NAM journal最新文献

We applied the TempO-LINC® platform to generate single-cell transcriptomic (SCTr) profiles of ∼40,000 HepaRG cells exposed to etoposide, brefeldin A, cycloheximide, rotenone, tBHQ, troglitazone, and tunicamycin at three concentrations for 24 hours. SCTr enabled a detailed analysis of adaptive stress response pathways (SRPs), including the unfolded protein response (UPR), oxidative stress response (OSR), heat shock response (HSR), and DNA damage response (DDR). Troglitazone upregulated lipid metabolism genes (PLIN2, ACOX1) along with HSR and UPR activation, with co-expression of DNAJA1, HSP90AA1, and DDIT3 in subsets of cells. Brefeldin A and tunicamycin strongly induced UPR markers (HSPA5, SYVN1, LMF2, PDIA4) in subsets of cells, with some also expressing apoptotic (DDIT3, CASP8) and autophagic (SQSTM1) genes, indicating diverse stress responses. Rotenone activated GDF15, TRIB3, and DDIT3 in a fraction of cells, accompanied by PLIN2 and mild UPR induction, reflecting heterogeneous mitochondrial stress responses. We scored individual cells using literature-derived SRP gene signatures to characterize overall stress phenotypes and clustered them using a generalized Jaccard metric. The clustering revealed five phenotypic groups spanning cell states associated with homeostasis, adaptive responses, terminal outcomes, autophagy, and apoptosis. By systematically analyzing the distributions of cells in different states across treatments, we visualized dynamic shifts in cellular subpopulations responding to chemicals, revealing early stress responses and potential transitions to cell death. Our findings suggest the utility of SCTr in decoding stress states that could provide possible insights into transitions between cellular adaptive and terminal transitions involved in toxicity.

This study aims to elucidate the mechanisms underlying bisphenols (BPs)-induced neurodegeneration and their contribution to neurodegenerative diseases. Focusing on four major disorders-Alzheimer's Disease, Parkinson's Disease, Amyotrophic Lateral Sclerosis, and Huntington's Disease-we systematically examined key molecular pathways potentially perturbed by BPs during disease progression. Preliminary toxicological profiling of four representative BPs was conducted using ProTox-3.0, ADMETlab 3.0, and the Xundrug database. Subsequent target identification involved integrated analyses of multiple bioinformatics resources, including CHEMBL and STITCH. Protein-protein interaction networks constructed with STRING and Cytoscape identified core targets such as HSP90AA1, ESR1, BCL2, and PTGS2. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses further revealed critical biological processes, including enzyme binding and heme binding, as well as key pathways associated with BPs neurotoxicity, such as chemical carcinogenesis-receptor activation, chemical carcinogenesis-DNA adducts, and arachidonic acid metabolism. Molecular docking studies demonstrated strong binding affinities between BPs and core targets, supported by low free energy values. Molecular dynamics simulations further validated stable binding conformations and dynamic interactions. Additionally, we analyzed regulatory networks of mRNA-miRNA-lncRNA interactions for core targets. In summary, our findings establish a novel multi-target and multi-pathway framework for BPs-induced neurodegeneration, revealing synergistic effects of pathways including carcinogenic signaling activation and metabolic dysregulation. This study advances understanding of environmental neurotoxicity and provides a foundation for developing preventive strategies against neurodegenerative diseases.

In the search for alternatives to replace in vivo studies, the application of assessment frameworks involving New Approach Methodologies (NAMs) often leads to the simultaneous availability of multiple pieces of evidence of different quality. When integrated into defined data structures, combinations of NAMs can generate answers to complex toxicological questions. However, if they are not integrated correctly, a collection of NAM results may produce misleading results that complicate the assessment process or lead to wrong conclusions. To support transparent decision-making in situations in which multiple NAMs are applied to generate results for the same toxicological question, we developed TOXTRUST (www.github.com/phi-grib/TOXTRUST) - an open-source computational tool integrating the mathematical framework of the Dempster-Shafer Theory (DST). In this article, we briefly describe the DST framework for the integration of NAM results to define the scope of its application and data requirements. This is followed by a description of the functionalities and infrastructure of TOXTRUST. Lastly, we illustrate how TOXTRUST can be applied to any endpoint with binary end-results, with a focus on the generation and interpretation of results expressed through probability bounds.

Objective: This study aims to investigate the potential mechanisms by which EDCs, recognized as emerging pollutants, induce metabolic disorders leading to metabolic diseases in humans.

Methods: Network toxicology and molecular docking techniques were employed to elucidate the molecular mechanisms underlying EDCs-induced pathogenesis of the six diseases. Potential targets associated with EDCs and these diseases were identified using databases such as PubChem, ChEMBL, Super-PRED, GeneCards, OMIM, and TTD. STRING analysis and Cytoscape software were further utilized to determine core targets most significantly linked to these metabolic disorders. GO and KEGG pathway enrichment analyses were performed on the core targets using the DAVID database. Finally, molecular docking was conducted to validate the binding affinities between EDCs and core target proteins.

