It is logistically impossible to perform toxicity testing of all chemicals in all species. Therefore, chemical safety evaluations must find ways to make inferences about toxicity in untested species based on observations from model species. Development of bioinformatics tools that create scientific lines of evidence for extrapolation of toxicity knowledge across species are gaining traction in decision-making surrounding chemical safety. These approaches take advantage of computational power and allow for incorporation of large-scale omics data to infer conservation of biological pathways across species to enable predictions of chemical susceptibility and further understand potential toxicity. As comparative toxicity data from such approaches are integrated into regulatory decisions to support surrogacy challenges it is important that the tools being developed for and applied in regulation meet necessary standards identified by hazard and risk assessment practitioners. The International Consortium to Advance Cross Species Extrapolation in Regulation (ICACSER), which consists of researchers, tool developers, and regulatory decision-makers seeks to highlight the foundational role of bioinformatics for enhanced scientific evidence to inform chemical safety decisions across species. To articulate essential standards and define best practices in development and application of bioinformatics approaches specific to this context, three existing tools are examined, and their key attributes are highlighted and connections to the adverse outcome pathway framework are discussed. These practices are intended to act as a guide for current and future tool development.

Traumatic brain injury (TBI) is a major global health issue, with approximately 27 million new cases annually. TBI significantly increases the risk of developing post-traumatic epilepsy (PTE), which may emerge weeks to years after the initial injury. Currently, effective therapies for both TBI and PTE remain limited. Extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) have demonstrated anti-inflammatory and neuroprotective effects in preclinical TBI models. VivaZome's proprietary cells (VZT-PCs) share characteristics with MSCs, including similar surface markers and immunomodulatory properties. Native EVs from VZT-PCs are enriched in microRNAs with known anti-inflammatory properties. This study assessed the therapeutic effect of VZT-PC-derived EVs in a murine model of TBI. Adult C57BL/6 mice underwent controlled cortical impact (CCI) and received either vehicle control or VZT-PC-EVs. Functional recovery was assessed using the Rotarod test and the Elevated Plus maze (EPM) test. After 4 weeks post TBI induction, PTE was evaluated using pentylenetetrazol challenge and video-electroencephalography. Native EV-treated TBI mice showed improved performance in Rotarod trials, where latency to fall was significantly increased at 24 h post injury, indicating enhanced motor coordination. Exploratory behavior in the EPM was also improved following EV administration, indicating mice exhibited reduced anxiety-like behavior. EV-treated mice exhibited a trend toward reduced seizure frequency and increased latency to generalized seizures, though not statistically significant. Collectively, these findings support the therapeutic potential of VZT-PC-derived native EVs in enhancing motor function and possibly reducing epileptogenicity following moderate TBI, highlighting VZT-PCs as a promising source of EVs for brain injury therapy.

The adverse outcome pathway (AOP) framework contributes to understanding how specific and measurable biological perturbations cause adverse effects on human and environmental health. Recognizing the value of AOPs to support regulatory decisions around the world, the Organisation for Economic Co-operation and Development (OECD) launched the AOP Programme in 2012, which sought to promote and guide the development of AOPs to ensure their suitability for the downstream applications in the context of regulatory safety assessment. The OECD published the initial guidance on AOP development and assessment in 2013, which has been expanded as practices have evolved and matured. Adverse outcome pathway development requires adherence to specific principles and considerations for identifying and describing key events (KEs) and representing and assessing the weight of evidence for the key event relationships. Ultimately, the structured and consistent application of the principles helps build confidence in the applicability of the knowledge represented in the AOP for decision-making in the regulatory context. To assist new AOP developers, in 2019, the OECD introduced a coaching program. This program primarily aims to pair novices with experienced AOP developers (i.e., coaches). International partnerships in the coaching program contribute to harmonizing and promoting AOP development according to OECD guidance. Coaches have also helped to identify and initiate “gardening” efforts that remove redundant/synonymous KEs in the AOP-Wiki, allowing for improved AOP network creation, promoting the reuse of extensively reviewed KEs, and ensuring the development of high-quality AOPs. The AOP Coaching Program represents the latest international activity to ensure that AOPs are developed in a consistent manner that is designed to enhance their use for supporting public health decisions around the world.

