Toxicology and Applied Pharmacology

Background The placenta serves a critical role in nutrient uptake and waste elimination for the developing fetus. The placenta is also responsible for the uptake and/or exchange of xenobiotics, including medications, between the maternal and fetal bloodstreams. An estimated 40-80% of women take medications or drugs during pregnancy for a variety of conditions. Very little is understood about fetal drug and nutrient exposure during pregnancy and how it may change over the course of fetal development. Objective This study aimed to characterize the abundance of transport proteins in placental tissue, which are important in modulating fetal nutrient and drug exposure, over the duration of pregnancy. Mass spectrometry-based global proteomic analysis revealed trends in the expression of thousands of proteins throughout gestation. Focusing on the membrane-associated proteome enabled an increased emphasis on the solute carrier and ATP-binding cassette families of transporter proteins that are critical for nutrient and xenobiotic transport across the maternal-fetal barrier. Study Design Using data-independent acquisition proteomics, relative abundance of proteins in placental tissue samples was profiled across all three trimesters of pregnancy (Trimester 1 = 16, Trimester 2 = 9, and Term = 9). Membrane fractions were generated to enrich membrane-associated proteins for proteomic analysis. Placental samples were grouped into randomized batches for membrane fraction generation and mass spectrometry analysis. Proteomic search results from each batch were imported into the R programming environment from Skyline, concatenated, and normalized as one data set for downstream analysis. Results A total of 6,331 proteins were detected across all samples with 4,210 proteins identified in every sample. Pathway analysis revealed that as gestational age increases, membrane-associated proteins involved in more complex metabolic pathways increase in relative abundance while those involved in extracellular remodeling events and simple organic ion transport tended to decrease. A total of 139 solute carrier and ATP-binding cassette transport proteins were identified in all samples, and 80 were identified in every sample. In general, membrane-associated proteins, including solute carrier and ATP-binding cassette transport proteins, were significantly enriched in placental tissue collected during early gestation compared to term placental tissue. Conclusion This study presents a comprehensive profiling of membrane-associated proteomic changes during gestation and identifies significant gestational age associated abundance changes at the protein level in several transport protein families. The application of data-independent acquisition global proteomic techniques enabled in-depth analysis of thousands of proteomic changes across pregnancy in a single experiment. These data provide critical information to support future studies into the understanding of fetal exposure to xenobiotics and nutrients circulating in the maternal bloodstream.

Tenuazonic acid (TeA) is an emerging Alternaria mycotoxin frequently detected in food and feed commodities, raising concerns about its toxicological relevance. Chronic oral exposure to TeA has been reported to induce dysplastic alterations in the esophageal mucosa of mice, while human biomonitoring data indicate an association between TeA exposure and esophageal cancer, although a causal relationship has not yet been established. At a mechanistic level, the effects of TeA in esophageal cells remain poorly characterized. Therefore, this study investigated the impact of TeA on cytotoxicity, oxidative stress, DNA damage, mitochondrial homeostasis, cell-cycle distribution and transcriptomic stress responses in human esophageal KYSE-510 cells. TeA induced a concentration-dependent reduction in metabolic activity and total protein content after 24 h exposure to 0.1-100 M. Significant cytotoxicity was measured starting from 20 M. At sub-cytotoxic concentrations, TeA triggered rapid ROS formation within 5-30 min exposure and induced formamidopyrimidine-DNA glycosylase (FPG) sensitive DNA damage after 1 h exposure (5-7.5 M), indicating oxidative DNA lesions. In addition, TeA altered mitochondrial morphology after 4 h exposure at 7.5 M, manifested by shrinkage of the mitochondrial network area and perinuclear redistribution, while mitochondrial respiration showed only a non-significant tendency towards reduced respiratory capacity. RNA sequencing after 6 h exposure to 10 M TeA revealed oxidative stress-associated transcriptional changes, impaired antioxidant and stress-adaptive responses, and p53-associated stress signaling. Furthermore, TeA induced significant G2/M phase accumulation after 24 h exposure to 1-10 M.

