搜索结果：Toxicological sciences : an official journal of the Society of Toxicology

BACKGROUND: Detailed, comprehensive, and timely reporting on population health by underlying causes of disability and premature death is crucial to understanding and responding to complex patterns of disease and injury burden over time and across age groups, sexes, and locations. The availability of disease burden estimates can promote evidence-based interventions that enable public health researchers, policy makers, and other professionals to implement strategies that can mitigate diseases. It can also facilitate more rigorous monitoring of progress towards national and international health targets, such as the Sustainable Development Goals. For three decades, the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) has filled that need. A global network of collaborators contributed to the production of GBD 2021 by providing, reviewing, and analysing all available data. GBD estimates are updated routinely with additional data and refined analytical methods. GBD 2021 presents, for the first time, estimates of health loss due to the COVID-19 pandemic. METHODS: The GBD 2021 disease and injury burden analysis estimated years lived with disability (YLDs), years of life lost (YLLs), disability-adjusted life-years (DALYs), and healthy life expectancy (HALE) for 371 diseases and injuries using 100 983 data sources. Data were extracted from vital registration systems, verbal autopsies, censuses, household surveys, disease-specific registries, health service contact data, and other sources. YLDs were calculated by multiplying cause-age-sex-location-year-specific prevalence of sequelae by their respective disability weights, for each disease and injury. YLLs were calculated by multiplying cause-age-sex-location-year-specific deaths by the standard life expectancy at the age that death occurred. DALYs were calculated by summing YLDs and YLLs. HALE estimates were produced using YLDs per capita and age-specific mortality rates by location, age, sex, year, and cause. 95% uncertainty intervals (UIs) were generated for all final estimates as the 2·5th and 97·5th percentiles values of 500 draws. Uncertainty was propagated at each step of the estimation process. Counts and age-standardised rates were calculated globally, for seven super-regions, 21 regions, 204 countries and territories (including 21 countries with subnational locations), and 811 subnational locations, from 1990 to 2021. Here we report data for 2010 to 2021 to highlight trends in disease burden over the past decade and through the first 2 years of the COVID-19 pandemic. FINDINGS: Global DALYs increased from 2·63 billion (95% UI 2·44-2·85) in 2010 to 2·88 billion (2·64-3·15) in 2021 for all causes combined. Much of this increase in the number of DALYs was due to population growth and ageing, as indicated by a decrease in global age-standardised all-cause DALY rates of 14·2% (95% UI 10·7-17·3) between 2010 and 2019. Notably, however, this decrease in rates reversed during the first 2 years of the COVID-19 pandemic, with increases in global age-standardised all-cause DALY rates since 2019 of 4·1% (1·8-6·3) in 2020 and 7·2% (4·7-10·0) in 2021. In 2021, COVID-19 was the leading cause of DALYs globally (212·0 million [198·0-234·5] DALYs), followed by ischaemic heart disease (188·3 million [176·7-198·3]), neonatal disorders (186·3 million [162·3-214·9]), and stroke (160·4 million [148·0-171·7]). However, notable health gains were seen among other leading communicable, maternal, neonatal, and nutritional (CMNN) diseases. Globally between 2010 and 2021, the age-standardised DALY rates for HIV/AIDS decreased by 47·8% (43·3-51·7) and for diarrhoeal diseases decreased by 47·0% (39·9-52·9). Non-communicable diseases contributed 1·73 billion (95% UI 1·54-1·94) DALYs in 2021, with a decrease in age-standardised DALY rates since 2010 of 6·4% (95% UI 3·5-9·5). Between 2010 and 2021, among the 25 leading Level 3 causes, age-standardised DALY rates increased most substantially for anxiety disorders (16·7% [14·0-19·8]), depressive disorders (16·4% [11·9-21·3]), and diabetes (14·0% [10·0-17·4]). Age-standardised DALY rates due to injuries decreased globally by 24·0% (20·7-27·2) between 2010 and 2021, although improvements were not uniform across locations, ages, and sexes. Globally, HALE at birth improved slightly, from 61·3 years (58·6-63·6) in 2010 to 62·2 years (59·4-64·7) in 2021. However, despite this overall increase, HALE decreased by 2·2% (1·6-2·9) between 2019 and 2021. INTERPRETATION: Putting the COVID-19 pandemic in the context of a mutually exclusive and collectively exhaustive list of causes of health loss is crucial to understanding its impact and ensuring that health funding and policy address needs at both local and global levels through cost-effective and evidence-based interventions. A global epidemiological transition remains underway. Our findings suggest that prioritising non-communicable disease prevention and treatment policies, as well as strengthening health systems, continues to be crucially important. The progress on reducing the burden of CMNN diseases must not stall; although global trends are improving, the burden of CMNN diseases remains unacceptably high. Evidence-based interventions will help save the lives of young children and mothers and improve the overall health and economic conditions of societies across the world. Governments and multilateral organisations should prioritise pandemic preparedness planning alongside efforts to reduce the burden of diseases and injuries that will strain resources in the coming decades. FUNDING: Bill & Melinda Gates Foundation.