The long-term sequelae of severe penetrating traumatic brain injury (TBI) include neurological and psychiatric disability, impaired cognitive function, and the development of post-traumatic epilepsy. The present study evaluated the therapeutic effects of intravenous immunoglobin (IVIg), a well-established immunomodulatory treatment, in a controlled cortical impact model of severe TBI in mice. The beneficial effects of IVIg treatment on acute neurological status, motor function, anxiety level, and spatial learning ability were demonstrated by reduced Neurological Severity Scores, increased Rotarod latency and cumulative movement durations in open-field tests, and improved active place avoidance performance. IVIg treatment also significantly reduced brain tissue loss, which was examined using Nissl staining at 16 weeks after TBI. Furthermore, brain microRNAs (miRNAs) were profiled to identify the biological pathways potentially associated with the actions of IVIg treatment using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis. To identify potential peripheral biomarkers reflecting the changes in the brain, differentially expressed miRNAs in plasma and brain samples from the same animals were compared. Our immunostaining results showed that IVIg treatment significantly attenuated the upregulation of IL-1β and complement 3 (C3) and altered the activation of microglia and astrocytes. This proof-of-concept study provided strong evidence for the beneficial effects of IVIg treatment in severe penetrating TBI.

Adverse outcome pathways (AOPs) provide evidence for demonstrating and assessing causality between measurable toxicological mechanisms and human or environmental adverse effects. AOPs have gained increasing attention over the past decade and are believed to provide the necessary steppingstone for more effective risk assessment of chemicals and materials and moving beyond the need for animal testing. However, as with all types of data and knowledge today, AOPs need to be reusable by machines, i.e., machine-actionable, in order to reach their full impact potential. Machine-actionability is supported by the FAIR principles, which guide findability, accessibility, interoperability, and reusability of data and knowledge. Here, we describe why AOPs need to be FAIR and touch on aspects such as the improved visibility and the increased trust that FAIRification of AOPs provides.

C1 human-derived C1 esterase inhibitor (C1-INH) is a U.S. Food and Drig Administration?approved drug with anti-inflammatory actions. In the present study, we investigated the therapeutic effects of C1-INH on acute and chronic neurobehavioral outcomes and on seizures in the chronic stage in a mouse traumatic brain injury (TBI) model. Adult male CD1 mice were subjected to controlled cortical impact and randomly allocated to receive C1-INH or vehicle solution 1?h post-TBI. Effects of C1-INH treatment on inflammatory responses and brain damage after TBI were examined using the Cytometric Bead Array, C5a enzyme-linked immunosorbent assay, Fluoro-Jade C staining, and Nissl staining. Neurobehavioral outcomes after TBI were assessed with modified neurological severity scores, the rotarod and open field tests, and the active place avoidance task. Video-electroencephalographic monitoring was performed in the 15th and 16th weeks after TBI to document epileptic seizures. We found that C1-INH treatment reduced TNF? expression and alleviated brain damage. Treatment with C1-INH improved neurological functions, increased locomotor activity, alleviated anxiety-like behavior, and exhibited an effect on seizures in the chronic stage after TBI. These findings suggest that C1-INH has beneficial effects on the treatment of TBI.

A potent effector of innate immunity, the complement system contributes significantly to the pathophysiology of traumatic brain injury (TBI). This study investigated the role of the complement cascade in neurobehavioral outcomes and neuropathology after TBI. Agents acting at different levels of the complement system, including 1) C1 esterase inhibitor (C1-Inh), 2) CR2-Crry, an inhibitor of all pathways acting at C3, and 3) the selective C5aR1 antagonist, PMX205, were administered at 1?h post-TBI. Their effects were evaluated on motor function using the rotarod apparatus, cognitive function using the active place avoidance (APA) task, and brain lesion size at a chronic stage after controlled cortical impact injury in C5-sufficient (C5+/+) and C5-deficient (C5?/?) CD1 mice. In post-TBI C5+/+ mice, rotarod performance was improved by CR2-Crry, APA performance was improved by CR2-Crry and PMX205, and brain lesion size was reduced by PMX205. After TBI, C5?/? mice performed better in the APA task compared with C5+/+ mice. C5 deficiency enhanced the effect of C1-Inh on motor function and brain damage and the effect of CR2-Crry on brain damage after TBI. Our findings support critical roles for C3 in motor deficits, the C3/C5/C5aR1 axis in cognitive deficits, and C5aR1 signaling in brain damage after TBI. Findings suggest the combination of C5 inhibition with C1-Inh and CR2-Crry as potential therapeutic strategies in TBI.