BACKGROUND: Perfluorooctane sulfonate (PFOS) is a widely distributed persistent organic pollutant in the environment and has been associated with an increased risk of atherosclerosis. However, the underlying pathogenic mechanisms remain largely unclear. This study aimed to investigate the effects of PFOS on atherosclerosis and its associated gut-vascular axis. METHODS: Pseudo-germ-free mouse models and fecal microbiota transplantation (FMT) were used to determine the role of the gut microbiota in PFOS-induced atherosclerosis. Metagenomic sequencing was performed to characterize alterations in gut microbial composition following PFOS exposure, and targeted metabolomics was used to assess bile acid profiles in the ileum and plasma. Transcriptomic analysis of Bacteroides caecimuris (B.caecimuris) was conducted to explore the reasons for the increased abundance of B.caecimuris after PFOS exposure. In addition, intestinal transcriptomics and ChIP-qPCR were performed to validate transcriptional regulation within the FXR-TLR3 signaling axis. RESULTS: Among 127 participants with paired serum and fecal samples, including 82 patients undergoing coronary angiography with Gensini scores (GS score), fecal PFOS levels were significantly associated with lipid profiles and GS score, whereas serum PFOS showed no such association. Mechanistically, PFOS exposure promotes intestinal enrichment of B. caecimuris by upregulating its tolC gene, thereby enhancing efflux capacity. This microbial shift was accompanied by reduced levels of tauro-ursodeoxycholic acid (TUDCA) and aberrant activation of intestinal FXR signaling. Further analyses demonstrated that FXR activation upregulated TLR3 expression and promoted inflammatory responses and atherosclerosis progression via the TLR3-NF-{kappa}B signaling axis. Both intestinal epithelial-specific FXR deficiency (Fxr{Delta}IE) and TUDCA supplementation significantly suppressed pathway activation and alleviated disease phenotypes.Functional experiments identified TLR3 as a key downstream effector of FXR. Overexpression of TLR3 abolished the protective effects observed in Fxr{Delta}IE mice. Moreover, pharmacological inhibition of TLR3 using CU CPT-4a significantly improved established atherosclerotic lesions in vivo. CONCLUSIONS: This study identifies a gut microbiota-driven FXR-TLR3 signaling axis that mediates PFOS-induced atherosclerosis. These findings provide new mechanistic insights into environmentally induced cardiovascular disease and suggest potential targets for risk assessment and therapeutic intervention.

An acetaminophen (APAP) overdose is the leading cause of drug-induced hepatotoxicity and acute liver failure (ALF) in the United States. While N-acetylcysteine (NAC), is highly effective when administered early after an overdose, its efficacy decreases with delayed administration. Since most patients present late to the clinic, there is an urgent need for novel late-acting therapeutic options to prevent progression to ALF. We previously demonstrated the benefit of delayed activation of the Adenosine A2B Receptor (A2BAR) in attenuating APAP-induced hepatotoxicity and this study focuses on its effects on liver recovery after injury. Fasted male C57BL/6J mice were treated with 300 mg/kg APAP, followed by activation of A2BAR 6 or 9 h later and sacrifice 24, 48 or 72 h post-APAP with evaluation of liver injury, the innate immune response and liver regeneration. Delayed activation of A2BAR significantly enhanced liver recovery, with accelerated repopulation of the liver by Kupffer cells, increased macrophage migration to the necrotic areas and their faster resolution. A2BAR activation also upregulated lipid metabolism related genes in non-parenchymal cells and cell proliferation and metabolism genes in hepatocytes. Remarkably, genes such as Cidec and Plin2, crucial for lipid droplet formation, were upregulated, indicating that A2ABR activation enhances lipid metabolism which plays a key role in providing energy for liver regeneration. Overall, these findings highlight the potential of A2BAR activation not only in protecting against liver injury, but also in promoting and accelerating liver regeneration by modulating the innate immune responses and metabolic pathways.

ObjectiveThe aryl hydrocarbon receptor (AhR) pathway primarily mediates pollutant responses by activating xenobiotic metabolism enzymes like cytochrome P450 1A1 and 1A2 (CYP1A). Although AhR has also been implicated in systemic metabolic dysfunction and is inducible in pancreatic islets, its role in islet physiology remains unclear. MethodsWe analyzed a publicly available bulk human islet transcriptomic dataset to identify pathways associated with CYP1A1 expression. We also assessed islet responses to the pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and glucolipotoxicity (GLT) in vitro using two mouse models: a global Cyp1a1/1a2 double knockout (CypKO) model, which disrupts canonical AhR-CYP1A signaling in whole islets, and a {beta}-cell-specific Ahr knockout ({beta}AhrKO) model, which abolishes AhR signaling selectively in {beta}-cells. We then examined the role of {beta}-cell Ahr in early adaptation to high-fat diet (HFD) feeding in vivo. ResultsXenobiotic and nutrient metabolism pathways were enriched in donors with high CYP1A1 expression. Global Cyp1a1/1a2 deletion increased susceptibility of female mouse islets to TCDD-induced impairments in insulin secretion but had minimal effects on GLT responses in either sex. In contrast, {beta}-cell Ahr deletion did not affect islet responses to TCDD, but exacerbated GLT-induced islet dysfunction in male islets and increased baseline insulin secretion in both vehicle- and GLT-exposed female islets in vitro. Lastly, {beta}-cell Ahr deletion prevented adaptive HFD-induced hyperinsulinemia in both sexes in vivo. ConclusionIslet AhR signaling shapes responses to chemical and nutrient stressors in a context- and sex-dependent manner. While the canonical AhR-CYP1A axis supports female islet resilience to TCDD, {beta}-cell AhR signaling more broadly regulates nutrient stress responses in both sexes.