Multi-omic investigations into environmental effects on health and disease are aided by inclusion of microbial microbiomes with assessment of mirobes producing metabolites that differentially modulate host organ functions. The gut microbiome is key because many environmental toxicants enter the body orally and may disrupt gut microbes that help digest food, as well as the microbiome-gut-brain axis, which produces regulatory metabolites with systemic effects. Environmental stressors may differentially alter brain development and function, even among identical twins, in that over time, there may be divergence due to epigenetic effects from the environment, including microbes within the microbiome. The diversity of microbiomes is presented as playing a key role in the influence of organs on each other, health, and the development of disorders. The gut microbes and their metabolites may cause mitochondria to produce less ATP and more reactive oxygen species (ROS). The metabolites produced by microbes during the digestion of foods can nourish or harm a person's cellular and molecular functions and vary depending on each person's exposome. The detrimental effects of environmental stressors are discussed, focusing on how altered levels of neuropeptides, neurotransmitters, and the inflammatory/anti-inflammatory balance affect health and disorders. During ATP production, dysfunctional mitochondria may produce more ROS, which can lead to inflammation and oxidative stress, causing cell damage and disrupting products needed for neuronal development, connections, and functions. The balance between inflammatory/anti-inflammatory biomarkers and metabolites and between oxidants/antioxidants is discussed in relation to some clinical connections; for example, the proportions of CD4 and CD8 T cells in HIV patients and the ROS-to-glutathione ratio in inflammatory bowel disease and septic patients. These imbalances are reviewed regarding brain development and functions leading to anxiety, depression, and dementia. The integration of multi-omics, dysbiosis, and mitochondrial dysfunction with a person's clinical evaluation is discussed to inform the formulation of prevention measures and therapeutic interventions regarding environmental effects on the microbiome-gut-brain axis and physical and mental health.

Firefighters are exposed to high levels of toxic chemicals while fighting fires, and previous studies have established these men and women have a significantly elevated risk for various cancers. Improved risk management for firefighters requires the identification of biomarkers indicative of physiological response. Micro-RNAs (miRNAs) have emerged as promising noninvasive prognostic and diagnostic biomarkers for various diseases. Here, we isolated miRNA from the urine of a large dataset of firefighters, collected pre- and post-fire exposure, as well as from healthy non-firefighter controls. miRNA was analyzed by microarray using the Affymetrix GeneChip miRNA 4.0 array. Analysis revealed 23 human miRNAs were significantly up-regulated and 25 significantly down-regulated in firefighters compared with control samples (analysis 1). Gene targets of these miRNAs were analyzed using the Online Database for Annotation, Visualization, and Integrated Discovery (DAVID) and found to cluster in several pathways and disease associations with smoking, cancer, and inflammatory diseases. Furthermore, we performed a longitudinal analysis of samples from firefighters that provided a sample prior to a fire exposure and immediately after a fire (analysis 2). This analysis found 20 miRNAs that were significantly up-regulated post-fire exposure. Of these, 5 were also up-regulated in firefighters vs control samples (hsa-miR-1268b, hsa-miR-4433b-3p, hsa-miR-4253, hsa-miR-6824-5p, and hsa-miR-3188). Again, analysis of gene targets of these miRNAs found association of mostly the same pathways and disease processes found in analysis 1. These findings are consistent with epidemiological evidence for increased risks associated with firefighting and offer a proof of concept and framework for the use of miRNA in urine as biomarkers for health risk assessment associated with firefighting.

