Toxicol. Sci. [JOURNAL]
Sun
200 papers
Developmental exposure to a PFAS mixture impairs the anamnestic response to influenza A virus infection in mice.
Post CM, Agyemang JJ, McDonald KA … +2 more , McDonough CA, Lawrence BP
Toxicol Sci · 2026 Jun · PMID 42184288 · Full text
Developmental exposure to per- and polyfluoroalkyl substances (PFAS) has been linked to reduced antibody responses to childhood vaccines, but the underlying mechanisms remain unclear. Antibody production relies on intera...
Developmental exposure to per- and polyfluoroalkyl substances (PFAS) has been linked to reduced antibody responses to childhood vaccines, but the underlying mechanisms remain unclear. Antibody production relies on interactions between various immune cell types, and it is unknown which are affected by PFAS exposure during development. To investigate this in a human-health-relevant system, an in vivo model was established to delineate effects of developmental exposure to a mixture of 4 PFAS commonly found in human serum: Perfluorooctanoic acid (PFOA), perfluorooctane sulfonate, perfluorohexane sulfonate, and perfluorononanoic acid. Pregnant mice consumed water containing these PFAS throughout gestation and lactation. PFAS were measured in both mothers and offspring, and an exposure that did not cause signs of overt toxicity was selected. The immune response to influenza A virus (IAV) infection was assessed in male and female offspring. Results showed that developmental PFAS exposure reduced IAV-specific antibody levels in both sexes. However, it diminished T follicular helper cells and germinal center B cells-critical for antibody production-in only female offspring. These findings highlight possible sex-specific immune effects and identify potential cellular mechanisms behind reduced antibody levels. Because these immune cells are essential for antibody production in humans, this study provides valuable insights into how PFAS exposure may impact human health.
Stress-activated pathways mediate PFAS effects on human placental syncytiotrophoblast cells.
Kozai K, Varberg KM
Toxicol Sci · 2026 Jun · PMID 42172632 · Full text
Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants associated with placenta-mediated pregnancy complications, including preeclampsia, fetal growth restriction, and preterm birth. The syncy...
Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants associated with placenta-mediated pregnancy complications, including preeclampsia, fetal growth restriction, and preterm birth. The syncytiotrophoblast (STB), which forms the placental barrier at the maternal-fetal interface and is directly exposed to maternal blood, is a primary site of PFAS exposure. Although PFAS induce STB apoptosis, the upstream stress-signaling pathways involved remain poorly defined. Here, we investigated stress-responsive signaling mechanisms mediating PFAS-induced STB cell death. STB differentiated from human trophoblast stem cells were exposed to vehicle or an environmentally relevant mixture of 5 PFAS (perfluorooctanoic acid, perfluorooctanesulfonic acid, perfluorohexane sulfonate, perfluorononanoic acid, and perfluorodecanoic acid; 0.0138 to 34.5 µM) for 3 or 6 h. Cytotoxicity, apoptosis, mitochondrial membrane potential, and stress-signaling pathway activation were assessed by lactate dehydrogenase release, immunoblotting, JC-10 assay, and reverse transcription-quantitative PCR. PFAS mixtures did not induce cytotoxicity at 3 h but significantly increased cytotoxicity at 6 h at 34.5 µM, coinciding with the induction of cleaved caspase-3, cleaved poly(ADP-ribose) polymerase, and NOXA. The pan-caspase inhibitor z-VAD-FMK prevented cytotoxicity, indicating caspase-dependent apoptosis. PFAS exposure reduced mitochondrial membrane potential and activated the integrated stress response (ISR), as evidenced by eukaryotic initiation factor 2α phosphorylation, activating transcription factor 4 (ATF4) induction, and increased ATF4 target gene expression. In parallel, c-Jun N-terminal kinase (JNK) signaling was activated, as evidenced by JNK phosphorylation and induction of immediate-early genes (JUN, FOS, EGR1). Pharmacologic inhibition of the ISR modestly attenuated PFAS-induced cytotoxicity, whereas pharmacologic inhibition of JNK rescued cytotoxicity and apoptotic signaling. Together, these findings identify JNK-driven stress signaling as the dominant mediator of PFAS-induced STB apoptosis, with a secondary contribution from the ISR.
Safety evaluation of glutaric acid as a novel pharmaceutical salt for first-in-human dosing.
Franklin KM, Aboulmali A
Toxicol Sci · 2026 Jun · PMID 42166747 · Publisher ↗
Salt selection is critical in pharmaceutical development. Glutaric acid, an endogenous dicarboxylic acid, offers potential advantages as a counterion but is underexplored in pharmaceutical applications. The nonclinical s...