Early life exposure to environmental chemicals can interfere with myelin formation in the developing brain, leading to neurological disorders. The Proteolipid Protein 1 (Plp1), Myelin Basic Protein (Mbp) and 2′,3′-Cyclic Nucleotide 3′Phosphodiesterase (Cnp) genes expressed in oligodendrocytes and involved in myelination processes can be useful biomarkers of potential developmental neurotoxicity. In an earlier study, we concluded that the reduction in the expression levels of Mbp splice variants in juvenile rat cerebellum following perinatal methylmercury (MeHg) exposure were compatible with an overall reduction of mature oligodendrocytes population. This observation prompted us to analyze the expression of Plp1 and Cnp in developing rat cerebellum to further confirm and investigate the toxic effects of MeHg on vulnerable oligodendrocytes. Splice variants of Plp1 in human and of Cnp in mouse are curated in NCBI RefSeq database, but not for rat. Lack of annotation of splice variants can pose significant challenge for the reliable quantification of gene expression levels in toxicological studies. Therefore, we applied a "comparative sequence analysis" approach, relying on annotated splice variants in human/mouse and on evolutionary conservation of intron–exon structures, to identify additional splice variants of Plp1 and Cnp in rat. Then, we confirmed their identity by nucleotide sequencing and characterized their temporal expression patterns during brain development by RT-PCR. The measurement of total transcripts and individual splice variants of Plp1 and Cnp in the cerebellum of MeHg-exposed rat pups revealed a relatively similar level of reduction in their expression levels. This study further confirms that perinatal exposure to MeHg can impact oligodendrocytes in pups. Based on these observations, we conclude that monitoring the expression of these oligodendrocyte-enriched genes can be useful to identify toxic chemicals affecting myelination.

The need to test chemicals in a timely and cost-effective manner has driven the development of new alternative methods (NAMs) that utilize in silico and in vitro approaches for toxicity prediction. There is a wealth of existing data from human studies that can aid in understanding the ability of NAMs to support chemical safety assessment. This study aims to streamline the integration of data from existing human cohorts by programmatically identifying related variables within each study. Study variables from the Atherosclerosis Risk in Communities (ARIC) study were clustered based on their correlation within the study. The quality of the clusters was evaluated via a combination of manual review and natural language processing (NLP). We identified 391 clusters including 3,285 variables. Manual review of the clusters containing more than one variable determined that human reviewers considered 95% of the clusters related to some degree. To evaluate potential bias in the human reviewers, clusters were also scored via NLP, which showed a high concordance with the human classification. Clusters were further consolidated into cluster groups using the Louvain community finding algorithm. Manual review of the cluster groups confirmed that clusters within a group were more related than clusters from different groups. Our data-driven approach can facilitate data harmonization and curation efforts by providing human annotators with groups of related variables reflecting the themes present in the data. Reviewing groups of related variables should increase efficiency of the human review, and the number of variables reviewed can be reduced by focusing curator attention on variable groups whose theme is relevant for the topic being studied.

Adverse Outcome Pathways (AOP) provide structured frameworks for the systematic organization of research data and knowledge. The AOP framework follows a set of key principles that allow for broad application across diverse disciplines related to human health, including toxicology, pharmacology, virology and medical research. The COVID-19 pandemic engages a great number of scientists world-wide and data is increasing with exponential speed. Diligent data management strategies are employed but approaches for systematically organizing the data-derived information and knowledge are lacking. We believe AOPs can play an important role in improving interpretation and efficient application of scientific understanding of COVID-19. Here, we outline a newly initiated effort, the CIAO project (https://www.ciao-covid.net/), to streamline collaboration between scientists across the world toward development of AOPs for COVID-19, and describe the overarching aims of the effort, as well as the expected outcomes and research support that they will provide.

