exposure of BJ cells to CPS significantly increased the level of DNA damage, except for 20 mJ/cm 2 UVB. In contrast, pre-exposure of HaCaT cells, to CPS prior to UVB radiation did not cause any significant changes. A decrease in caspase 3/7 activity was observed in HaCaT cells pre-exposed to 250 µM CPS and 5 mJ/cm 2 UVB. Meanwhile, no statistically significant changes were observed in fibroblasts. In HaCaT cells, pre-exposure to CPS resulted in a statistically significant increase in ROS production. Also, in BJ cells, similar results were obtained except for 20 mJ/cm 2 . Interestingly, CPS seems to inhibited IL-6 production in HaCaT and BJ cells exposed to UVB (in the case of HaCaT cells for all UVB doses, while for BJ cells only at 15 and 20 mJ/cm 2 )., Conclusions: In conclusion, the present study indicates that CPS may contribute to the increased UVB-induced toxicity in skin cells, which was likely due to the induction of ROS formation along with the generation of DNA damage. However, further studies are required to gain better understanding of the mechanisms involved., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: The present study aimed to evaluate the potential differences in the biological effects of two types of spherical silver particles of 20 and 200 nm (Ag20 and Ag200), and of PVP-coated silver nanowires (AgNWs) with a diameter of 50 nm and length up to 50 μm, using a complex 3D model representative for the alveolar barrier cultured at air-liquid interface (ALI). The alveolar model was exposed to 0.05, 0.5 and 5 μg/cm 2 of test compounds at ALI using a state-of-the-art exposure system (Vitrocell™Cloud System). Endpoints related to the oxidative stress induction, anti-oxidant defence mechanisms, pro-inflammatory responses and cellular death were selected to evaluate the biocompatibility of silver particles and nanowires (AgNMs) and to further ascribe particular biological effects to the different morphologic properties between the three types of AgNMs evaluated., Results: Significant cytotoxic effect was observed for all three types of AgNMs at the highest tested doses. The increased mRNA levels of the pro-apoptotic gene CASP7 suggests that apoptosis may occur after exposure to AgNWs. All three types of AgNMs increased the mRNA level of the anti-oxidant enzyme HMOX-1 and of the metal-binding anti-oxidant metallothioneins (MTs), with AgNWs being the most potent inducer. Even though all types of AgNMs induced the nuclear translocation of NF-kB, only AgNWs increased the mRNA level of pro-inflammatory mediators. The pro-inflammatory response elicited by AgNWs was further confirmed by the increased secretion of the 10 evaluated interleukins., Conclusion: In the current study, we demonstrated that the direct exposure of a complex tetra-culture alveolar model to different types of AgNMs at ALI induces shape- and size-specific biological responses. From the three AgNMs tested, AgNWs were the most potent in inducing biological alterations. Starting from 50 ng/cm 2 , a dose representative for an acute exposure in a high exposure occupational setting, AgNWs induced prominent changes indicative for a pro-inflammatory response. Even though the acute responses towards a dose representative for a full-lifetime exposure were also evaluated, chronic exposure scenarios at low dose are still unquestionably needed to reveal the human health impact of AgNMs during realistic conditions.

Background: The increased incorporation of silver nanoparticles (Ag NPs) into consumer products makes the characterization of potential risk for humans and other organisms essential. The oral route is an important uptake route for NPs, therefore the study of the gastrointestinal tract in respect to NP uptake and toxicity is very timely. The aim of the present study was to evaluate the effects of Ag NPs and ions on a Caco-2/TC7:HT29-MTX intestinal co-culture model with mucus secretion, which constitutes an important protective barrier to exogenous agents in vivo and may strongly influence particle uptake., Methods: The presence of the mucus layer was confirmed with staining techniques (alcian blue and toluidine blue). Mono and co-cultures of Caco-2/TC7 and HT29-MTX cells were exposed to Ag NPs (Ag 20 and 200 nm) and AgNO3 and viability (alamar blue), ROS induction (DCFH-DA assay) and IL-8 release (ELISA) were measured. The particle agglomeration in the media was evaluated with DLS and the ion release with ultrafiltration and ICP-MS. The effects of the Ag NPs and AgNO3 on cells in co-culture were studied at a proteome level with two-dimensional difference in gel electrophoresis (2D-DIGE) followed by Matrix Assisted Laser Desorption Ionization - Time Of Flight/ Time Of Flight (MALDI-TOF/TOF) mass spectrometry (MS). Intracellular localization was assessed with NanoSIMS and TEM., Results: The presence of mucus layer led to protection against ROS and decrease in IL-8 release. Both Ag 20 and 200 nm NPs were taken up by the cells and Ag NPs 20 nm were mainly localized in organelles with high sulfur content. A dose- and size-dependent increase in IL-8 release was observed with a lack of cytotoxicity and oxidative stress. Sixty one differentially abundant proteins were identified involved in cytoskeleton arrangement and cell cycle, oxidative stress, apoptosis, metabolism/detoxification and stress., Conclusions: The presence of mucus layer had an impact on modulating the induced toxicity of NPs. NP-specific effects were observed for uptake, pro-inflammatory response and changes at the proteome level. The low level of overlap between differentially abundant proteins observed in both Ag NPs and AgNO3 treated co-culture suggests size-dependent responses that cannot only be attributed to soluble Ag.

