Archives of Toxicology最新文献

Calcium/calmodulin-dependent protein kinases (CaMKs) are important proteins in the calcium signaling cascade response pathway, which can broadly regulate biological functions in vivo. Multifunctional CaMKs play key roles in neural development, including neuronal circuit building, synaptic plasticity establishment, and neurotrophic factor secretion. Currently, four familial proteins, calcium/calmodulin-dependent protein kinase I (CaMKI), calcium/calmodulin-dependent protein kinase II (CaMKII), eukaryotic elongation factor 2 kinase (eEF2K, popularly known as CaMKIII) and calcium/calmodulin-dependent protein kinase IV (CaMKIV), are thought to have been the most extensively studied during neurodevelopment. Although their spatial structures are extremely similar, as well as the initial starting point of activation, both require the activation of calcium and calmodulin (CaM) complexes to be involved in the process, and the phosphorylation sites and modes of each member are different. Furthermore, due to the high structural similarity of CaMKs, their members may play synergistic roles in the regulation of neural development, but different CaMKs also have their own means of regulating neural development. In this review, we first describe the visualized protein structural forms of CaMKI, CaMKII, eEF2K and CaMKIV, and then describe the functions of each kinase in neurodevelopment. After that, we focus on four main mechanisms of neurodevelopmental damage caused by CaMKs: CaMKI/ERK/CREB pathway inhibition leading to dendritic spine structural damage; Ca²⁺/CaM/CaMKII through induction of mitochondrial kinetic disorders leading to neurodevelopmental damage; CaMKIII/eEF2 hyperphosphorylation affects the establishment of synaptic plasticity; and CaMKIV/JNK/NF-κB through induction of an inflammatory response leading to neurodevelopmental damage. In conclusion, we briefly discuss the pathophysiological significance of aberrant CaMK family expression in neurodevelopmental disorders, as well as the protective effects of conventional CaMKII and CaMKIII antagonists against neurodevelopmental injury.

Reproductive toxicity is one of the important issues in chemical safety. Traditional laboratory testing methods are costly and time-consuming with raised ethical issues. Only a few in silico models have been reported to predict human reproductive toxicity, but none of them make full use of the topological information of compounds. In addition, most existing atom-based graph neural network methods focus on attributing model predictions to individual nodes or edges rather than chemically meaningful fragments or substructures. In current studies, we develop a novel fragment-based graph transformer network (FGTN) approach to generate the QSAR model of human reproductive toxicity by considering internal topological structure information of compounds. In the FGTN model, the compound is represented by a graph architecture using fragments to be nodes and bonds linking two fragments to be edges. A super molecule-level node is further proposed to connect all fragment nodes by undirected edges, obtaining global molecular features from fragment embeddings. The FGTN model achieved an accuracy (ACC) of 0.861 and an area under the receiver operating characteristic curve (AUC) value of 0.914 on nonredundant blind tests, outperforming traditional fingerprint-based machine learning models and atom-based GCN model. The FGTN model can attribute toxic predictions to fragments, generating specific structural alerts for the positive compound. Moreover, FGTN may also have the capability to distinguish various chemical isomers. We believe that FGTN can be used as a reliable and effective tool for human reproductive toxicity prediction in contribution to the advancement of chemical safety assessment.

