Ping Xiao

Hepatocellular carcinoma (HCC) is characterized by a poor prognosis, largely driven by metastasis and therapy resistance, processes closely linked to epithelial–mesenchymal transition (EMT) and ferroptosis. Neodymium nitrate (Nd(NO₃)₃), a predominant neodymium species, has shown antitumor potential, yet its specific effects and underlying mechanisms in HCC remain poorly understood. This study investigated whether Nd(NO₃)₃ exerts anti-malignant effects in Hep-G2 cells through the Nrf2–ferroptosis axis. We treated Hep-G2 cells with Nd(NO₃)₃ and assessed proliferation (CCK-8), migration, invasion (wound-healing and Transwell assays), EMT markers (E-cadherin, Vimentin), Nrf2 and GPx4 expression (Western blotting, qRT-PCR), and ferroptosis indicators (ROS, MDA, GSH, Fe²⁺/Fe³⁺). Low-dose Nd(NO₃)₃ significantly inhibited migration, invasion, and EMT-associated marker expression without affecting proliferation and promoted ferroptotic phenotypes, as evidenced by elevated ROS, MDA, and ferrous iron expression; decreased GSH levels; and downregulated GPx4 expression. Mechanistically, Nd(NO₃)₃ downregulated Nrf2 expression. Nrf2 overexpression reversed the suppression of migration, invasion, EMT-associated markers, and ferroptotic phenotypes, while the ferroptosis inhibitor deferoxamine rescued migration and invasion. These findings suggest that Nd(NO₃)₃ exerts anti-malignant effects in Hep-G2 cells by targeting Nrf2 to coordinately suppress EMT and promote ferroptosis, identifying the Nrf2–ferroptosis axis as a potential therapeutic target in HCC cells. However, these findings are based on in vitro experiments in a single Hep-G2 cell line with n = 3 biological replicates, which limits statistical power and generalizability; the results should therefore be interpreted as preliminary evidence requiring further validation in additional cell lines and in vivo models.

Background/Objective: Bisphenol AF (BPAF), a fluorinated analogue of bisphenol A, is an environmental contaminant associated with hepatotoxicity and metabolic disruption. However, the systematic molecular mechanisms linking early transcriptional events to metabolic dysfunction in the liver remain poorly defined. The aim of this study is to elucidate the association between BPAF exposure and hepatic lipid accumulation by integrating transcriptomics, cellular metabolomics, and targeted phenotypic assays. Methods: We performed RNA-sequencing on livers from mice exposed to BPAF (0.1–10 mg/kg/day, 28 days), and performed non-targeted metabolomics on AML12 murine hepatocytes co-cultured with RAW264.7 macrophages in a Transwell system (0–2500 nM BPAF, 48 h). Key metabolic pathways were identified through integrated bioinformatics and validated using enzymatic assays, qRT-PCR, Western blotting, and phenotypic staining (lipid droplets, ROS). Results: Multi-omics integration revealed significant disruption of PPAR signaling and the tricarboxylic acid (TCA) cycle. A striking dose-dependent accumulation of succinate was observed in exposed cells, concomitant with a significant inhibition of succinate dehydrogenase (SDH) activity (52% reduction at 2500 nM, p < 0.001). Transcriptomic data confirmed the downregulation of mitochondrial fatty acid β-oxidation genes. Phenotypic validation indicated that BPAF exposure is associated with oxidative stress, pro-inflammatory cytokine release (TNF-α, IL-6), and pronounced intracellular lipid droplet accumulation in hepatocytes. Conclusions: This study suggests that BPAF exposure is associated with SDH dysfunction, TCA cycle arrest, and lipid dysregulation. Whether BPAF directly inhibits SDH or acts through upstream mitochondrial targets warrants further structural and kinetic investigation.

Rare earth elements (REEs) are critical to modern industries but pose growing health risks due to increasing environmental release, and neodymium nitrate (Nd(NO₃)₃), a representative REE compound, lacks comprehensive toxicological data. To address this, we developed an integrated computational toxicology framework combining artificial intelligence-enhanced in vitro to in vivo extrapolation (AIVIVE), physiologically based pharmacokinetic (PBPK) modeling, quantitative in vitro to in vivo extrapolation (QIVIVE), and high-throughput toxicokinetic (HTTK) validation for mechanism-based risk assessment. A closed-loop "model-informed experimental design" was employed, where AIVIVE, using conditional generative adversarial networks (cGAN) predicted toxicity pathways from multiomics data, experimental determination of key toxicokinetic parameters (plasma protein binding and partition coefficients) calibrated HTTK predictions, and a PBPK-QIVIVE framework incorporating nonlinear features extrapolated in vitro EC₅₀ to human equivalent doses (HED), with Monte Carlo simulation and Sobol sensitivity analysis quantifying uncertainty. Results showed AIVIVE predicted transcriptomic responses with high fidelity (cosine similarity = 0.9986) and identified p53, apoptosis, and ferroptosis pathways with >85% accuracy. Experimental calibration revealed significant nonlinearity: plasma unbound fraction (fu) exhibited a U-shaped concentration dependence (0.556 at 1 μg/mL → 0.176 at 10 μg/mL → 0.965 at 100 μg/mL), while cellular partition coefficients (K) displayed an inverted U-shape (0.048-0.079). HTTK substantially underestimated fu (∼15-fold) and partition coefficients (2.4-5.5-fold). The integrated framework predicted a median HED of 0.032 mg/kg/day (95% CI: 0.012-0.098), with an 18% probability of exceeding the high-risk threshold (0.1 mg/kg/day). Sensitivity analysis identified fu (65%), K (22%), and EC₅₀ (11%) as the dominant uncertainty sources. Probabilistic integration with exposure data indicated a high safety margin for the general population but concerns for mining area residents (100% probability of margin of exposure 3 for neodymium nitrate.