Salt selection is critical in pharmaceutical development. Glutaric acid, an endogenous dicarboxylic acid, offers potential advantages as a counterion but is underexplored in pharmaceutical applications. The nonclinical safety of glutaric acid as a pharmaceutical counterion was assessed using LY3972406 hemi-glutarate hemi-hydrate (HG) as a model, with direct comparison to the LY3972406-hydrochloride (HCl) salt. Comprehensive safety assessments included genetic toxicology (Ames, in vitro and in vivo micronucleus tests), hazard evaluation (phototoxicity, and dermal and corneal irritation), and repeat-dose toxicity studies in rats and dogs. Comparative studies evaluated LY3972406-HCl versus LY3972406-HG, and chronic studies assessed glutaric acid alone and as part of the LY3972406-HG salt. Glutaric acid was nongenotoxic. LY3972406-HG showed no phototoxicity and only mild, nonadverse dermal and ocular irritation in hazard assessments. Glutaric acid was well-tolerated by rats and dogs in repeat-dose toxicity studies, with no treatment-related adverse effects. When LY3972406-HG was administered at glutaric acid equivalent doses up to 31 mg/kg/d, all findings were nonadverse, and likely attributable to the active pharmaceutical ingredient LY3972406. Glutaric acid demonstrates a favorable nonclinical safety profile with no genotoxicity and was tolerated at up to 40 mg/kg/d in chronic toxicity studies. Direct comparison with HCl salt confirmed equivalent safety and exposure profiles, supporting glutaric acid as a viable alternative to HCl in pharmaceutical development. Glutaric acid is a promising pharmaceutical counterion that lacks regulatory safety data. This first comprehensive Good Laboratory Practice-compliant nonclinical safety evaluation supports the viability of glutaric acid as an alternative pharmaceutical counterion to hydrochloride salts in drug development.
Technical and biological factors driving inter-individual body burden of arsenic species in murine models of human arsenic exposure.
Wang L, Wang Q, Wolfe TM … +5 more , Pinkham NV, Erickson R, Yoshinaga M, McDermott TR, Walk ST
Toxicol Sci · 2026 Jun · PMID 42155039 · Full text
Arsenic is one of the most important environmental toxicants, requiring advanced analytical techniques to resolve individual species. There is little consensus on arsenic speciation methodology for in vivo studies. The o...
Arsenic is one of the most important environmental toxicants, requiring advanced analytical techniques to resolve individual species. There is little consensus on arsenic speciation methodology for in vivo studies. The objectives of this study were to generate a robust framework for arsenic speciation in murine models of human exposure and evaluate factors influencing the levels of arsenobetaine, inorganic arsenite, dimethylarsinate, monomethylarsonate, and inorganic arsenate resolved by high-performance liquid chromatography (HPLC)-inductively coupled plasma mass spectrometry (ICPMS). Enzyme-assisted digestion by papain vs. pepsin and maceration by bead beating vs. mechanical homogenization were evaluated using chemical standard spiking experiments. Dose-controlled mouse exposures to inorganic arsenite were conducted, and species detected in urine and bladder tissue were compared. Species in stool, liver, and bladder were compared between groups of mice eating a standard vs. purified diet, fasted vs. unfasted mice, and conventional vs. germ-free mice. Finally, between-lab differences in HPLC-ICPMS instrumentation/quantification procedures were evaluated. These comparisons led to several important conclusions, including: Significant conversion of inorganic arsenate to arsenite by papain, significant inorganic arsenate background in bead-beating lysing matrix, significant arsenobetaine in mice eating standard but not purified chow, significant correlation between species detected in urine and bladder, significant correlation of results between laboratories that differed in absolute quantification, and large inter-individual variability between mice of the same treatment group. Finally, diet type and the presence of a microbiome had the largest effect on arsenic species levels. Our results provide a benchmark for evaluating arsenic species in murine models, including adequate sample sizes for powering studies to avoid erroneous conclusions.
Cigarette smoke extract suppresses peptide transporter 2 function in alveolar epithelial cells via aryl hydrocarbon receptor signaling.
Kawami M, Nakanishi M, Furuichi Y … +3 more , Yumoto R, Uchida Y, Takano M
Toxicol Sci · 2026 May · PMID 42155038 · Publisher ↗
Cigarette smoke is a major risk factor for chronic respiratory diseases, yet the molecular mechanisms underlying its effects on pulmonary defense remain incompletely understood. Although transporters such as P-glycoprote...
Cigarette smoke is a major risk factor for chronic respiratory diseases, yet the molecular mechanisms underlying its effects on pulmonary defense remain incompletely understood. Although transporters such as P-glycoprotein and CFTR have been implicated in smoking-related dysfunction, little is known about the impact of cigarette smoke on peptide transporter 2 (PEPT2), which plays a dual role in xenobiotic disposition and host defense by mediating the uptake of di- and tripeptides, bacterial fragments, and peptide-like drugs. Here, we investigated the effects of cigarette smoke extract (CSE) on the functional expression of PEPT2 in alveolar epithelial cells. Using the human H441 cell line, we found that CSE reduced PEPT2 activity and mRNA expression in a concentration-dependent manner. Among signaling inhibitors, only PD98059 reversed the CSE-induced suppression, whereas U0126 had no effect. Consistently, the AhR ligand benzo[a]pyrene decreased PEPT2 expression and function, and this effect was abolished by AhR antagonists. Other AhR ligands, including leflunomide and omeprazole, produced similar inhibitory effects. These findings indicate that CSE suppresses PEPT2 in an AhR-dependent manner, whereas the underlying transcriptional mechanisms remain to be clarified. These findings suggest that PEPT2 is a previously unrecognized target of cigarette smoke-related transporter regulation and are consistent with the involvement of AhR signaling in its suppression. These findings provide insight into potential alterations in epithelial transport function and their possible implications for pulmonary defense and drug disposition in smokers.