The cGAS–STING pathway plays an essential role in the activation of tumor immune cells. Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants with potential carcinogenicity, and their exposure is associated with the development of colorectal cancer. However, the impacts of genetic factors in the cGAS‒STING pathway and gene‒environment interactions on colorectal cancer remain understudied. We used logistic regression models and interaction analysis to evaluate the impact of genetic variants on colorectal cancer risk and gene‒environment interactions. We analysed the expression patterns of candidate genes based on the RNA-seq data. Molecular biology experiments were performed to investigate the impact of PAHs exposure on candidate gene expression and the progression of colorectal cancer. We identified the susceptibility locus rs3750511 in the cGAS‒STING pathway, which is associated with colorectal cancer risk. A negative interaction between TRAF2 rs3750511 and PAHs exposure was also identified. Single-cell RNA-seq analysis revealed significantly elevated expression of TRAF2 in colorectal cancer tissues compared with normal tissues, especially in T cells. BPDE exposure increased TRAF2 expression and the malignant phenotype of colorectal cancer cells. The treatment also further increased the expression of the TRAF2 downstream gene NF-κB and decreased the expression of Caspase8. Our results suggest that the genetic variant of rs3750511 affects the expression of TRAF2, thereby increasing the risk of colorectal cancer through interaction with PAHs. Our study provides new insights into the influence of gene‒environment interactions on the risk of developing colorectal cancer.

Fullerene whiskers (FLW)s are thin rod-like structures composed of C₆₀ and C₇₀ fullerene (FL). The shape of FLWs suggests potential toxic effects including carcinogenicity to the lung and pleura, similar to effects elicited by asbestos and multi-walled carbon nanotubes (MWCNT)s. However, no long-term carcinogenic studies of FL or FLW have been conducted. In the present study we investigated the pulmonary and pleural carcinogenicity of FL and FLW. Twelve-week-old male F344 rats were administered 0.25 or 0.5 mg FL, FLW, MWCNT-7, and MWCNT-N by intra-tracheal intra-pulmonary spraying (TIPS). Acute lung lesions and carcinogenicity were analyzed at 1 and 104 weeks after 8 doses/15 days TIPS administration. At week 1, FLW, MWCNT-7, and MWCNT-N significantly increased alveolar macrophage infiltration. Expression of Ccl2 and Ccl3, reactive oxygen species production, and cell proliferation were significantly increased by administration of MWCNT-7 and MWCNT-N but not FL or FLW. At week 104, the incidence of bronchiolo-alveolar adenoma plus adenocarcinoma was significantly increased in the MWCNT-7 and MWCNT-N groups, and the incidence of mesothelioma was significantly increased in the MWCNT-7 group. No significant induction of pulmonary or pleural tumorigenesis was observed in the FL or FLW groups. The number of 8-OHdG-positive cells in the alveolar epithelium was significantly increased in the MWCNT-7 and MWCNT-N groups but not in the FL or FLW groups. FL and FLW did not exert pulmonary or pleural carcinogenicity in our study. In addition, oxidative DNA damage was implicated in MWCNT-induced lung carcinogenesis, suggesting that it may be a useful initial marker of carcinogenicity.

Chemotherapeutics are most often used to treat cancer, but side effects, drug resistance, and toxicity often compromise their effectiveness. In contrast, phytocompound plumbagin possesses a distinct pleiotropic nature, targeting multiple signaling pathways, such as ROS generation, cell death, cellular proliferation, metastasis, and drug resistance, and is shown to enhance the efficacy of chemotherapeutic drugs. Plumbagin has been shown to act synergistically with various chemotherapeutic drugs and enhance their efficacy in drug-resistant cancers. The pleiotropic nature is believed to be due to plumbagin's unique structure, which contains a naphthoquinone ring and a hydroxyl group responsible for plumbagin's various biological responses. Despite limitations such as restricted bioavailability and delivery, recent developments aim to address these challenges and harness the potential of plumbagin as an anticancer therapeutics. This review delves into the structural aspect of the plumbagin molecule contributing to its pleiotropic nature, explores the diverse mechanism that it targets, and discusses emerging strategies to overcome its limitations.