Per- and polyfluoroalkyl substances (PFAS) are persistent toxicants with immunological, metabolic and epithelial effects, but their relevance to inflammatory skin disease remains unclear. We developed a computational toxicology framework to test whether perfluoroalkyl sulfonate programs, especially perfluorooctanesulfonic acid (PFOS) and perfluorohexanesulfonic acid (PFHxS), converge with psoriasis-associated keratinocyte inflammation. Exposure transcriptomes were derived from GSE236956, in which human embryonic stem cell-derived epithelial-lineage models were exposed to 10 M PFAS for 8-16 days. Six PFAS were prioritized using descriptors, Tanimoto similarity, toxicology evidence, adverse outcome pathway (AOP)-like key events, exposure differentially expressed gene burden and read-across support. PFAS signatures were integrated with psoriasis bulk transcriptomes, single-cell RNA sequencing, keratinocyte-state mapping, regulator and communication inference, spatial transcriptomics and token-aware Geneformer-compatible virtual perturbation. PFOS ranked highest in integrated prioritization, followed by PFHxS and perfluorooctanoic acid. PFHxS produced a smaller but directionally informative signature within a PFOS-dominant perfluoroalkyl sulfonate footprint. The shared PFOS and PFHxS program converged with psoriasis through inflammatory keratinocyte, epidermal-stress, cytoskeletal and lipid-related modules. Single-cell and spatial analyses localized the program to activated keratinocytes and inflammatory epidermal niches, with strong spatial co-localization with inflammatory keratinocyte and epidermal stress scores. Virtual perturbation prioritized S100A9, S100A8, KRT16, IL36G, CCL20, CXCL8, FABP5, KRT17, FOS, JUN and NFKBIZ as candidate effectors. These findings support an exposure-informed, experimentally testable hypothesis linking persistent perfluoroalkyl sulfonate programs to keratinocyte inflammatory niches in psoriasis.

PurposeEstrogen deficiency drives bone loss through interacting endocrine, oxidative, inflammatory and bone-remodeling disturbances. Ergothioneine (EGT) is a diet-derived thiol/thione antioxidant whose effects on the estrogen-deficient skeleton are unknown. We evaluated whether EGT, alone or combined with vitamin K2, vitamin D3 and magnesium L-threonate, attenuates the skeletal and systemic consequences of ovariectomy (OVX) in mice. MethodsForty-eight female C57BL/6J mice underwent sham surgery or OVX and received daily oral gavage for 12 weeks of vehicle, alendronate (1.53 mg/kg), EGT (30 mg/kg/day), EGT with vitamin K2 (40 {micro}g/kg/day) and vitamin D3 (500 IU/kg/day), or EGT with vitamin K2, magnesium L-threonate (350 mg/kg/day) and vitamin D3 (n = 5-6 analysed per group). Outcomes included the uterine index, tibial micro-computed tomography, distal-femoral histology, and serum bone turnover markers (CTX-I, PINP, osteocalcin), sex hormones, TNF-, IL-6, SOD and MDA. OVX lowered the uterine index and induced tibial trabecular deterioration, with increased CTX-I, decreased PINP and osteocalcin, elevated TNF- and IL-6, reduced SOD and increased MDA (all P < 0.01 vs sham). Alendronate restored tibial micro-CT bone-volume fraction (BV/TV) and trabecular number (P < 0.01 vs OVX). The EGT-based regimens did not significantly restore tibial micro-CT BV/TV, trabecular thickness or trabecular number (all P > 0.05 vs OVX), but significantly increased trabecular area on distal-femoral histology (OVX 7.6% vs 14.2-15.0% across regimens; P < 0.05 vs OVX) and lowered CTX-I, TNF-, IL-6 and MDA while raising SOD and partially restoring PINP and osteocalcin (P < 0.05-0.01 vs OVX). Because the histological and micro-CT endpoints were assessed at different skeletal sites, structural interpretation is cautious. Apparent increases in serum estradiol were assay-dependent and are regarded as exploratory. Ergothioneine-based nutritional regimens improved the systemic oxidative, inflammatory and bone-turnover environment of estrogen-deficient bone loss and preserved distal-femoral trabecular area on histology, although tibial three-dimensional microarchitecture by micro-CT was not restored. Because the histological and micro-CT endpoints were assessed at different skeletal sites, the structural interpretation is necessarily cautious. These findings support further evaluation of EGT as a dietary adjunct, with mechanistic and dose-optimization studies warranted.