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.

A chronic bioassay investigating radiofrequency (RF) carcinogenicity, intentionally designed to be conducted simultaneously in Korea and Japan, using the same research protocol and experimental environment. The study aimed to assess the potential carcinogenicity of Code Division Multiple Access (CDMA)-modulated 900 MHz RF signals at a whole-body specific absorption rate (SAR) of 4 W/kg, which is the reference level of the international human safety guideline, and to verify the key findings from the National Toxicology Program (NTP) study at that SAR level. Two reverberation chamber systems were used for RF exposures, and the same study protocols were followed. Male Harlan Sprague-Dawley (Hsd:Sprague Dawley SD) rats were randomly assigned to cage-control, sham-exposed, or RF-exposed groups. The exposure started on gestational day 5 and lasted for 18 h and 20 min each day, with 10-min on/off cycles. The project included a 28-d toxicity study, a 2-yr carcinogenicity study, and a 14-wk genotoxicity test. Histopathological evaluations were conducted in a partially blinded manner. The results were independently analyzed and submitted separately based on each country's research findings. In the Korean study, no statistically significant changes in tumor incidence or survival rates were observed. No significant RF-related effects were detected in the heart, brain, or adrenal glands. No changes in body temperature. Genotoxicity tests showed no evidence of DNA damage or mutation. In conclusion, the Korean part found that long-term exposure to CDMA-modulated 900 MHz RF was neither carcinogenic nor genotoxic at a SAR of 4 W/kg in male rats. An international animal study was jointly conducted as a chronic bioassay in Japan and Korea to evaluate the carcinogenicity of mobile phone RF signals and to verify key findings from the NTP study using identical protocols and exposure systems.

Exposure to environmental pollutants during key stages of development increases the risk of disease later in life. One such toxicant with growing evidence of this response is the air pollutant, ozone (O3). Exposure to O3 during the implantation receptivity period in rats affects the metabolic status of offspring at adolescence, which may increase their susceptibility to subsequent environmental exposures. Herein, we studied the impacts of maternal O3 exposure on postnatal systemic responses to O3 in male and female offspring. Following peri-implantation O3 exposure (0.8 ppm for 4 h/d on gestation days 5 and 6), offspring were exposed to O3 for 1 d/wk on postnatal weeks 5 to 7. After the final exposure, metabolic effects were analyzed by circulating hormones and clinical chemistries, as well as hepatic lipid status and transcriptomic alterations. By and large, male offspring from O3-exposed dams were more greatly impacted than those from air-exposed dams. This included increased hepatic lipid mobilization, increased circulating glucose, and a robust number of differentially expressed genes (2,348). Interestingly, many of these transcriptomic differences were attributed to maternal O3 exposure, with 1,741 of these genes sharing directional similarity with postnatally exposed air littermates. Females, on the other hand, reported minimal baseline effects of maternal O3 exposure (108). However, postnatal O3 exposure in female offspring substantially increased these differences to 947 genes. Collectively, this work supports the growing evidence that early pregnancy exposure to O3 alters the metabolic development of the offspring. Furthermore, postnatal exposure to environmental stressors reveals hepatic susceptibilities that are sexually dimorphic.