Late gestational exposure to flutamide alters stromal composition and immune landscape in the rat mammary gland during pre-puberty, peri-puberty, and adulthood.
Tovar Parra D, McDermott A, Cardot J … +12 more , Juarez MN, Joao F, El Omri-Charai R, Berthiaume L, Dhawan B, Aghigh A, Breton Y, Légaré F, Delbès G, Pelletier M, Audet-Walsh É, Plante I
Toxicol Sci · 2026 May · PMID 42155037 · Full text
Perinatal development of the mammary gland is regulated by hormonal signals that influence cell proliferation, extracellular matrix remodeling, immune cell recruitment, and intracellular signaling. Although the role of e...
Perinatal development of the mammary gland is regulated by hormonal signals that influence cell proliferation, extracellular matrix remodeling, immune cell recruitment, and intracellular signaling. Although the role of estrogen in mammary gland development is well established, the impact of androgens remains less understood. To address this gap, we inhibited androgen signaling in utero using the anti-androgen flutamide (FLU) and investigated the effects on mammary gland development in rats. Using an integrative strategy combining histology, transcriptomics, lipidomics, cytokine profiling, and high-resolution imaging, mammary tissue were analyzed at pre-puberty (postnatal days [PND] 21), peri-puberty (PND46), and adulthood (PND90). FLU exposure induced subtle, yet significant, alterations in mammary morphology and molecular signatures. At PND21, the FLU-exposed group exhibited an increased number of adipocytes with reduced size. Transcriptomic analysis revealed differentially expressed genes at PND21 and enrichment in pathways related to androgen response and immune signaling, but minimal changes at later developmental stages. Lipidomic profiling showed transient disruption in long-chain fatty acid composition at early developmental stages. Cytokine profiling revealed a reduced adaptive immune response at PND46 and PND90, and second-harmonic generation imaging demonstrated changes in collagen fiber orientation and density across all developmental stages. These data indicate that prenatal androgen signaling is essential for proper stromal development and the establishment of early transcriptional networks in the mammary gland, with only minor long-term effects on glandular architecture in adult nulliparous females.
Gestational and lactational exposure to phthalates and nanoplastics induces inflammation and oxidative imbalance in the prostate of adult rats.
Rocha VA, Cavalheiro M, Souza PV … +10 more , Magosso N, Fioretto MN, Moreira MF, Pinha VC, Aquino AM, Quadreli DH, Fernandes GSA, Justulin LA, Flaws JA, Scarano WR
Toxicol Sci · 2026 May · PMID 42155030 · Publisher ↗
The intrauterine microenvironment is highly susceptible to environmental disturbances, which may result in persistent effects on offspring health. Given that plastic-derived contaminants can cross the placental barrier a...
The intrauterine microenvironment is highly susceptible to environmental disturbances, which may result in persistent effects on offspring health. Given that plastic-derived contaminants can cross the placental barrier and act as endocrine-disrupting chemicals, this study evaluated the effects of perinatal exposure to an environmentally relevant phthalate mixture (PM) and nanoplastics (NPs) on the prostate of adult rats. Pregnant Sprague-Dawley rats were allocated into 6 experimental groups: Ctrl (vehicle), T1 (20 μg/kg/d PM), T2 (200 mg/kg/d PM), T3 (NPs: 1.0 mg/kg/d), T4 (20 μg PM + NPs), and T5 (200 mg PM + NPs). Treatments were administered orally from gestational day 10 to postnatal day (PND) 21. Male offspring were euthanized on PND120, and ventral prostate samples were collected. Histological evaluation revealed increased inflammatory foci and stromal expansion in all exposed groups (T1-T5), along with reduced luminal compartment in T1, T3, and T4 compared with controls. All treated groups showed increased total and degranulated mast cells and enhanced androgen receptor immunoreactivity. Tumor necrosis factor (Tnf) expression was increased in all exposure groups, whereas Rela gene expression was elevated in the T4 and T5 groups. Oxidative stress analysis demonstrated increased lipid peroxidation and glutathione S-transferase (GST) activity in all treated animals. Catalase (CAT) activity was reduced in T1, T3, and T5, whereas superoxide dismutase (SOD) activity increased in T3, T4, and T5. Elevated levels of reduced, oxidized, and total glutathione (GSH, GSSG, tGSH) were observed in T5. Overall, perinatal exposure to PM and NPs induced persistent inflammation and altered redox status in the prostate, increasing susceptibility to pathological disorders.
Benzene-induced pulmonary dysfunction in diabetic mice.