Background: Randomized controlled trials have shown that computed tomographic (CT) screening reduces lung cancer mortality. Improved identification of at-risk groups, by leveraging non-smoking risk factors, could help refine screening selection. Aim: To evaluate polygenic risk scores (PRSs) and ambient air pollution (AAP) exposure for risk stratification in the NELSON lung cancer screening cohort. Methods: Two PRSs (PRS-McKay/PRS-Byun) and several AAPs (including nitrogen dioxide, ozone, and particulate matter [PM]) were assessed in the NELSON lung cancer screening trial (N=7,364). PRSs were validated in the Rotterdam Study (N=11,493). Associations with lung cancer, mortality, screening results, and discriminative ability to distinguish lung cancer were evaluated. Results: PRS-McKay and PRS-Byun were associated with lung cancer (odds ratio [OR] per SD [95%CI]: 1.22 [1.08-1.37] and 1.28 [1.13-1.44], respectively) and lung cancer-specific mortality (OR [95%CI]: 1.24 [1.05-1.47], for both), but not with non-lung cancer mortality (OR [95%CI]: 1.01 [0.94-1.10] and 1.03 [0.95-1.12], respectively). Exposure to PM2.5 was associated with lung cancer (OR [95%CI]: 1.11 [1.01-1.22]). PM constituents were associated with adenocarcinoma, particularly PM10 (OR [95%CI]: 1.16 [1.01-1.32]) and ultra-fine particles (OR [95%CI]: 1.16 [1.04-1.30]). PRS and AAP added modestly to the discriminative ability for lung cancer on top of pack-years, age, and sex (area under the curve [95%CI]: 0.659 [0.624-0.695] vs. 0.643 [0.608-0.679]). Conclusions: PRSs and exposure to PM were associated with lung cancer in a high-risk screening population. The primary potential of PRSs may reside in refining lung cancer screening selection toward individuals at higher risk of dying from lung cancer specifically.

Alternaria mycotoxins represent an emerging concern due to their frequent occurrence in food and feed. However, available toxicological data remain limited. Under the current EU regulatory framework, and in line with the EFSA/ECHA/JRC guidance for the identification of endocrine disruptors (EDs), assessment of endocrine activity relies on standardized assays performed according to OECD Test Guidelines (TGs) for the estrogen-, androgen- and steroidogenesis- (EAS) modalities. Within the framework of the European Partnership for the Assessment of Risks from Chemicals (PARC), standardized in vitro methods of regulatory relevance were performed for six chemically characterized Alternaria toxins, aiming to address current regulatory gaps on EAS-mediated activity. Alternariol (AOH), alternariol monomethyl ether (AME), tenuazonic acid (TeA), altertoxin-I (ATX-I), tentoxin (TEN) and altenuene (ALT) were assessed over a broad concentration range, from 0.001 up to 60 M, depending on cytotoxicity and solubility profile of each compound. Our findings indicate estrogenic activity for AOH (PC50: 3.9 - 4.6 {micro}M) and AME (PC50: 5.2 - 8.5 {micro}M) in the estrogen receptor transactivation assay (OECD TG 455), as well as an anti-estrogenic activity for ATX-I (IC30: 0.27 - 0.37 {micro}M). Minimal positive responses were observed at high concentrations for AOH (from the dose of 3 {micro}M) and for AME (from the dose of 10 {micro}M) in the agonistic part of the androgen receptor transactivation assay (OECD TG 458), which may also reflect glucocorticoid receptor activation. No effects on estradiol or testosterone production were observed for any of the tested Alternaria compounds in the steroidogenesis assay (OECD TG 456).

Research on targeted genetic therapies for myocardial diseases, such as cardiomyopathies, currently focuses on (r)AAVs as the delivery method. Despite substantial efforts and advances in animal trials, predicting biodistribution and transduction efficacy in human tissue remains challenging due to interspecies differences in tissue tropism and the difficulty of accurately assessing alternative delivery routes and vector differences. The application in humans has also proven challenging, in part due to severe adverse events associated with systemic administration of (r)AAVs. This necessitates implementing alternative trial designs and stringent evaluation methods that minimize harm or risk to patients. Applying a predesigned vector carrying a gene-editing tool to a normothermic machine-perfused living beating heart in an ex vivo setting could overcome conventional obstacles and limitations. This can serve as a basis for safe and effective gene-therapy testing and assist in evaluating effects at the molecular level. Boxed-Breathing-Heart is a translational trial assessing the feasibility of ex vivo gene editing and gene translation in normothermic machine-perfused human hearts. Human hearts explanted from cardiomyopathy patients undergoing heart transplantation are donated for research and immediately placed in an Organ Care System, where they are surgically connected. The viability of the heart is maintained through normothermic perfusion of system solutions and donor blood. A predesigned AAV containing a CRISPR-Cas system is infused into the circulation and dispersed throughout the tissue via coronary perfusion. The changes at the cellular and molecular levels are assessed continuously via frequent sequential myocardial biopsies. Furthermore, after the pre-planned 72-hour perfusion, the heart is sectioned and analyzed using spatial and single-cell omics. The aim is to provide a proof-of-concept for genetic therapeutic options delivered to the human heart via AAV in an ex vivo perfusion setup. In summary, Boxed-Breathing-Heart provides an ex vivo translational platform for evaluating targeted cardiac gene therapies, enabling molecular analysis directly in human hearts and accelerating clinical translation without posing risks to patients.