Per- and polyfluoroalkyl substances (PFAS) are a diverse class of highly fluorinated persistent synthetic chemical pollutants. Major routes of human exposure include ingestion of contaminated drinking water and foods including dairy. Consumption of PFAS-contaminated milk and dairy is especially concerning for infants and children who are particularly sensitive and most highly exposed. Here, we report findings of quantitative analysis of PFAS binding to β-lactoglobulin (β-Lg), the major whey protein in bovine milk, using differential scanning fluorimetry to determine binding affinities for 17 PFAS; except for uncharged fluorotelomer alcohols, β-Lg bound each PFAS congener tested, supporting a key role of charged functional groups in binding. The perfluoroalkyl carboxylic acid trifluoroacetic acid (TFA) bound with lowest affinity (Kd=8.6 mM) and long-chain congeners PFNA, PFDA, and PFUnDA bound with highest affinities. Evidence of significant cooperative binding was found for TFA, PFDA, PFUnDA, and PFOS. Molecular docking was used to define molecular mechanisms of PFAS binding by β-Lg and across the calycin super family of lipocalins and fatty acid-binding proteins. All calycins were predicted to bind PFAS in the calyx domain with ΔG of binding ranging from -5.3 to -9.4 kcal/mol, revealing that the binding affinity for many PFAS is greater than those for binding albumin. In total, this study has identified the calycin protein superfamily as PFAS-binding proteins, most of which have well-characterized functions related to key endocrine and toxicological pathways associated with the adverse consequences of PFAS exposure.

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.

Glyphosate is an herbicide found worldwide in glyphosate-based formulations (GBFs). Although glyphosate appears to have a low toxicity profile for humans and mammals, conflicting reports exist regarding the risk for cancer in humans. US-EPA and European regulatory agencies have described glyphosate as unlikely to pose a carcinogenic hazard to humans. However, the International Agency for Research on Cancer (IARC) classified glyphosate as "probably carcinogenic to humans (Group 2A)," citing "mechanistic data provide strong evidence for genotoxicity and oxidative stress." Given these discrepancies, the Division of Translational Toxicology at NIEHS designed an experimental strategy to expand mechanistic evidence and address critical gaps within existing literature (e.g. mechanistic evaluations of glyphosate alongside GBFs, inclusion of context-defining positive controls). Cell morphology, viability, H2O2, and γH2AX formation were assayed in human keratinocytes (HaCaT), previously cited by IARC, and human hepatocytes (HepaRG) to derive benchmark concentrations and fold-change response metrics. Our findings revealed glyphosate alone was weakly and inconsistently bioactive for oxidative stress and DNA damage when compared with positive controls. In contrast, most of the 13 GBFs evaluated were more clearly bioactive with no apparent correlation to varied glyphosate concentrations. Hierarchical clustering of biological responses revealed some bioactive GBFs to cluster near well-characterized positive controls for oxidative stress, whereas 4 GBFs clustered more similarly to negative controls and glyphosate. Collectively, this study provides a robust dataset with context-defining results that advance our understanding of the hazard potential of GBFs while revealing that glyphosate is likely not a primary driver of oxidative stress from GBF exposures.

Doxorubicin (Dox) is a potent chemotherapeutic with known vascular toxicity and connective-tissue damage. Endothelial cells (EC) and fibroblasts crosstalk is essential for vascular homeostasis and extracellular matrix (ECM) remodeling. This study aimed to explore whether Dox induces endothelial-to-mesenchymal transition (EndMT) and the paracrine effects of Dox-exposed EC on fibroblasts activation, senescence, and ECM synthesis. Human umbilical vein endothelial cells (HUVECs) were treated with Dox, and conditioned medium (CM) from EC was applied to human dermal fibroblasts for short- and long-term culture. Dox induced EndMT in ECs. Fibroblasts exposed to CM from Dox-treated EC exhibited early activation with increased fibroblast activation protein (FAP) and α-smooth muscle actin (α-SMA) at day 3, followed by a progressive senescent phenotype marked by elevated p21 and reduced Lamin B1 at day 21. ECM formation was impaired, with reduced collagen and increased transcriptional expression of matrix-degrading enzymes (MMP1 and MMP9). Cytokines profiling of the CM revealed decreased interleukin-1β (IL-1β), C-C motif ligand 2 (CCL2), and C-X-C motif ligand 10 (CXCL10), and elevated interleukin-6 (IL-6) levels. These findings demonstrate that exposure of EC to Dox induced endothelial dysfunction and elicited pathological paracrine signaling, driving fibroblast activation, myofibroblast transition, senescence, and ECM disruption. This mechanism may underlie Dox-related skin aging and delayed wound healing, and emphasizes the importance of endothelial dysfunction in chemotherapy-associated connective tissue damage and impaired repair.

