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Abstract: The rapid development of new chemicals and consumer products has raised concerns about their potential genotoxic effects on human health, including DNA damage leading to serious diseases. For such new chemicals and pharmaceutical products, international regulations require genotoxicity data, initially obtained through in vitro tests, followed by in vivo experiments, if needed. Traditionally, laboratory animals have been used for this purpose, however, they are costly, ethically problematic, and often unreliable due to species differences. Therefore, innovative more accurate in vitro testing approaches are rapidly being developed to replace, refine and reduce (3R) the use of animals for experimental purposes and to improve the relevance for humans in toxicology studies. One of such innovative approaches are in vitro three-dimensional (3D) cell models, which are already being highlighted as superior alternatives to the two-dimensional (2D) cell cultures that are traditionally used as in vitro models for the safety testing of chemicals and pharmaceuticals. 3D cell models provide physiologically relevant information and more predictive data for in vivo conditions. In the review article, we provide a comprehensive overview of 3D hepatic cell models, including HepG2, HepG2/C3A, HepaRG, human primary hepatocytes, and iPSC-derived hepatocytes, and their application in the field of genotoxicology. Through a detailed literature analysis, we identified 31 studies conducted between 2007 and April 2024 that used a variety of standard methods, such as the comet assay, the micronucleus assay, and the γH2AX assay, as well as new methodological approaches, including toxicogenomics, to assess the cytotoxic and genotoxic activity of chemicals, nanoparticles and natural toxins. Based on our search, we can conclude that the use of in vitro 3D cell models for genotoxicity testing has been increasing over the years and that 3D cell models have an even greater potential for future implementation and further refinement in genetic toxicology and risk assessment.
Keywords: genotoxicity, advanced 3D in vitro models, hepatic cells, spheroids, comet assay, micronucleus assay, genotoxicology, toxicological studies
Published in DiRROS: 14.11.2024; Views: 270; Downloads: 120
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Abstract: Aim: Parvalbumin (PV) is a primary calcium buffer in mouse fast skeletal musclefibers. Previous work showed that PV ablation has a limited impact on cytosolicCa2+ ([Ca2+]cyto) transients and contractile response, while it enhances mitochon-drial density and mitochondrial matrix-free calcium concentration ([Ca2+]mito).Here, we aimed to quantitatively test the hypothesis that mitochondria act tocompensate for PV deficiency.Methods: We determined the free Ca 2+ redistribution during a 2 s 60 Hz tetanicstimulation in the sarcoplasmic reticulum, cytosol, and mitochondria. Via a re-action–diffusion Ca 2+ model, we quantitatively evaluated mitochondrial uptakeand storage capacity requirements to compensate for PV lack and analyzed pos-sible extracellular export.Results: [Ca 2+]mito during tetanic stimulation is greater in knock-out (KO)(1362 ± 392 nM) than in wild-type (WT) (855 ± 392 nM), p < 0.05. Under the as-sumption of a non-linear intramitochondrial buffering, the model predicts an ac-cumulation of 725 μmoles/Lfiber (buffering ratio 1:11 000) in KO, much higherthan in WT (137 μmoles/Lfiber, ratio 1:4500). The required transport rate via mi-tochondrial calcium uniporter (MCU) reaches 3 mM/s, compatible with availableliterature. TEM images of calcium entry units and Mn2+ quenching showed a greater capacity of store- operated calcium entry in KO compared to WT. However,levels of [Ca 2+]cyto during tetanic stimulation were not modulated to variations ofextracellular calcium.Conclusions: The model-based analysis of experimentally determined calciumdistribution during tetanic stimulation showed that mitochondria can act as abuffer to compensate for the lack of PV. This result contributes to a better under-standing of mitochondria's role in modulating [Ca2+]cyto in skeletal muscle fibers.
Keywords: calcium, mitochondria, mouse skeletal muscle fibers, parvalbumin, reaction-diffusion model
Published in DiRROS: 13.11.2024; Views: 169; Downloads: 2920
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Keywords: monitoring, stanje gozdov, osutost dreves, poškodovanost dreves, rast dreves, meteorološke meritve, ozon, pritalna vegetacija, fenološka opazovanja, foliarni popis, zračne usedline
Published in DiRROS: 12.11.2024; Views: 153; Downloads: 0
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Published in DiRROS: 12.11.2024; Views: 191; Downloads: 565
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Abstract: Questions The long-term response of understorey vegetation to increasing tree mortality has rarely been addressed in resurvey studies. For two Quercus-dominated forest types, we asked: (a) How did overstorey alterations, induced by canopy mortality, affect understorey diversity and composition? (b) Is there a signal of global change effects on understorey communities? (c) Are these assemblages experiencing a homogenization process? Location Five sites in Quercus robur (QR) and four sites in Q. petraea (QP) forests, Slovenia. Methods We studied changes in vascular plants in the understorey layer from 1992/1993 to 2023 across 45 permanent 20 m × 20 m plots in QR and QP forests, respectively. Vegetation surveys were carried out following the standard Braun-Blanquet method. We compared original surveys with recent resurveys using multivariate analysis, ecological indicator values (EIV), plant traits and methods that quantify changes in individual species. Results Since the early 1990s, tree layer cover decreased from 95% to an average of 55% in QR, whereas it remained relatively high (77%) in QP plots. This resulted in denser understorey vegetation and a significant increase in plot-level species richness in QR forests, but a slight decrease in QP forests. The extensive loss of canopy cover and disturbance effects in QR forests caused significant changes in species composition. Species turnover in QR was driven by colonization of new disturbance-tolerant taxa characterized by ruderal traits, whereas the compositional shift in QP was to a greater extent due to species losses. We detected a process of vegetation thermophilization (increase in EIV-temperature), suggesting an effect of rapid climatic warming. Understorey communities are now more similar to each other than 30 years ago, indicating a decrease in beta-diversity (floristic homogenization). Conclusions Despite some common trends, vegetation responses were forest type-specific. Our study presents evidence of understorey vegetation changes triggered by increased canopy mortality (a strong local driver particularly in QR plots) and also points to the signal of global change symptoms (thermophilization, homogenization), which acted rather independently from the observed decline in tree layer cover.
Keywords: floristic homogenization, forest disturbances, permanent vegetation plots, Quercus robur, Quercus petraea, resurvey study, Slovenia, thermophilization, tree layer cover
Published in DiRROS: 11.11.2024; Views: 201; Downloads: 210
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