BACKGROUND: Erythrophagocytosis constitutes a major pathogenic mechanism of metabolic dysfunction associated fatty liver disease (MAFLD). Our previous research established a quantitative thin-layer chromatography (TLC) technique for sphingomyelin, revealing reduced levels in the red blood cells (erythrocytes) of patients with metabolic dysfunction associated fatty liver disease (MAFLD). This reduction was accompanied by erythrocyte sphingosine accumulation, a driver of pro-inflammatory erythrophagocytosis, though sphingosine 1-phosphate release remained stable. To better understand erythrocyte sphingosine metabolism, we adapted our quantitative TLC method to analyze sphingosine within the erythrocyte-conditioned media (ECM) of MAFLD patients. Methodology Separation was performed on 10X10cm Silica gel 60 F254 plates using a mobile phase of chloroform, methanol, acetic acid, and water (60:50:1:4 v/v/v/v). The dynamic range, linearity, and range of linearity were assessed by analysing sphingosine levels from 0.1 to 10microg/spot. We validated the system precision and sensitivity by performing triplicate analyses of sphingosine standards (1.25, 2.5, and microg). The limits of detection and quantification were derived from the calibration curve slope and standard deviation (3.3 XSD/slope for LOD; 10 XSD/slope for LOQ). Accuracy was assessed via recovery tests at 100%, 200%, and 300% of a 2.5microg load. We confirmed specificity by evaluating the retention factors against other lipid species. This protocol was applied to Folch-extracted lipids from the ECM (5 X 107 cells/ml) of four MAFLD patients and four healthy controls, spiked with 5microg of sphingosine. Findings The calibration model, based on combined Green and Blue color intensities, followed the linear equation y = -11.171x + 353.25(R2 = 0.94). Interday precision values were 0.21%, 1.65%, and 0.44%, while recovery rates (accuracy) ranged from 94.5% to 98.7%. The measured LOD and LOQ were 0.75microg and 1.21microg, respectively. The sensitivity was calculated at 90ng. Statistical analysis showed no significant variance in sphingosine concentrations in erythrocyte-conditioned media between the MAFLD group and the control group. Summary The described thin layer chromatography is accurate, precise, sensitive, with good limits of detection and quantification, and most importantly is low-cost and time-efficient. Using this method, we show that while erythrocytes of MAFLD patients exhibit sphingosine accumulation, the utilisation of exogenous sphingosine from their erythrocytes is not affected. This suggests that the metabolic shift may be driven by increased sphingosine supply from the plasma.

Chemotherapy treatment of colorectal cancers (CRC) using cisplatin (CDDP) encounters problems of drug resistance by the cancer cells and cytotoxicity to normal cells, highlighting the urgent need for joint therapeutical strategies. Selenium-enriched rapeseed extracts exhibit anti-cancer effects but the bioactive components and mechanisms remain unclear. Here, we applied different solvents to fractionate the extracts from Selenium-enriched rapeseed and found that the water extract (WE) fraction significantly enhanced the cytotoxic effect of CDDP on cancer cells but no damage on normal cells. HPLC-ICP-MS analysis revealed that methylselenocysteine (MSC) and selenocystine (SeCys2) were the main selenium speciation in WE. Through cell biology and integrative multi-omics analysis, we found a synergistic anti-CRC cell effect when combining CDDP with MSC, sulforaphane (SFN), celastrol (Cel), Indole-3-carbinol (I3C), -linolenic acid (ALA) or linoleic acid (LA). We propose that the CDDP-WE combination treatment holds the promise for improving curative efficacy for chemo-refractory CRC patients in the future.