BSTP0204A is a T-cell dependent-bispecific (TCB) antibody targeting 6-transmembrane epithelial antigen of the prostate 1 (STEAP1) that induces T-cell mediated killing of STEAP1 expressing cancer cells. STEAP1 is considered an attractive target for prostate cancer due to its high expression in the prostate and prostate cancer. Characterization of BSTP0204A showed potent T-cell-mediated killing in vitro and anti-tumor activity in mouse xenograft models against prostate cancer cell lines with both moderate and high STEAP1 expression. Analysis of STEAP1 protein expression in human and monkey tissues confirmed low STEAP1 expression outside of the prostate, suggesting a low potential for on-target/off-tumor toxicity. However, administration of BSTP0204A in a repeat-dose toxicity study in cynomolgus monkeys revealed adverse vascular inflammation that was inconsistent with STEAP1 expression observed in normal tissues. Additional assessments of STEAP1 expression in the vascular lesions from the toxicity study in monkeys and in human inflammatory disease conditions showed increased STEAP1 expression associated with inflammation and/or injury in both species. Furthermore, upregulation of STEAP1 was observed in both human and monkey primary cells in the presence of inflammatory stimuli. These findings suggest that systemic inflammation induced by T-cell activation following BSTP0204A treatment may have resulted in increased STEAP1 expression, inducing additional inflammation and tissue damage. This work demonstrated the need to understand not only baseline target expression for T-cell-engaging therapies, but also expression under conditions such as inflammation, injury, or disease.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is one of the most prevalent liver disorders, affecting approximately one-third of the global adult population. The disease begins with hepatic fat accumulation (steatosis) and can progress to inflammation, fibrosis, and hepatocellular carcinoma. To elucidate the complex mechanisms underlying MASLD, we have developed a novel mathematical model that integrates glucose and lipid metabolisms, oxidative stress, insulin signaling and insulin resistance, and cytokines functions. We demonstrated that variations in extracellular fatty acid and lactate levels, along with changes in the activities of key glycolytic and triglyceride-synthesizing enzymes observed in actual patients, exert a substantial impact on oxidative stress and subsequent cellular damage. Moreover, this model enabled us to evaluate daily metabolic dynamics associated with protein expression patterns specific to steatotic livers. Importantly, it also allowed simulation of cytokine release from hepatocytes into the bloodstream (autocrine and endocrine effects) and the impact of locally elevated cytokines concentrations derived from immune cells (paracrine effects). Our model revealed the dynamics of the early stages of MASLD progression in response to alterations in blood metabolite levels, hepatic enzyme activities, insulin profiles, and cytokine patterns. Furthermore, we identified specific combinations of these factors that may mitigate hepatic fat accumulation or oxidative stress, highlighting the importance of patient-specificity. This study presents the first mechanistic framework grounded in experimental data to describe the crosstalk among hepatic metabolism, insulin, and cytokines, serving as a powerful tool for elucidating disease mechanisms and developing therapeutic strategies.

While acetylcholinesterase (AChE) is the primary inhibitory target for organophosphate (OP) insecticides and chemical warfare nerve agents, research supports the concept that the wide range of systems affected by OPs are not solely dependent on the anticholinesterase activity of OPs but are attributable to the inhibition of other serine hydrolases. Oxime reactivators play an integral role in the treatment of acute OP exposure by returning function to OP-inhibited AChE. Our laboratory has synthesized and patented a platform of oxime reactivators (US Patent 9,227,937) that have provided central neuroprotection through returning function to OP-inhibited AChE in the brain and preservation of neuronal and glial structures from damage in a rat model; the current U.S. approved oxime, pralidoxime (2-PAM), does not provide central neuroprotection. Thus, returning function to other OP-inhibited serine hydrolases by these novel oximes could provide additional secondary neuroprotection. Rat brain proteins were studied using activity-based protein profiling (ABPP) for serine hydrolase targets of a highly relevant sarin surrogate, nitrophenyl isopropyl methylphosphonate (NIMP), alone or in combination with Oxime 20, our lead novel AChE reactivator. ABPP indicated that NIMP significantly inhibited 6 serine hydrolases and of these were the well-known serine hydrolases fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), which are important in endocannabinoid signaling. Oxime 20 administration after NIMP treatment showed only limited remediation of the inhibition experienced by these targets. The impact of this study is the identification of secondary serine esterase targets that could be explored for esterase reactivating therapeutics for the treatment of OP poisoning.