Background: The increasing body of evidence suggest that nanomaterials toxicity is associated with generation of oxidative stress. In this paper we investigated the role of respiration in silver nanoparticles (AgNPs) generated oxidative stress and toxicity. Since cancer cells rely on glucose as the main source of energy supply, glucose availability might be an important determinant of NPs toxicity., Methods: AgNPs of 20 nm nominal diameter were used as a model NPs. HepG2 cells were cultured in the media with high (25 mM) or low (5.5 mM) glucose content and treated with 20 nm AgNPs. AgNPs-induced toxicity was tested by neutral red assay. Generation of H2O2 in mitochondria was evaluated by use of mitochondria specific protein indicator HyPer-Mito. Expression of a 77 oxidative stress related genes was assessed by qPCR. The activity of antioxidant enzymes was estimated colorimetrically by dedicated methods in cell homogenates., Results: AgNPs-induced dose-dependent generation of H2O2 and toxicity was observed. Toxicity of AgNPs towards cells maintained in the low glucose medium was significantly lower than the toxicity towards cells growing in the high glucose concentration. Scarceness of glucose supply resulted in upregulation of the endogenous antioxidant defence mechanisms that in turn alleviated AgNPs dependent ROS generation and toxicity., Conclusion: Glucose availability can modify toxicity of AgNPs via elevation of antioxidant defence triggered by oxidative stress resulted from enhanced oxidative phosphorylation in mitochondria and associated generation of ROS. Presented results strengthen the idea of strong linkage between NPs toxicity and intracellular respiration and possibly other mitochondria dependent processes.

Background: Respirable crystalline silica (silicon dioxide; SiO₂, quartz) particles are known to induce chronic inflammation and lung disease upon long-term inhalation, whereas non-crystalline (amorphous) SiO₂ particles in the submicrometre range are regarded as less harmful. Several reports have demonstrated that crystalline, but also non-crystalline silica particles induce IL-1β release from macrophages via the NALP3-inflammasome complex (caspase-1, ASC and NALP3) in the presence of lipopolysaccharide (LPS) from bacteria. Our aim was to study the potential of different non-crystalline SiO₂ particles from the nano- to submicro-sized range to activate IL-1β responses in LPS-primed RAW264.7 macrophages and primary rat lung macrophages. The role of the NALP3-inflammasome and up-stream mechanisms was further explored in RAW264.7 cells., Results: In the present study, we have shown that 6 h exposure to non-crystalline SiO₂ particles in nano- (SiNPs, 5-20 nm, 50 nm) and submicro-sizes induced strong IL-1β responses in LPS-primed mouse macrophages (RAW264.7) and primary rat lung macrophages. The primary lung macrophages were more sensitive to Si-exposure than the RAW-macrophages, and responded more strongly. In the lung macrophages, crystalline silica (MinUsil 5) induced IL-1β release more potently than the non-crystalline Si50 and Si500, when adjusted to surface area. This difference was much less pronounced versus fumed SiNPs. The caspase-1 inhibitor zYVAD and RNA silencing of the NALP3 receptor reduced the particle-induced IL-1β release in the RAW264.7 macrophages. Furthermore, inhibitors of phagocytosis, endosomal acidification, and cathepsin B activity reduced the IL-1β responses to the different particles to a similar extent., Conclusions: In conclusion, non-crystalline silica particles in the nano- and submicro-size ranges seemed to induce IL-1β release from LPS-primed RAW264.7 macrophages via similar mechanisms as crystalline silica, involving particle uptake, phagosomal leakage and activation of the NALP3 inflammasome. Notably, rat primary lung macrophages were more sensitive with respect to silica-induced IL-1β release. The differential response patterns obtained suggest that silica-induced IL-1β responses not only depend on the particle surface area, but on factors and/or mechanisms such as particle reactivity or particle uptake. These findings may suggest that bacterial infection via LPS may augment acute inflammatory effects of non-crystalline as well as crystalline silica particles.

The erythrocytes of 12-month old Sod1 (-/-) mice showed an increased level of reactive oxygen species (ROS), as estimated by the degree of dihydroethidine and dihydrorhodamine oxidation, and the increased level of Heinz bodies. No indices of severe oxidative stress were found in the red blood cells and blood plasma of Sod1 (-/-) mice as judged from the lack of significant changes in the levels of erythrocyte and plasma glutathione, plasma protein thiol and carbonyl groups and thiobarbituric-acid reactive substances in the blood plasma. However, a decreased erythrocyte lifespan, increased reticulocyte count and splenomegaly were noted, indicating the importance of superoxide dismutase for maintaining erythrocyte viability. The levels of erythrocyte ROS and Heinz bodies and the reticulocyte count were indistinguishable in Sod1 (+/+) and Sod1 (+/-) mice, suggesting that a superoxide dismutase activity decrease to half of its normal value may be sufficient to secure the protective effects of the enzyme.