Hussain A, Srivastava S, Zelko IN
Toxicol Sci · 2026 May · PMID 42152223 · Full text
Benzene is among the most produced chemicals in the United States, and epidemiological evidence links benzene exposure to impaired pulmonary function; yet, the influence of pre-existing metabolic disease on susceptibilit...
Benzene is among the most produced chemicals in the United States, and epidemiological evidence links benzene exposure to impaired pulmonary function; yet, the influence of pre-existing metabolic disease on susceptibility remains poorly defined. We investigated the effects of inhaled benzene on pulmonary mechanics and surfactant homeostasis in diabetic mice with varying glycemic severity. Mildly hyperglycemic KK and severely hyperglycemic, insulin-resistant KK-Ay mice were exposed to filtered air or 50 ppm benzene (6 h/d, 5 d/wk) for 2 wk. Pulmonary function was assessed by the forced-oscillation technique, and bronchoalveolar lavage fluid was analyzed for surfactant proteins and lipid composition by lipidomics; whole-lung transcriptomes were profiled by RNA sequencing. Benzene increased respiratory system resistance and Newtonian resistance in KK-Ay mice without altering tissue elastance or compliance in either strain. Methacholine challenge revealed exaggerated bronchoconstriction in benzene-exposed KK-Ay mice, with significant increases in Rn and tissue damping absent in benzene-exposed KK controls. Surfactant protein B was reduced by benzene in both strains, whereas SP-C was diminished in benzene-exposed KK-Ay mice compared with benzene-exposed KK controls. Lipidomic profiling demonstrated benzene-induced surfactant lipid remodeling characterized by elevated triradylglycerols, increased phosphatidylcholine-to-phosphatidylethanolamine ratio, and reduced phosphatidylserine, with specific lipid classes correlating with airway resistance. Transcriptomic analysis identified enrichment of neurotransmitter transport, ion-channel, and calcium-regulated exocytosis pathways, alongside upregulation of phase II detoxification enzymes. These findings demonstrate that in severe hyperglycemic KK-Ay mice, benzene induced airway hyperresponsiveness through convergent disruption of surfactant protein/lipid homeostasis, identifying poorly controlled diabetes as a risk factor for benzene-related respiratory toxicity.
Integrating experimental data and mechanistic modeling to assess potential lead exposure from tampon use.
Ellison CA, Doyle PR, Haven CA … +3 more , McClenathan DM, Obringer CM, Woeller KE
Toxicol Sci · 2026 May · PMID 42097300 · Full text
Trace levels of lead (Pb) have been reported in tampons, prompting concerns about potential exposure during menstrual product use. However, the presence of a chemical in a product does not necessarily translate to biolog...
Trace levels of lead (Pb) have been reported in tampons, prompting concerns about potential exposure during menstrual product use. However, the presence of a chemical in a product does not necessarily translate to biologically relevant exposure. In this study, we characterized the distribution and binding of Pb in menstrual fluid and developed a deterministic, compartmental mass-balance model to evaluate the release and fate of Pb that may be present as an inadvertent trace impurity in tampons. Our experimental measurements demonstrated that Pb preferentially partitions to the red blood cell (RBC) fraction of menstrual fluid. The remaining Pb was distributed in the plasma fraction, where the majority was protein-bound. These distribution and binding characteristics were broadly comparable to those reported for systemic blood. Utilizing these data, a mechanistic model was parameterized to describe the release of Pb from a tampon into menstrual fluid, partitioning between menstrual fluid compartments, reabsorption into the tampon during fluid uptake, and potential permeation across vaginal tissue. Under the conservative assumptions evaluated here, model simulations for a 4-h tampon wear scenario predicted that the majority of theoretically released Pb is reabsorbed into the tampon, with only a very small fraction (<1 ng; <0.3%) available for potential absorption into vaginal tissue. Sensitivity and alternative release scenario analyses demonstrated that predicted tissue uptake remained minimal across various plausible conditions. Collectively, these findings underscore the importance of integrating chemical presence data with physiological context and mechanistic modeling to inform exposure assessment and support science-based evaluation of product safety.
Contemporary Review: Temporal Dynamics of Neuroinflammation in Response to Environmental Exposure.
Baird B, Noor S, Leng S … +2 more , Ottens AK, Campen MJ
Toxicol Sci · 2026 Jun · PMID 42097296 · Full text
Neuroinflammation is the brain's immune response to injury or disease and can negatively impact neurological functions, ranging from mood, cognition, learning, and memory, as well as promote neurodegenerative disease and...