The potential carcinogenic and genotoxic effects of radiofrequency electromagnetic fields, particularly those emitted by mobile communication systems, have raised public health concerns. A previous study by the U.S. National Toxicology Program suggested increased incidences of gliomas and cardiac schwannomas in rats exposed to high levels of RF radiation. To evaluate these findings, an international collaborative study was initiated between Japan and Korea. Male Hsd:Sprague Dawley SD rats were exposed to 900 MHz CDMA-modulated RF-EMFs at a whole-body specific absorption rate of 4 W/kg for 18 h and 20 min daily over 2 yr. The study included a 28-d preliminary toxicity study, genotoxicity assays (alkaline comet and micronucleus tests), and a 2-yr carcinogenicity assessment. All procedures followed OECD guidelines and Good Laboratory Practice. No statistically significant increases in the incidences of neoplastic or non-neoplastic lesions were found in any major organ, including the brain, heart, and adrenal glands. Genotoxicity assays revealed no evidence of DNA damage or chromosomal aberrations in RF-exposed rats. A higher survival rate in the RF-exposed group, likely due to lower body weight and food consumption, was observed. This study, performed in Japan, jointly planned and executed by Japan and Korea, provides strong evidence that long-term exposure to 900 MHz RF-EMFs did not produce reproducible carcinogenic or genotoxic effects in male rats. Combined with data from the Korean counterpart study, these results are expected to contribute to future international assessments of the carcinogenic potential of electromagnetic radiation.

Why and how does cancer start? Building from a Symposium at the 2025 Society of Toxicology meeting, we convened a group of international experts to answer this seemingly simple question. As experimental evidence has evolved, perspectives on cancers' origins have shifted from the accumulation of DNA mutations in single cells to complex processes involving signals from an altered tissue microenvironment which promote tumorigenesis. Carcinogen exposures impact the biology of the microenvironment in complex and tissue-specific ways. These changes can include the infiltration of inflammatory cells that produce growth factors, neo-angiogenesis, morphological changes, and immune tolerance that avoids immune-mediated elimination. In this in-depth review, we discuss the evidence linking chemical-driven microenvironmental changes in the development of a range of solid and liquid tumors. We discuss specific phenotypic alterations, such as selection pressure driving clonal expansion and cellular plasticity and reacquisition of stem cell states, linked to carcinogen-induced changes in the microenvironment. We describe assays and biomarkers which can allow us to experimentally assess links between chemical exposures, the microenvironment, and cancer phenotypes. We end by discussing how understanding the role of the microenvironment and malignancy in toxicology is essential for accurate cancer hazard evaluation, development of next-generation risk assessment frameworks, identifying new strategies for cancer prevention, and improving patient care.