Results: EDCs may potentially induce metabolic disorders by modulating cellular expression, influencing apoptosis and proliferation, and regulating related signaling pathways. Notably, a close interrelationship was observed among lipid metabolism disorders and atherosclerosis, Alzheimer's disease, type 2 diabetes mellitus, osteoporosis, hyperuricemia, and non-alcoholic fatty liver disease.

Conclusion: This study provides novel insights into the mechanisms through which EDCs induce metabolic diseases in humans and highlights correlations among distinct disorders, thereby establishing a theoretical foundation for disease prevention and therapeutic strategies.

While peer review during manuscript or grant application forms the cornerstone of scientific evaluation and assessment, this process is also prone to biases and confounding. Animal methods bias is a specific form of peer review bias where a preference for animal-based research methods or a lack of expertise in nonanimal-based methods undermines the quality and fairness of assessments of nonanimal studies. In this study, we conducted a survey to assess the challenges and experiences of researchers in India to publish peer-reviewed publications and apply for grants based on projects using nonanimal methods. A cross-sectional survey with 19 questions was completed by 186 respondents working in various biological fields in India. Decision logic was used to route respondents through the survey, which led to varying number of responses per question. Fifty-six per cent of respondents (39 out of 70) said they have been asked by manuscript reviewers to add animal experiments to their otherwise nonanimal-based studies. Respondents reported complying to 24 % of these requests on average. Respondents indicated the primary impacts of requests for additional experiments were publishing in lower impact factor journals and manuscript rejection. In addition, 57 % (47 of 83 respondents) felt that the lack of animal experiments in their grant proposal negatively influenced its evaluation. Respondents were also asked about key factors influencing their use of animal and nonanimal methods, which revealed that some perceived animal methods as necessary to validate nonanimal methods and as more reliable for mimicking biological complexity, while others perceived nonanimal methods as more physiologically relevant and practically advantageous. This survey provides preliminary evidence of animal methods bias experiences during publishing and funding peer review faced by Indian researchers.

The search for an alternative in vivo diagnostic method for Clostridium perfringens toxinotype D (CPD) aimed to evaluate the susceptibility of the nematode Caenorhabditis elegans CF512 CPD as a substitute for conventional biological models. The LD50 of the activated filtrate from CPD strain 426 was determined using a 96-well plate method over 23 h. We performed seroneutralization test with polyclonal anti-CPD toxin to determine the ND50. We observed significant differences between the toxin as the only factor and the toxin combined with polyclonal antibodies between 11 and 14 h after the test began. The evaluation of CPD toxin toxicity, as indicated by the survival or death of the nematode, suggests the potential utility of this model for future diagnostic applications. Although these initial findings provide insight into the CPD toxin's effect in this model, other toxins in the filtrate caused delayed effects. Future investigations employing purified toxins are expected to yield more precise data regarding mortality and neutralization. This study constitutes the inaugural evaluation of CPD effects within the C. elegans model, thereby offering direct implications for the biological diagnosis of this toxin and potentially enhancing the accuracy and efficacy of enterotoxemia detection and treatment.

Botulinum neurotoxin (BoNT), commonly referred to as 'Botox', is widely used in medical and aesthetic applications across the globe. As a biological product, BoNT requires rigorous batch potency testing to ensure safety, and is currently reliant on the ethically and scientifically contentious Mouse Lethality Bioassay (MLB). Despite severe suffering of the mice, the reproducibility crisis, and translatability issues of this Lethal Dose (LD50) test, its use persists due to regulatory inertia and other systemic barriers. Recent advancements in new approach methodologies (NAMs), particularly cell-based assays (CBAs), offer promising non-animal alternatives for regulatory potency testing. This paper seeks to provide an update on the transition to non-animal NAMs with a primary focus on Europe as a key player in the production of BoNT products. Namely, it describes the (accepted) alternatives to the MLB, the distribution of BoNT products and use of mice in the European Union and United Kingdom, and attempts to understand the number of mice used per year for BoNT alone, by analyzing Non-Technical Summaries (NTSs) from known testing facilities across the UK, Germany, and Ireland. This paper synthesizes findings from publicly available resources to provide actionable insights into overcoming barriers to implementing ethical and scientifically robust alternatives for regulatory testing of BoNT products. Based on NTS data, the MLB has likely ceased in Germany but persists in the UK and Ireland. Our findings show that even when companies have developed CBAs, MLB use may continue for various factors such as testing for reference standards, comparability testing/validation of CBAs, a lack of universal alternatives, and compliance with regulations for market authorizations of new BoNT products. Finally, predicting the number of mice used for BoNT potency testing was strikingly difficult, especially given the severity of the LD50 test, demonstrating the need for greater transparency in actual animal use for (severe) procedures and the limited applicability of public data to predict actual animal use for more specific research questions.