Background & Aims: Activation of apoptosis signal-regulating kinase 1 (ASK1), a ubiquitous redox-sensitive kinase, results in inflammation, apoptosis, and fibrosis, key common pathways in human liver disease. SRT-015 is a novel, small molecule inhibitor of ASK1. This study evaluated the in vitro efficacy of SRT-015, compared it to other ASK1 inhibitors, and determined the in vivo efficacy of SRT-015 across multiple acute and chronic liver disease models. Methods: In vitro studies determined the kinase potency and selectivity of SRT-015, and cellular studies were used to demonstrate direct mechanisms of action. The cardiac hERG channel inhibition was assessed and PK determined in rodents and nonhuman primates. In vivo studies evaluated SRT-015 efficacy in rodent models of drug-induced hepatotoxicity (acetaminophen (APAP) overdose), alcohol-associated liver disease (ALD), metabolic-disease associated steatohepatitis (MASH) and cholestatic disease (bile duct ligation, BDL). Results: SRT-015, was demonstrated a selective ASK1 kinase, and SRT-015 treatment directly inhibited fibrosis, apoptosis and inflammation in activated human fibroblasts, hepatocytes and PBMCs, respectively without safety signals or hERG inhibition. Other ASK1 inhibitors had safety concerns or limited functional activity. Liver and kidney selective PK were observed for SRT-015 in all species evaluated. In vivo, SRT-015 treatment was efficacious in the acute mouse APAP overdose and ALD model significantly (P<0.05) decreasing serum ALT. Using a therapeutic diet-induced obesity (DIO)-MASH model with biopsy-verified fibrosis, SRT-015 treatment significantly (P<0.05) inhibited DIO-induced liver enzymes, hepatomegaly, fibrosis, inflammation, and apoptosis independent of body weight loss whereas treatment with selonsertib was ineffective. In a rat cholestatic model, SRT-015 treatment significantly (P<0.05) decreased fibrosis and stellate cell activation. Conclusions: These findings support SRT-015 as a potential therapeutic for human liver diseases of any etiology.

Depleted uranium (DU) is an environmental contaminant with a 30 g/L (ppb; parts per billion) EPA maximum contaminant level (MCL) for drinking water. The mining of uranium and use of DU in modern weapons underly human exposure that disproportionally impacts military and tribal communities in the United States. Uranium's radiotoxic characteristics are understood, but its chemical hazards much less so. In zebrafish (Danio rerio) and human cell cultures we test the hypothesis that exposure to DU negatively impacts cellular function and development through disruption of mitochondrial metabolism. Using a novel shrapnel model with TEM/SEM+EDS, we showed uranium microparticles caused proximity-dependent mitochondrial disruption. In waterborne exposure paradigms, larval movement was reduced and hatching delayed as a result of reduced movement and not enzyme deficiencies in response to 18 ppb DU, below the MCL. Increased DNA damage accumulation was detected in exposed larva and cells. DNA-damage quantitative PCR of DU-exposed larvae showed increased damage in the ahr1 locus (nuclear gene) and decreased mitochondrial DNA (mtDNA) copy number, but mtDNA damage levels varied across experiments. Mitochondrial function was assessed using a resazurin-based assay in the presence and absence of antioxidants and showed diminished cytoplasmic reductive capacity. DU exposure alone did not enrich antioxidant gene expression, contrasting with arsenic exposure, a known ROS-inducer and Nrf2-activator. Sulforaphane (SFN), a potent Nrf2-activator, did not blunt the effects of DU exposure, despite activation of antioxidant response element (ARE) genes (gstp and gss), but did blunt the effects of arsenic exposure. The most enriched transcript in DU-exposed larvae coded for slingshot protein phosphatase (ssh), further exploration revealed ssh1b as the zebrafish-specific ortholog activated in response to DU, and inhibition using an identified SSH1 inhibitor, Sennoside A, partially rescued the metabolic and hatching defects observed. Our data points to a cytotoxic mechanism in which DU disrupts mitochondrial function through ssh1b enrichment that impairs normal mitophagy, leading to decreased cellular reductive potential independent of either ROS production or ARE-activation. Our results suggest that health impacts from DU exposure may be directly linked to impaired mitochondrial functions.

Background and aims: Longitudinal data and tracking of serum lipoprotein(a) (Lp(a)) concentrations through childhood?s development from infancy to adolescence are lacking. We aimed to establish the strength of the tracking phenomenon from early infancy to adolescence and to examine whether a heart-healthy dietary intervention and individual dietary components influence serum concentrations of Lp(a). Methods: 1062 healthy children aged 7 months were recruited and randomized into control (N=522) and intervention (N=540) groups in the Special Turku Coronary Risk Factor Intervention Project (STRIP). Serum Lp(a) concentration was measured at 10 age points (0.7, 1.3, 2, 3, 4, 5, 9, 11, 13, and 15 years) and median serum Lp(a) levels were studied longitudinally. Tracking across age points was studied using Spearman's rank-order correlation. Sex differences and the effects of the dietary intervention and individual dietary components were analyzed with linear mixed-effects models for repeated measures. Results: A total of 7018 Lp(a) measurements were analyzed. Median serum Lp(a) concentrations increased from infancy until approximately age 13 years. After age 13, median Lp(a) declined in boys (-15.1%) but was largely unchanged in girls. Girls had higher Lp(a) concentrations at all age points (P=0.01). Spearman's correlation analysis indicated a strong tracking between age points in both sexes (r=0.854-0.956). Achievement of at least one dietary fat quality goal of the intervention corresponded to a 2.5% increase in serum Lp(a) concentration (P=0.0004). Higher sucrose intake was associated with modestly higher Lp(a), whereas fiber intake showed no association. At age 15 years, 16.2% of all participants with available measurements had elevated Lp(a) ([&ge;] 30mg/dL). Conclusions: A rising trend was observed in median serum Lp(a) concentrations from infancy to adolescence. Due to strong tracking, these findings suggest that early-life measurements may provide valuable insight for longitudinal cardiovascular risk assessment. Heart-healthy diet does not meaningfully influence serum Lp(a).