Prolonged exposure to ozone causes lung injury and persistent inflammation, pathologies associated with emphysema and asthma. Herein, we characterized inflammatory cells in the lungs using a murine model of prolonged ozone exposure, with the long-term goal of assessing their role in disease pathogenesis. Mice were exposed to air or ozone (1.5 ppm, 2 h, 2×/wk, 6 wk). Bronchoalveolar lavage fluid (BAL) and cells and lung tissue were collected 24 h after the final exposure. Alveolar/bronchiolar hyperplasia, epithelial degeneration, and mononuclear cell infiltration were observed following ozone exposure; BAL protein, cells, fibrinogen, and SP-A and SP-D were also increased, along with markers of oxidative stress, and impaired pulmonary function. Flow cytometric analysis of infiltrating myeloid cells revealed that after ozone exposure, the majority of these cells were mature infiltrating macrophages. These were comprised mainly of anti-inflammatory/profibrotic macrophages, with a smaller number of proinflammatory macrophages. Proinflammatory genes (Il1β, Ccl3, Ccl17, Ccl22, Tnfα) and NF-κB activity were increased in BAL cells from ozone-exposed mice (>97% macrophages); profibrotic genes (Mmp12, Mmp28, Tgfβ), but not anti-inflammatory genes (Il10, Arg1), were also upregulated. Following ozone exposure, glycolytic activity and oxidative phosphorylation increased in BAL cells, consistent with proinflammatory and profibrotic activation, respectively. These findings are important as they provide a rationale for evaluating the role of inflammatory macrophages in the pathophysiological response to prolonged ozone exposure. Impact statement: These studies are significant as they may lead to the identification of novel therapeutic approaches for reducing inflammatory lung disease caused by long-term exposure to inhaled ozone.

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 characterised 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 conclude, 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.

Clonal expansion (CE) of cells carrying cancer driver mutations (CDMs) is being developed as a biomarker of cancer risk. CE in lung of MutaMouse males treated with 0, 6.25, 12.5, and 25 mg/kg/d benzo[b]fluoranthene (B[b]F) by gavage for 90 and 180 d was assessed by CarcSeq. DNA regions encompassing mouse correlates of human hotspot CDMs were PCR amplified, attaching 18-base unique molecular identifiers (UMIs) during the PCR. Following library preparation and sequencing, UMI-defined read families were assembled to produce single-strand consensus sequences (SSCSs). Recovered mutants with mutant fractions (MFs) ≥10-4 were stratified based on their occurrence in lung-specific or nonlung driver sequences and CE was assessed on a per mouse basis as median absolute deviation in mutant fraction (MAD). A significant, dose-dependent increase in MAD was observed for lung-specific MFs after 180 d of B[b]F treatment, a duration that did not cause a significant increase in lung lesions. Dose- and treatment duration-related increases in MF were observed for Egfr, the mouse correlate of a known human lung tumor driver gene. MF and mutation counts were significantly decreased in response to longer treatment duration for some nonlung drivers, suggesting negative selection. Importantly, the normalized trinucleotide mutation spectrum derived from CDMs reflects amplification of preexisting spontaneous mutations, distinct from those induced by B[b]F mutagenesis. These results show CarcSeq detects CE of preexisting cancer driver gene mutants induced by the genotoxic carcinogen B[b]F and suggest a CE endpoint may be useful for evaluating cancer risk associated with tumor promoters or complete carcinogens.

Wildfire smoke (WFS) exposures are becoming more common, and firefighters and community members are often exposed to WFS for days to weeks. Controlled studies assessing the effects of repeated exposure to WFS and woodsmoke (WS; as a surrogate for WFS) or compared acute versus repeated exposure have evaluated a limited number of timepoints and endpoints using rodent models. Here, we leveraged differentiated primary human bronchial epithelial cells (HBECs) to test the hypothesis that different molecular responses occur upon acute versus repeat exposures to WS. HBECs (n = 4 donors) were exposed to 22 µg/cm2 red oak WS condensate for an acute, 4-h exposure, or repeat exposures of 4 h/d, 3 d/wk, across 2 wk. Membrane permeability, cell viability, and transcriptional responses were measured at the end of each exposure paradigm, and secreted proteins were measured throughout the repeat exposure. Acute exposure significantly increased expression of genes involved in fibrosis and immune response, whereas repeated exposure significantly decreased expression of genes involved in tissue repair and remodeling. Secreted protein responses were similar to transcriptomic responses and demonstrated temporal variation in response to exposure. This study supports the feasibility of using HBECs to evaluate acute and repeat WS exposures and indicates differential responses from these exposures with direct relevance to pulmonary disease processes, including those involved in fibrosis, asthma, and chronic obstructive pulmonary disease. These findings highlight the need for future studies to better understand molecular responses to repeated smoke exposures.