Neuroinflammation is the brain's immune response to injury or disease and can negatively impact neurological functions, ranging from mood, cognition, learning, and memory, as well as promote neurodegenerative disease and even psychiatric conditions. Different environmental factors may trigger a neuroinflammatory response; the mechanisms of this activation may influence the pattern of activation, the immune subsets involved, and the persistence of the response. The duration of neuroinflammation may significantly outlast exposures, creating a potential unique vulnerability for rare or intermittent exposures. The neuroinflammatory response can be classified into acute, subacute, and chronic phases, with subacute neuroinflammation representing a transient state between acute and chronic inflammation. This phase, lasting from days to weeks, is denoted by metabolic disruptions, cognitive impairments, and peripheral immune activation. Environmental exposures, such as air pollution, pesticides, and heavy metals, and social factors impact oxidative stress, glial activation, and blood-brain barrier disruption, leading to neuronal injury and cognitive decline. Notably, exposures like diesel fumes and wildfire smoke have been shown to induce neuroinflammation, subsequently impacting memory and learning, and exacerbating mental health conditions such as post-traumatic stress disorder, depression, and anxiety. Triggering of reactive astrogliosis via impacts on the blood-brain barrier function may be the most common non-specific manner that environmental toxicants drive neuroinflammation, but direct outcomes from compounds that readily access the brain are also possible.
RENAsym, a quantitative systems toxicology platform model, designed to predict drug-induced kidney injury indicates polymyxin B and valproate overdose toxicity potential while simultaneously demonstrating acetaminophen and valproate safety at therapeutic levels.
Vallejo C, Bhargava P, Hamzavi N … +10 more , McDaniel M, Lakhani VV, Qi T, Battista C, Clemens L, Shoda LKM, Kenz ZR, Tallapaka S, Gebremichael Y, Woodhead JL
Toxicol Sci · 2026 Jun · PMID 42093084 · Full text
RENAsym is a quantitative systems toxicology (QST) modeling platform designed to predict drug-induced kidney toxicity. The model includes the proximal tubule cell (PTC) life cycle, regeneration of PTCs, sodium reabsorpti...
RENAsym is a quantitative systems toxicology (QST) modeling platform designed to predict drug-induced kidney toxicity. The model includes the proximal tubule cell (PTC) life cycle, regeneration of PTCs, sodium reabsorption, nephron death, and drug-induced injury and inflammation within the proximal tubules. The mechanisms of drug-induced toxicity included are overproduction of reactive oxygen species and mitochondrial dysfunction. Inputs into the QST model include the drug's exposure in the kidney and results from in vitro toxicity assays used to parameterize the toxicity mechanisms included in the model. The QST model has a wide array of outputs including fraction of viable PTCs, fraction of viable nephrons, and serum creatinine concentration over time that allow the user to assess the extent of kidney injury induced by a drug. Polymyxin B, valproate, and acetaminophen were three compounds used to test RENAsym's ability to correctly classify compounds as either nephrotoxic or safe. The model successfully identified polymyxin B and valproate as drugs with the potential to cause kidney injury and acetaminophen as a drug that does not cause kidney injury at therapeutic levels, providing increased confidence that the model can be used to indicate kidney injury liability using in vitro inputs before dosing in clinical settings.
RNAseq analysis demonstrates polarization state-dependent responses to ambient particulate matter in a particle composition-specific manner.
Smyth TR, Brocke S, Wu W … +2 more , An Z, Jaspers I
Toxicol Sci · 2026 May · PMID 42063226 · Full text
Macrophages are key innate immune cells which exist on a spectrum between pro-inflammatory (M1) and pro-resolutory (M2) states while retaining the ability to reprogram following exposure to new stimuli. Particulate matte...
Macrophages are key innate immune cells which exist on a spectrum between pro-inflammatory (M1) and pro-resolutory (M2) states while retaining the ability to reprogram following exposure to new stimuli. Particulate matter (PM) exposure significantly alters macrophage function, leading to increasing respiratory infection morbidity and mortality. However, the influence of macrophage polarization on PM responses remains poorly understood. We hypothesized that human macrophages would demonstrate polarization state- and seasonality-specific responses to airborne PM collected from Xinxiang, China. CD14+CD16- monocytes were differentiated into macrophages and polarized using established methods with or without PM co-exposure followed by RNA sequencing (RNAseq). Macrophage expression profiles were primarily determined by inflammatory (M1) versus naive (M0) and alternative activation (M2), regardless of PM exposure. However, differential expression analysis using polarization state-specific reference groups uncovered distinct gene and pathway expression patterns. Directly polarized M0- > M1 macrophages exhibited the fewest unique differentially expressed genes (DEGs) but displayed similar pathway level activity to reprogrammed M2- > M1 cells. M2 macrophages showed the highest number of unique DEGs suggesting increased chemotactic pathway activity. Conversely, M0 cells exhibited greater expression of major inflammatory cytokines and chemokines. Although polarization state was the primary driver of gene and pathway responses, expression levels varied depending on PM collection dates and correlated strongly with individual PM components in M0 and M2 cells. These findings highlight the need to consider both particle seasonality and macrophage polarization state when studying PM's impact on macrophages as these factors may contribute to the negative health outcomes associated with PM exposure during respiratory infections.
High-concentration MEHP triggers mtDNA depletion in undifferentiated HepaRG and C2C12 cultures and disrupts mitochondrial homeostasis in both HepaRG culture states.
Rahman MM, Khadka P, Young CKJ … +4 more , Wang J, McCormick EM, Falk MJ, Young MJ
Toxicol Sci · 2026 May · PMID 42054602 · Full text
Phthalates are often called "everywhere chemicals" because they are widely used in consumer products and are detectable in the environment and humans. One of the most studied phthalates, di-2-ethylhexyl phthalate (DEHP),...