Air pollution exposure has been associated with increased risk of developing mental health problems. It is possible that individuals at high genetic risk for psychopathology may be more vulnerable to these effects; however, this question remains to be investigated. We leveraged longitudinal data from n=10,620 participants from the Adolescent Brain Cognitive Development Study to first investigate sex-stratified associations of particulate matter (PM2.) exposure and genetic risk with mental health trajectories across 9-16 years including internalizing symptoms and psychotic like experiences (PLEs). Additionally, we tested whether genetic risk for schizophrenia (PRS-SCZ) and major depressive disorder (PRS-MDD) exacerbate the association with PM2. exposure and change in symptoms over time. PM2. exposure was associated with lower decreases in PLEs over time in females (p-FDR=0.005), with no effects on internalising symptom trajectories in either sex. Genetic influences were sex-specific, with higher PRS-SCZ and PRS-MDD linked to greater increases in internalising symptoms in females (p-FDR=0.009; p-FDR=0.022) and higher PRS-MDD associated with greater decreases in PLEs in males (p-FDR=0.001). In females we also observed an interaction between PM2. and PRS-MDD on PLEs trajectories (p-FDR=0.048) such that those with high genetic risk and high PM2.5 exposure demonstrated increases in PLEs over time. Our results suggest that PM2. exposure and polygenic risk for depression jointly shape mental health during adolescence. This underscores the potential of interventions aimed at lowering air pollution during sensitive periods of neurodevelopment in improving adolescent mental health.

IntroductionPrenatal dibutyl phthalate (DBP) exposure is associated with increased risks of adverse fetal outcomes as well as metabolic and cardiovascular diseases in the offspring in a fetal sex-specific manner. However, mechanisms underlying these prenatal DBP exposure-associated adverse fetal/offspring outcomes are unclear. We hypothesize that environmentally relevant low-dose prenatal DBP exposure dysregulates fetal-placental vascular function and lipid metabolism in a fetal sex-specific manner, thereby impairing placental efficiency and programming adverse offspring metabolic outcomes. MethodsFemale CD-1 adult mice (8-10wks) were orally dosed with vehicle or an environmentally relevant low-dose DBP (0.1 g/kg/day) daily from 30 days pre-pregnancy through gestational day (GD) 18.5. Fetal-placental vascular hemodynamics of these dams were examined using high-frequency ultrasound at multiple timepoints. The effect of prenatal environmentally relevant low-dose DBP exposure on placental efficiency, spatial transcriptomic profiles, lipid homeostasis, and placental vascular endothelial cells function in male and female fetuses were evaluated at gestational day (GD) 18.5. ResultsThe prenatal low-dose DBP exposure dysregulated the fetal-placental vascular hemodynamic indices from mid-to late gestation. DBP exposure impairs placental efficiency in male, but not female placenta at GD18.5. Further, female placentas exhibited fetal labyrinth vasculature-specific transcriptomic adaptations that preserves placental efficiency and endothelial function. In contrast, male placentas exhibited minimum transcriptomic adaptation, together with compromised placental efficiency and endothelial function associated with lipotoxic lipid profile. ConclusionsIn conclusion, prenatal low-dose DBP exposure dysregulates placental vascular function and lipid homeostasis in a fetal sex-specific manner, with male fetuses being more susceptible to DBP exposure.