Targeting neuroinflammation is a novel frontier in the prevention and treatment of epilepsy. A substantial body of evidence supports a key role for neuroinflammation in epileptogenesis, the pathological process that leads to the development and progression of spontaneous recurrent epileptic seizures. It is also well recognized that traumatic brain injury (TBI) induces a vigorous neuroinflammatory response and that a significant proportion of patients with TBI suffer from debilitating post-traumatic epilepsy. The complement system is a potent effector of innate immunity and a significant contributor to secondary tissue damage and to epileptogenesis following central nervous system injury. Several therapeutic agents targeting the complement system are already on the market to treat other central nervous system disorders or are well advanced in their development. The purpose of this review is to summarize findings on complement activation in experimental TBI and epilepsy models, highlighting the potential of drug repurposing in the development of therapeutics to ameliorate post-traumatic epileptogenesis.

In Alzheimer’s disease (AD) glial fibrillary acidic protein (GFAP) is expressed by reactive astrocytes surrounding β‐amyloid (Aβ) plaques, whereas brain‐derived neurotrophic factor (BDNF) levels are typically reduced. We compared the expression of GFAP, BDNF, and its precursor proBDNF in the dorsal hippocampus of two transgenic AD mouse models. APPSwe YAC mice expressing the APPSwe transgene on a yeast artificial chromosome (YAC) were assessed at age 4 and 21 months, and APPSwe/PS1dE9 mice co‐expressing mutant amyloid precursor protein (APPSwe) and presenilin‐1 (PS1dE9) were assessed at age 4 and 9 months. Significantly increased (1.4‐fold) GFAP expression was observed in APPSwe YAC c.f . wild‐type (Wt) mice aged 21 months, when Aβ deposition was first evident in these mice. In APPSwe/PS1dE9 mice aged 4 and 9 months, GFAP expression was significantly increased (1.6‐ and 3.1‐fold, respectively) c.f. Wt mice, and was associated with robust Aβ deposition at 9 months. BDNF expression was significantly lower in 4‐ and 21‐month old APPSwe YAC mice (0.8‐ and 0.6‐fold, respectively) c.f. age‐matched Wt mice, whereas proBDNF expression was significantly higher (10‐fold) in the APPSwe YAC c.f. Wt mice aged 21 months. In APPSwe/PS1dE9 mice aged 4 months, BDNF expression was significantly lower (0.4‐fold) c.f . age‐matched Wt mice and was equivalent to that in 9‐month old mice of both genotypes; proBDNF expression mirrored that of BDNF in this strain. These findings support a role for reactive astrocytes and neuroinflammation, rather than BDNF, in the spatial memory deficits previously reported for APPSwe YAC and APPSwe/PS1dE9 mice.

New approach methodologies (NAMs) in chemical safety evaluation are being explored to address the current public health implications of human environmental exposures to chemicals with limited or no data for assessment. For over a decade since a push toward "Toxicity Testing in the 21st Century," the field has focused on massive data generation efforts to inform computational approaches for preliminary hazard identification, adverse outcome pathways that link molecular initiating events and key events to apical outcomes, and high-throughput approaches to risk-based ratios of bioactivity and exposure to inform relative priority and safety assessment. Projects like the interagency Tox21 program and the US EPA ToxCast program have generated dose-response information on thousands of chemicals, identified and aggregated information from legacy systems, and created tools for access and analysis. The resulting information has been used to develop computational models as viable options for regulatory applications. This progress has introduced challenges in data management that are new, but not unique, to toxicology. Some of the key questions require critical thinking and solutions to promote semantic interoperability, including: (1) identification of bioactivity information from NAMs that might be related to a biological process; (2) identification of legacy hazard information that might be related to a key event or apical outcomes of interest; and, (3) integration of these NAM and traditional data for computational modeling and prediction of complex apical outcomes such as carcinogenesis. This work reviews a number of toxicology-related efforts specifically related to bioactivity and toxicological data interoperability based on the goals established by Findable, Accessible, Interoperable, and Reusable (FAIR) Data Principles. These efforts are essential to enable better integration of NAM and traditional toxicology information to support data-driven toxicology applications.