Brain exposure to toxic agents is a key condition for neurotoxicity hazards in the human central nervous system (CNS). Therefore, reliable prediction of brain penetration is needed to confirm the toxicological safety of compounds to be developed. However, current regulatory risk assessments of neurotoxicity are mainly performed in rodents, which entail several critical issues associated with animal studies. Consequently, authorities worldwide have highlighted the importance of harmonized, human-relevant new approach methodologies (NAMs) such as human induced pluripotent stem cell (hiPSC)-derived models to estimate substrate permeabilities concerning the blood-brain barrier (BBB). This study aimed to redesign a commercially available human BBB model and permeability testing method for future regulatory neurotoxicity studies, considering the in vivo architecture and functionality as well as potential for standardization and higher throughput. Model validation should be achieved by correlation of in vitro permeability to human clinical positron emission tomography (PET) data to enable reliable prediction of in vivo brain penetration. Accordingly, cryopreserved hiPSC-derived BBB models were set up within five days in 96-transwells®, quality-controlled for in vivo barrier properties and exposed to seven pharmaceuticals analogously to clinical PET studies (exposure concentrations: 5 µM to 10 µM for 60 min). The results indicated that the redesigned in vitro permeability assessment model allows to emulate human in vivo brain penetration (Spearman rank correlation coefficient: 0.964). This suggests that utilizing novel commercially available, cryopreserved hiPSC-derived BBB cells could be an important starting point for a standardizable NAMs application to be implemented in regulatory neurotoxicity hazard assessment.

A new R package, ctxR, is presented. ctxR is an open-source R Client package used to interact with the Computational Toxicology and Exposure Application Programming Interfaces (CTX APIs) in R. The CTX APIs were developed by the US EPA's Center for Computational Toxicology and Exposure, and provide users transparent, reproducible access in a programmatic manner to the data surfaced on the CompTox Chemicals Dashboard (CCD) and other data sources. The utility of this package is demonstrated through a case study that compares traditional Points of Departure (PODs) and PODs based on New Approach Methods (NAMs), referred to as PODNAM, relative to estimates of external exposure as an indicator of level of risk. This analysis combines a wide variety of data and showcases the power of programmatic data access that ctxR can provide to the scientific community.

Liver cirrhosis and hepatocellular cancer are brought on by the hepatitis C virus (HCV). Since the virus's discovery, significant therapeutic advancements have been made. Recent studies have demonstrated the growing importance of plant compounds in the creation of novel, efficient, and reasonably priced anti-Hepatitis C virus therapies. The present study involved a thorough analysis of 35 Quinoline-2-yl (piperazin-1-yl) compounds. These compounds were computationally analyzed using in-silico methods like 2D-3D QSAR modeling and molecular docking, and their results were verified through the use of Density Functional Theory (DFT) calculations and ADMET characteristics assessment. The inhibitors were optimized using DFT based on a notion of B3LYP/6-31G* levels. The genetic function algorithm (GFA) was utilized to create the QSAR models. The best model was chosen based on its statistical fitness using the subsequent measurement parameters: R²trng = 0.8328, R²adj = 0.798036, Q²cv = 0.765153, and R²test = 0.6510 led to its selection and publication. A reliable 2D QSAR model was constructed and validated using an external validation test set, Y-randomization, variance inflation factor (VIF), mean effect (MF), and William's plot applicability domain (AD). The virtual screening of the compounds under study revealed that compound 29 was the top hit, with the highest mole dock score of -144.45 kcal/mol. A structure-based method was used to create six novel compounds with a higher affinity (SFBD3 -152.299 kcal/mol) for the NS5B polymerase. The food drug administration (FDA) drug Rib/Pig (-111.095 kcal/mol) has lower mole dock scores than the newly created compounds. Molecular dynamics (MD) simulations evaluated the binding stability and dynamic behavior of compounds 29 and SFD B3 with HCV NS5B polymerase (PDB ID: 3QK7) over 100 ns, using Desmond. Analyses of RMSD, RMSF, and hydrogen bonding revealed stable binding for both compounds. SFD B3 demonstrated superior stability (avg RMSD 1.92 Å, 2.91 H-bonds) compared to compound 29 (avg RMSD 2.32 Å, 0.94 H-bonds), indicating more favorable interaction dynamics within the NS5B active site. According to the synthetic accessibility score, the newly discovered compounds may be readily made in a wet lab, have excellent pharmacokinetic profiles, and have a bioavailability score of 0.55 based on the anticipated drug-likeness and ADMET properties. As potential Hepatitis C viral agents, the created molecule SFBD3, which is stable, has a higher affinity, and has the best pharmacokinetic characteristics, ought to be made in a wet lab.