Fluoride-induced toxicity (fluorosis) poses a significant health concern globally, affecting millions of individuals. Understanding the molecular mechanisms underlying fluorosis, particularly the role of microRNAs (miRNAs), is crucial for developing effective preventive and therapeutic strategies. This review explores the pivotal role of miRNAs in the pathogenesis of fluorosis, particularly examining its impact on both hard (skeletal and dental) and soft (brain, liver, kidney, heart, and reproductive organs) tissues. Skeletal fluorosis manifests as abnormal bone mineralization and structure, while dental fluorosis affects enamel formation. In vitro and in vivo studies suggest a significant involvement of miRNAs in the progression of these conditions. For skeletal fluorosis, miR-124, miR-155, and miR-200c-3p have been identified as key regulators, while miR-296-5p and miR-214-3p are implicated in dental fluorosis. Moreover, soft tissue fluorosis encompasses a spectrum of adverse effects on various organs, including the brain, liver, kidneys, heart, and reproductive system. In soft tissues, miRNAs, such as miR-124, miR-200c-3p, miR-132, and miR-34b-5p, have been linked to cellular damage and dysfunction. Notably, miRNAs exert their effects through the modulation of critical pathways involved in fluorosis pathology, including Wnt signaling, apoptosis, cell cycle, and autophagy. Understanding the regulatory roles of miRNAs in fluorosis pathogenesis holds promise for identifying biomarkers and therapeutic targets. However, further research is needed to elucidate the molecular mechanisms underlying miRNA-mediated responses to fluoride exposure. Integration of miRNA research into fluorosis studies could facilitate the development of diagnostic tools and therapeutic interventions, thus mitigating the detrimental effects of fluorosis on both hard and soft tissues.

Proteomic investigations result in high dimensional datasets, but integration or comparison of different studies is hampered by high variances due to different experimental setups. In addition, cell culture conditions can have a huge impact on the outcome. This study systematically investigates the impact of experimental parameters on the proteomic profiles of commonly used cell lines—A549, differentiated THP-1 macrophage-like cells, and NR8383—for toxicity studies. The work focuses on analyzing the influence at the proteome level of cell culture setup involving different vessels, cell passage numbers, and post-differentiation harvesting time, aiming to improve the reliability of proteomic analyses for hazard assessment. Mass-spectrometry-based proteomics was utilized for accurate protein quantification by means of a label-free approach. Our results showed that significant proteome variations occur when cells are cultivated under different setups. Further analysis of these variations revealed their association to specific cellular pathways related to protein misfolding, oxidative stress, and proteasome activity. Conversely, the influence of cell passage numbers on the proteome is minor, suggesting a reliable range for conducting reproducible biological replicates. Notable, substantial proteome alterations occur over-time post-differentiation of dTHP-1 cells, particularly impacting pathways crucial for macrophage function. This finding is key for the interpretation of experimental results. These results highlight the need for standardized culture conditions in proteomic-based evaluations of treatment effects to ensure reliable results, a prerequisite for achieving regulatory acceptance of proteomics data.

Man-made vitreous fibers (MMVF) comprise diverse materials for thermal and acoustic insulation, including stone wool. Depending on dimension, durability, and dose, MMVF might induce adverse health effects. Therefore, early predictive in vitro (geno)toxicity screening of new MMVF is highly desired to ensure safety for exposed workers and consumers. Here, we investigated, as a starting point, critical in vitro screening determinants and pitfalls using primary rat alveolar macrophages (AM) and normal rat mesothelial cells (NRM2). A stone wool fiber (RIF56008) served as an exemplary MMVF (fibrous vs. ground to estimate impact of fiber shape) and long amosite (asbestos) as insoluble fiber reference. Materials were comprehensively characterized, and in vivo-relevant in vitro concentrations defined, based on different approaches (low to supposed overload: 0.5, 5 and 50 µg/cm²). After 4–48 h of incubation, certain readouts were analyzed and material uptake was investigated by light and fluorescence-coupled darkfield microscopy. DNA-strand break induction was not morphology-dependent and nearly absent in both cell types. However, NRM2 demonstrated material-, morphology- and concentration-dependent membrane damage, CINC-1 release, reduction in cell count, and induction of binucleated cells (asbestos > RIF56008 > RIF56008 ground). In contrast to NRM2, asbestos was nearly inactive in AM, with CINC-1 release solely induced by RIF56008. In conclusion, to define an MMVF-adapted, predictive in vitro (geno)toxicity screening tool, references, endpoints, and concentrations should be carefully chosen, based on in vivo relevance, and sensitivity and specificity of the chosen cell model. Next, further endpoints should be evaluated, ideally with validation by in vivo data regarding their predictivity.