The histone variant H2A.J is implicated in radiation-induced senescence by promoting the transcription of inflammatory genes. While H2A.J expression increases in renal tubular epithelial cells (TECs) following ionizing radiation (IR), its functional role remains poorly understood. To investigate this, constitutive H2A.J knock-out (KO) mice and wild-type (WT) controls were subjected to CT-guided IR (fractionated whole-body or localized kidney IR). Kidneys were analyzed at acute, intermediate, and chronic stages using immunofluorescence, histochemistry, automated image analysis, and electron microscopy. In WT TECs, IR induced rapid chromatin incorporation and C-terminal serine phosphorylation of H2A.J. Conversely, KO kidneys exhibited significantly more severe histopathological damage, including tubular dilation, flattened epithelium, associated with increased apoptosis, and premature senescence, characterized by persistent DNA damage with lamin B1 loss. Notably, KO TECs displayed disrupted mitochondrial networks and reduced brush borders even at baseline, which were further exacerbated by IR. Unlike WT controls, KO kidneys developed progressive tubular atrophy and incipient fibrosis, indicating a failure in regenerative capacity. Our findings suggest that H2A.J loss impairs tubular regeneration due to defective mitochondrial activation, resulting in insufficient energy supply for coordinated repair. Collectively, these results identify H2A.J as a critical stress-adaptive histone variant essential for the epigenetic regulation of tissue repair following radiation-induced damage. One Sentence SummaryIn irradiated kidney, the loss of histone variant H2A.J impairs the chromatin-mediated adaptation of mitochondrial function in tubular epithelial cells, thereby exacerbating cellular stress - characterized by increased induction of apoptosis and senescence - and ultimately leading to tubular atrophy. Translational RelevanceAcute and chronic kidney injury are frequent complications of genotoxic cancer therapies. Chemo- and radiotherapy induce DNA lesions that trigger cell death and senescence, often leading to irreversible renal damage. However, renal regeneration can occur through the dedifferentiation, proliferation, and redifferentiation of surviving tubular epithelial cells (TECs). This repair process is governed by epigenetic mechanisms that regulate the DNA damage response (DDR) and adapt gene expression programs. Following ionizing radiation (IR), epigenetic remodeling involves the incorporation of histone variants that modulate chromatin accessibility for stress-responsive transcription factors. We identify the histone variant H2A.J as a constitutive component of renal TECs, significantly upregulated after exposure to ionizing radiation (IR). Using H2A.J knock-out (KO) mice, we demonstrate that its absence disrupts acute damage responses and prevents coordinated repair, severely impairing regeneration. Mechanistically, H2A.J deficiency compromises mitochondrial function under postirradiation metabolic stress, driving the transition from acute injury to chronic kidney disease via persistent inflammation and maladaptive tubulointerstitial repair. Targeting these epigenetic drivers offers a promising strategy for regenerating damaged kidney tissue in oncology.

High concentrations of steroidal hormone compounds are a growing source of concern for environmental pollution in aquatic ecosystems. In this study, we examine the effects of two estrogenic compounds (estriol and 17-ethinylestradiol) on fitness traits in the aquatic microcrustacean, Daphnia magna, a key bioindicator species for toxicology studies. The impacts were compared of two forms representing a natural and synthetic estrogenic compound. Growth and reproduction traits were assayed by exposing Daphnia to each estrogen type at four concentrations reflecting potential environmental exposure conditions up to acute toxicity levels (ranging from 0.1 - 50 {micro}g/L). Assaying the effects at a variety of concentrations is important given that it is known that hormone exposures can often result in non-monotonic responses. Both forms of estrogen impact a subset of the traits assessed, in some cases leading to beneficial changes and others causing harm. Estriol, the naturally-occurring estrogen, and EE2, the synthetic version, at high doses shift fitness traits in opposite directions such as adult growth rate as do at low doses for fecundity. In conclusion, our results support the need to assay a wide array of traits using multiple forms of steroidal hormones at a range of doses in order to assess non-monotonic patterns and their impact on an organismal fitness. In particular, assays that extend beyond the conventional measurements of lethality during acute exposure windows will be essential for understanding the impact of increased levels of hormone pollution on aquatic organisms and ecosystem health.

Tenuazonic acid (TeA) is one of the most frequently detected Alternaria mycotoxins in contaminated food. Despite its frequent occurrence, its immunomodulatory effects remain insufficiently characterized. Therefore, the present study investigated the impact of TeA on inflammatory signaling and cytokine regulation in monocytes and intestinal epithelial cell (IEC) models. NF-{kappa}B activity was assessed using a reporter gene assay in THP1-Lucia monocytes, while cytokine mRNA expression and protein secretion were quantified in Caco-2 and HCEC-1CT cells by qRT-PCR and ELISA, respectively. In THP-1 monocytes, TeA significantly suppressed lipopolysaccharide (LPS)-induced NF-{kappa}B activation in a concentration-dependent manner starting at 25 M, while cytotoxicity occurred only at concentrations [&ge;]100 M. In HCEC-1CT and differentiated Caco-2 cells, TeA increased IL-6, IL-8, and TNF- mRNA levels at non-cytotoxic concentrations ([&ge;]10 M). In Caco-2 cells, these transcriptional changes were accompanied by increased cytokine secretion, whereas HCEC-1CT cells showed only partial effects on the protein level after short-term exposure. Following prolonged incubation, TNF- secretion was increased and IL-6 and IL-8 secretion were slightly reduced. IL-10 remained unaffected under all conditions. Overall, TeA exerted cell type-dependent immunomodulatory effects characterized by immunoinhibitory activity in monocytes and pro-inflammatory responses in IECs, highlighting the complex immunotoxic potential of this Alternaria mycotoxin.