This study was undertaken to evaluate the use of ontology-based semantic mapping (OS-Mapping) in chemical toxicity assessment. Nineteen chemical-species phenotypic profiles (CSPPs) were constructed by ontologically annotating the toxicity responses reported in more than seven hundred published studies of ten chemicals on six vertebrate species. The CSPPs were semantically compared to more than 29,000 publicly available phenotypic profiles of genes, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways, and diseases based on a cross-species phenotype ontology. OS-Mapping was shown to differentiate chemical toxicities among themselves as well as within and across species. It also revealed cases of chemical by species interactions. In addition to confirming similar MOAs (mechanisms of action) for a few chemicals, OS-Mapping also generated novel insights into the MOAs underlying some seemingly different, yet phenotypically similar, classes of chemicals. The nature of a unified cross-species phenotype ontology and its representation of diverse knowledge domains allowed the construction of a complete phenotypic continuum for the 17α-ethynylestradiol_fathead minnow across the biological levels of organization, which complemented a similar one derived from the Comparative Toxicogenomics Database but based primarily on 17α-ethynylestradiol-induced molecular phenotypes. Overall, OS-Mapping has been demonstrated to offer a powerful approach to help bridge the gap between the molecular and non-molecular phenotypes of chemicals characterized by using high throughput or traditional omics methods and their apical endpoints of greater regulatory relevance, which are typically phenotypes found at the higher levels of biological organization. OS-Mapping also enables comparative toxicity assessment among chemicals, both within and across species. Furthermore, the semantic analysis of phenotypes can reveal additional novel MOAs for some well-known chemicals and discover candidate MOAs for chemicals that are less molecularly characterized. A full phenotypic continuum based on OS-Mapping will also be conducive to the future development of adverse outcome pathways. As phenomics continues to advance and the ontological annotation of literature becomes more automated, the power of OS-Mapping will be further enhanced.

A range of chemical exposures that resulted in the specific pathology of hepatic lipid dysfunction in rats were selected from DrugMatrix, a publicly available toxicogenomic database. Raw microarray data collected from these exposures were further analyzed using bioinformatic tools to generate a differentially expressed genes (DEGs) dataset associated with hepatic lipid dysfunction. Further analysis of the DEGs dataset resulted in 324 upregulated genes, and 275 genes that were down regulated. Meanwhile, 36 genes were either up regulated or down regulated in different chemical treatments. All identified genes were uploaded in the web application for Database for Annotation, Visualization and Integrated Discovery (DAVID) for gene ontology enrichments and to identify Kyoto Encyclopedia of Genes and Genome (KEGG) pathways. Some of the identified pathways included glycolysis/gluconeogenesis, steroid hormone biosynthesis, retinol metabolism, and metabolism of xenobiotics by cytochrome P450. The same DEGs dataset was also analyzed using Ingenuity Pathway Analysis (IPA) software. IPA identified several pathways including PXR/RXR activation, Aryl hydrocarbon receptor signaling, and xenobiotic metabolism signaling. Furthermore, the generated DEGs lists were uploaded into NCATS BioPlanet platform. Some of the identified pathways were related to fatty acid omega oxidation, lipid and lipoprotein metabolism, and adipogenesis. The enrichment and clarification of the pathways and biological networks obtained from the DEGs dataset provide prior knowledge on the underlying biological key events and molecular mechanisms for the computational development of putative adverse outcome pathways (AOPs) for hepatic lipid dysfunction as a precursor to hepatic steatosis.

Advancements in measurement technologies and modeling capabilities continue to result in an abundance of exposure information, adding to that currently in existence. However, fragmentation within the exposure science community acts as an obstacle for realizing the vision set forth in the National Research Council's report on Exposure Science in the 21st century to consider exposures from source to dose, on multiple levels of integration, and to multiple stressors. The concept of an Aggregate Exposure Pathway (AEP) was proposed as a framework for organizing and integrating diverse exposure information that exists across numerous repositories and among multiple scientific fields. A workshop held in May 2016 followed introduction of the AEP concept, allowing members of the exposure science community to provide extensive evaluation and feedback regarding the framework's structure, key components, and applications. The current work briefly introduces topics discussed at the workshop and attempts to address key challenges involved in refining this framework. The resulting evolution in the AEP framework's features allows for facilitating acquisition, integration, organization, and transparent application and communication of exposure knowledge in a manner that is independent of its ultimate use, thereby enabling reuse of such information in many applications.

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