Phthalates are often called "everywhere chemicals" because they are widely used in consumer products and are detectable in the environment and humans. One of the most studied phthalates, di-2-ethylhexyl phthalate (DEHP), is metabolized to mono-(2-ethylhexyl) phthalate (MEHP), which is known to disrupt metabolic processes through peroxisome proliferator-activated receptor (PPAR) signaling. However, accumulating evidence suggests that lipophilic phthalates also affect mitochondria, key regulators of oxidative metabolism, autophagy, and apoptosis. Based on previous observations that undifferentiated cells are more sensitive to a mitotoxic agent, we hypothesized that MEHP differentially affects mitochondrial function and mitochondrial DNA (mtDNA) maintenance across hepatic cell states. To test this, we used the human HepaRG hepatoma-derived cell line, which can be cultured in undifferentiated and differentiated states, and assessed viability and mitochondrial function following prolonged 6- and 12-d high-concentration MEHP treatments. Prolonged treatments reduced viability and altered bioenergetics in both states. Short treatments (1 to 3 d) reduced viability only in differentiated cultures and were associated with mtDNA depletion in undifferentiated cultures. In both states, MEHP increased the expression of the low-molecular-weight mitochondrial genome maintenance exonuclease (MGME1) isoform, altered the levels of autophagy-related factors, and induced apoptosis. In another mitochondrial-competent myoblast model (C2C12 cells), a high concentration of MEHP was associated with mtDNA depletion, whereas lower concentrations were associated with modest reductions in cell density without detectable mtDNA loss. These results demonstrate state-dependent mitochondrial responses to MEHP and indicate that a reduced endpoint cell density is a sensitive outcome occurring independently of, and at lower concentrations than, mtDNA depletion in undifferentiated cells.
Omeprazole activation of CD4+ and CD8+ T-cells through off-target covalent modification of cellular proteins.
Grice S, Albashtawy S, Wells G … +14 more , Hering L, Adair K, Sarsby J, Brownridge P, Ford M, Lloyd R, Hampson L, Wagner A, Sun Y, Liu H, Hammond S, Meng X, Zhang F, Naisbitt D
Toxicol Sci · 2026 May · PMID 42045007 · Full text
Targeted covalent inhibition of protein function is increasingly used as a therapeutic mode of action; however, there is a need to characterize off-target binding interactions and to understand whether this represents an...
Targeted covalent inhibition of protein function is increasingly used as a therapeutic mode of action; however, there is a need to characterize off-target binding interactions and to understand whether this represents an immunological risk. Given that the proton-pump inhibitor omeprazole exerts its mechanism of action through covalent inhibition, it serves as an ideal model to investigate the relationship between off-target protein binding and T-cell activation. Binding of omeprazole, omeprazole metabolites, and alternative proton-pump inhibitors to antigen-presenting cells and GST-pi was characterized by mass spectrometry. Omeprazole-responsive clones were generated and assessed in terms of cytokine secretion, pathways of T-cell activation, and crossreactivity with omeprazole metabolites, alternative proton-pump inhibitors, and unrelated drugs. Omeprazole stimulated CD4+ and CD8+ T-cell clones to proliferate and secrete cytokines and cytolytic molecules. HLA-restricted T-cell activation was dependent on processing of omeprazole protein adducts by antigen-presenting cells. Omeprazole-modified CYS-containing peptides derived from 36 off-target proteins were detected within antigen-presenting cells. Omeprazole metabolites and alternative protein pump inhibitors that form protein adducts also activated omeprazole-responsive T-cells. In conclusion, T-cells were activated with omeprazole via a hapten mechanism and exhibited considerable promiscuity to metabolites and structurally related drugs of the same pharmacological class. Similar off-target binding interactions may be a relevant concern for the increasing number of covalent inhibitor drugs receiving regulatory approval.
Emergent latent neurotoxic effects of manganese following nominal chronic exposures in human stem cell and Caenorhabditis elegans models.
Tang X, Martins AC, Tukker AM … +3 more , Kim H, Aschner M, Bowman AB
Toxicol Sci · 2026 Apr · PMID 42033779 · Full text
Behavioral deficits can emerge after the removal of manganese (Mn) exposures in humans and other mammals. Although epidemiological studies provide substantial evidence supporting latency, challenges reproducing such effe...