This study investigated the impact of PM_2.5 on promoting EMT in PM_2.5-induced pulmonary fibrosis (PF) development and explored molecular mechanisms of the IL-9/STAT3/Snail/TWIST1 signaling pathway in PF owing to PM_2.5. Four groups of male SD rats were formed: control (0 mg/kg.bw), low (1 mg/kg.bw), medium (5 mg/kg.bw), and high-dose (25 mg/kg.bw) PM_2.5 groups. Experimental rats were subjected to PM_2.5 exposure via intratracheal instillation, given once weekly for 16 weeks. 24 h after the final exposure, blood, BALF, and lung tissues were collected. Pulmonary epithelial cells underwent cultivation and exposure to varying PM_2.5 concentrations with/without inhibitors for 24 h, after which total protein was extracted for relevant protein assays. The findings demonstrated that PM_2.5 damaged lung tissue to different degrees and led to PF in rats. Rats subjected to PM_2.5 exposure exhibited elevated concentrations of IL-9 protein in both serum and BALF, and elevated levels of IL-9 and its receptor, IL-9R, in lung tissues, compared to control counterparts. Furthermore, PM_2.5-exposed groups demonstrated significantly augmented protein levels of p-STAT3, Snail, TWIST1, Vimentin, COL-I, and α-SMA, while displaying notably diminished levels of E-Cadherin compared to control group. The same findings were observed in PM_2.5-treated cells. In BEAS-2B cells co-treated with Stattic (STAT3 inhibitor) and PM_2.5, the opposite results occurred. Similar results were obtained for cells co-treated with IL-9-neutralizing antibody and PM_2.5. Our findings suggest PM_2.5 mediates PF development by promoting IL-9 expression, leading to STAT3 phosphorylation and upregulation of Snail and TWIST1 expression, triggering EMT occurrence and progression in lung epithelial cells.

In recent years, nitrosamine impurities in pharmaceuticals have been subject to intense regulatory scrutiny, with nitrosamine drug substance-related impurities (NDSRIs) treated as cohort of concern impurities, regardless of predicted mutagenic potential. Here, we describe a case study of the NDSRI N-nitroso-hydrochlorothiazide (NO-HCTZ), which was positive in the bacterial reverse mutation (Ames) test but is unstable under the test conditions, generating formaldehyde among other products. The mutagenic profile of NO-HCTZ was inconsistent with that expected of a mutagenic nitrosamine, exhibiting mutagenicity in the absence of metabolic activation, and instead aligned well with that of formaldehyde. To assess further, a modified Ames system including glutathione (3.3 mg/plate) to remove formaldehyde was developed. Strains used were S. typhimurium TA98, TA100, TA1535, and TA1537, and E. coli WP2 uvrA/pKM101. In this system, formaldehyde levels were considerably lower, with a concomitant increase in levels of S-(hydroxymethyl)glutathione (the adduct formed between glutathione and formaldehyde). Upon retesting NO-HCTZ in the modified system (1.6–5000 µg/plate), a clear decrease in the mutagenic response was observed in the strains in which NO-HCTZ was mutagenic in the original system (TA98, TA100, and WP2 uvrA/pKM101), indicating that formaldehyde drives the response, not NO-HCTZ. In strain TA1535, an increase in revertant colonies was observed in the modified system, likely due to a thiatriazine degradation product formed from NO-HCTZ under Ames test conditions. Overall, these data support a non-mutagenic designation for NO-HCTZ and demonstrate the value of further investigation when a positive Ames result does not align with the expected profile.