Behavioral deficits can emerge after the removal of manganese (Mn) exposures in humans and other mammals. Although epidemiological studies provide substantial evidence supporting latency, challenges reproducing such effects in alternative models have slowed mechanistic understanding. Here, we report in 2 systems, human-induced pluripotent stem cell (hiPSC)-derived and Caenorhabditis elegans, that prior chronic exposure elicits clear latent neurotoxic effects in gene expression and functional outcomes. To identify these effects and investigate underlying mechanisms, single-cell RNA sequencing was employed in hiPSC-derived cortical culture to provide comparisons of transcriptomic changes immediately following versus after cessation of chronic Mn exposures. Transcriptomic alterations revealed latent effects after cessation of elevated Mn that were not detected immediately following 40-day exposures. To confirm the reproducibility of the observed latent magnification of chronic Mn-induced neurotoxicity, behavioral endpoints were evaluated in C. elegans. We detected a significant amplification of 2 motor phenotypes after a period of exposure cessation. These data demonstrate, in 2 genetic and mechanistically tractable systems, the detection of novel latent neurotoxic effects not detected until the cessation of a chronic exposure at a magnitude well beyond the effects of the chronic Mn exposure itself. Identified alterations support a linkage between the latent effects following chronic Mn exposure and a broad range of neurodegenerative etiologies and provide insight into the cellular pathways involved. Using both in vitro and in vivo experimental models provides complementary evidence that substantially strengthens the robustness and translational relevance of these novel findings.
An environmentally relevant mixture of organophosphate esters induces cholesterol biosynthesis in THP-1 macrophages.
Giles BH, Robaire B, Mann KK
Toxicol Sci · 2026 May · PMID 42015365 · Full text
Organophosphate esters (OPEs), widely used as flame retardants and plasticizers, are environmental toxicants known to disrupt lipid metabolism. Although most studies have focused on individual OPEs, environmental exposur...
Organophosphate esters (OPEs), widely used as flame retardants and plasticizers, are environmental toxicants known to disrupt lipid metabolism. Although most studies have focused on individual OPEs, environmental exposures typically involve complex mixtures. Our previous studies demonstrated that a representative OPE mixture from Canadian household dust promotes cholesterol and lipid droplet accumulation in THP-1 macrophages. However, the molecular mechanisms underlying this lipid dysregulation remain unclear. Here, we employed tandem mass tag (TMT)-based quantitative proteomics to investigate how OPE mixtures alter protein expression and lipid regulation in macrophages. THP-1 macrophages were exposed to vehicle or environmentally relevant dilutions of the OPE mixture for 48 h. Lysates were subjected to TMT labeling and mass spectrometry. Bioinformatic analyses using STRING and Ingenuity Pathway Analysis identified 162 differentially expressed proteins, with unsupervised clustering highlighting cholesterol biosynthesis as a key pathway. Further validation via qPCR and upstream analysis implicated the sterol regulatory element-binding protein 2 (SREBP2) signaling axis in OPE-induced cholesterol biosynthesis. Functional assays revealed that atorvastatin-mediated HMG-CoA reductase inhibition, the rate limiting enzyme in cholesterol biosynthesis, prevents cholesterol buildup and lipid droplet formation in macrophages. These findings provide the first evidence that an environmentally relevant OPE mixture can induce cholesterol biosynthesis in human macrophages. These studies provide mechanistic evidence that an environmentally relevant mixture of organophosphate esters induces cholesterol biosynthesis in macrophages. These findings link real-world exposure to lipid pathways implicated in metabolic disease and support the need for updated regulatory standards that protect human health.
Integration of mechanistic and repeat dose toxicity data in the derivation of an oral reference dose for HFPO-DA.
Thompson CM, Heintz MM, Rogers SI … +2 more , Vincent MJ, Haws LC
Toxicol Sci · 2026 May · PMID 41968070 · Full text
HFPO-DA (ammonium 2,3,3,3-tetrafluoro-2-(heptafluoropropoxy)-propanoate) is a short-chain perfluoroether carboxylic acid used as a polymerization aid in the manufacture of some types of fluorinated polymers. Existing tox...
HFPO-DA (ammonium 2,3,3,3-tetrafluoro-2-(heptafluoropropoxy)-propanoate) is a short-chain perfluoroether carboxylic acid used as a polymerization aid in the manufacture of some types of fluorinated polymers. Existing toxicity criteria for HFPO-DA include chronic and subchronic oral reference dose (RfD) values based on liver effects in mice. New mechanistic data demonstrate that these liver effects are incontrovertibly linked to rodent-specific peroxisome proliferator-activated receptor alpha (PPARα) signaling pathways that lack human relevance. Therefore, it was critical to reevaluate existing HFPO-DA RfD values. A literature review was conducted to identify human and animal studies that might serve as the basis of updated toxicity criteria. Relevant studies were considered along with a newly completed chronic bioassay in mice. No epidemiological studies were determined acceptable for use in toxicity value development. The only neoplasms in the chronic mouse bioassay were in the liver and were considered PPARα related. The most sensitive extrahepatic noncancer endpoints in rodents involved placental lesions in reproductive and developmental toxicity studies and reduced testicular cellularity in mice following chronic exposure. Both effects resulted in RfD values of 0.001 mg/kg-d. Also presented are probabilistic risk assessment methods that resulted in a similar probabilistic RfD. These results indicate the need to update existing toxicity criteria for HFPO-DA.
Unveiling toxicological adverse outcomes: toward construction and simulation of large-scale networks.
Ikonomi N, Ketter N, Pepe MAA
Toxicol Sci · 2026 Apr · PMID 41967855 · Full text
Predicting toxicological adverse outcomes is crucial for advancing in silico toxicology strategies. Modern toxicology increasingly relies on systems biology approaches to model and interpret these outcomes. Adverse outco...
Predicting toxicological adverse outcomes is crucial for advancing in silico toxicology strategies. Modern toxicology increasingly relies on systems biology approaches to model and interpret these outcomes. Adverse outcome pathways (AOPs) focus on systems-level descriptions and causal linear relations among initiating, key, and adverse outcome events. Key characteristics (KC)-based topologies capture mechanistic breadth via interconnected property-based modules without assuming linear causality. From another perspective, emerging physiological maps dive deeper into toxicological mechanisms by mapping them at the detailed molecular level. To capture the dynamic nature of toxicological responses, especially their time- and dose-dependent behaviors, there is growing interest in integrating systems biology and mathematical modeling strategies. Although dynamic models have been applied to small-scale AOPs, larger regulatory networks remain largely unexplored from a dynamic perspective. In this review, we highlight recent efforts to combine systems and network biology approaches for predicting toxicological adverse outcomes, covering network construction, analysis, and dynamic predictions. We also explore the aspect of dynamically simulating large-scale molecular networks and its potential contribution to systems toxicology. Specifically, we charter the use of logic-based models (Boolean networks) as an integrative approach to understand molecular crosstalk and cellular phenotypes, highlighting the potential repurpose of existing models. To this end, we show 2 use cases on toxicological applications of Boolean network models. Finally, we prospectively discuss the importance and need of bridging molecular and systemic scales and integrating these modeling strategies with high-dimensional data sources, including omics and multi-omics datasets.
Withdrawal interval estimations of oxytetracycline and chlortetracycline in swine using physiologically based pharmacokinetic models: a global trade perspective.
Mi K, Enomoto H, Baynes RE … +1 more , Lin Z
Toxicol Sci · 2026 Apr · PMID 41965107 · Publisher ↗
Violative drug residues in animal-derived food are a global food safety concern. Physiologically based pharmacokinetic (PBPK) modeling is a valuable tool for predicting drug residues in edible tissues and determining wit...
Violative drug residues in animal-derived food are a global food safety concern. Physiologically based pharmacokinetic (PBPK) modeling is a valuable tool for predicting drug residues in edible tissues and determining withdrawal intervals (WDIs). This study aimed to develop a PBPK model for oxytetracycline (OTC) and chlortetracycline (CTC) in swine to determine WDIs based on different regulatory requirements of different countries. The models were calibrated and evaluated with the pharmacokinetic data after oral administration via feed and drinking water collected from the Food Animal Residue Avoidance Databank (FARAD). The models can accurately capture the observed kinetics in plasma and edible tissues (liver, muscle, kidney, and fat), and most of the model predictions were within a 3-fold factor of observed data (87.9% for OTC and 88.9% for CTC). WDIs of OTC and CTC were determined using the population PBPK models based on maximum residue limits (MRLs) from 13 countries or regions under the label dosage regimens. The models were converted to a web-based PBPK dashboard. The models are a useful tool for predicting tissue residues and estimating WDIs based on different MRLs across countries, thereby supporting food safety assessment and international trade of meat products derived from swine treated with OTC and CTC.
Repeated bronchoalveolar lavage variability and impact in a 2-week nonclinical toxicology study.
Hackshaw E, Nelson J, Boyle MH
Toxicol Sci · 2026 Apr · PMID 41965104 · Publisher ↗
Nonclinical inhalation studies often rely on systemic biomarkers of exposure and pharmacology for serial evaluations, since local pulmonary effects are generally limited to assessment by terminal procedures. In addition,...
Nonclinical inhalation studies often rely on systemic biomarkers of exposure and pharmacology for serial evaluations, since local pulmonary effects are generally limited to assessment by terminal procedures. In addition, invasive procedures raise concern with health impact for subsequent inhalation doses and artifacts in tissue histology that could confound interpretation of toxicity. To test a non-terminal lung monitoring technique for use in non-human primate repeat-dose inhalation toxicology studies, bronchoalveolar lavage was performed during the pretest phase (twice), following 7 d of treatment with saline or albuterol, and the day after the final dose. Evaluations included clinical observations and histopathology, as well as differential cell counts and quantification of protein, lactate dehydrogenase, and urea in bronchoalveolar lavage fluid. Animals sufficiently recovered from the procedure were dosed by conscious facemask inhalation several hours later, without incident, and had slightly lower lavage total cell counts. Bronchoalveolar lavage the day before necropsy was associated with mixed cell infiltrates or inflammation in 2 of 8 animals. Total white cell counts in lavage fluid were higher but more variable when collected as a terminal rather than survival procedure, although intra-animal variability between pretest occasions was notable as well. Urea concentration showed greater variability than total protein and lactate dehydrogenase, although values were near the method's limit of detection. In summary, this study showed that bronchoalveolar lavage can be performed as a serial evaluation in nonclinical studies, without benefit from multiple baseline occasions and with some effect on lung histopathology.
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