Background/Objectives: Bisphenol AF (BPAF) is a prevalent environmental contaminant with demonstrated metabolic and immunological toxicity. This study aimed to investigate whether VDAC1 (Voltage-Dependent Anion Channel 1) mediates BPAF-induced succinate dysmetabolism and inflammatory responses in macrophages, and to evaluate the therapeutic potential of VDAC1 silencing. Methods: RAW264.7 macrophages were exposed to BPAF (0–2500 nM, 24 h) with or without VDAC1 siRNA transfection. Succinate levels, SDH activity, mitochondrial function (complexes I–V, ATP, membrane potential), and inflammatory markers (TNF-α, IL-6, IL-1β, ROS, MDA) were quantified. A 90-day oral toxicity study in C57BL/6J mice (0–32 mg kg⁻¹) assessed systemic inflammation and hepatic ultrastructure. p38 MAPK/NF-κB signaling was evaluated by Western blot and immunofluorescence. Results: BPAF elevated succinate 2.3-fold and decreased SDH activity by 48%, coinciding with reduced mitochondrial membrane potential and ATP synthesis (p < 0.01). Inflammatory cytokines and ROS were markedly increased. VDAC1 siRNA reversed these perturbations, restored complex II activity, and blunted p38 MAPK/NF-κB activation. In vivo, BPAF dose-dependently increased serum TNF-α, IL-6 and IL-1β, promoted NF-κB nuclear translocation and mitochondrial swelling, without altering body or liver weight; VDAC1 knockdown mitigated these effects. Conclusions: VDAC1 orchestrates BPAF-elicited succinate accumulation and macrophage inflammation via p38 MAPK/NF-κB signaling. Targeted VDAC1 silencing alleviates metabolic and inflammatory injury, offering a promising therapeutic strategy against BPAF-related diseases.

Bisphenol AF (BPAF) exposure is increasingly linked to metabolic disorders, yet the molecular initiating events (MIE) and key events (KE) leading to hepatic steatosis remain unclear. We constructed an adverse outcome pathway (AOP) to mechanistically connect BPAF-triggered macrophage–hepatocyte crosstalk to liver fat accumulation. Male C57BL/6 mice received daily oral gavage of 0, 0.5, 4, or 32 mg kg⁻¹ BPAF for 90 d, and Transwell co-cultures of RAW264.7 macrophages and AML12 hepatocytes were used for in vitro validation. Targeted metabolomics, western blotting, and lipid staining quantified succinate, pathway proteins, and steatosis. BPAF dose-dependently increased serum succinate (BMD = 6901.95 nM) and hepatic triglyceride (TG) (BMD = 874.26 nM). Cryo-EM docking revealed BPAF binding to SUCNR1 at 2.9 Å, disrupting the inactive-state conformation. In co-culture, BPAF-exposed macrophages released succinate that bound hepatocyte SUCNR1, suppressed Akt phosphorylation, and activated JNK. These KEs led to a 40% increase in lipid droplets and elevated TG, total cholesterol (TC), and free fatty acids (FFA) without liver weight gain. We propose the first AOP for BPAF-induced hepatic steatosis: BPAF–SUCNR1 binding (MIE) → macrophage succinate release (KE1) → SUCNR1-mediated Akt inhibition/JNK activation (KE2–4) → hepatic lipid accumulation (KE5) → steatosis (AO). These findings provide mechanistic insight for chemical risk assessment of BPAF and structurally related bisphenols.

Bisphenol A （BPA） and its analogs are collectively termed bisphenol compounds （BPs）， which are predominantly utilized in the manufacturing of polycarbonate plastics and epoxy resins. BPs are ubiquitous in diverse environmental matrices， human tissues， and metabolic products. Extensive research has demonstrated that BPs exert adverse effects on the nervous， reproductive， immune， and metabolic systems. After exposure in humans， BPs are primarily excreted in urine. Consequently， the development of efficient and robust analytical methods for BPs quantification in urine is essential for assessing population exposure levels. In this study， solid supported liquid-liquid extraction （SLE） was combined with ultra-high performance liquid chromatography-tandem mass spectrometry （UHPLC-MS/MS） technology to establish a high-throughput determination method for 17 BPs in human urine. After enzymatic hydrolysis treatment of the samples， the steps of fully automatic SLE purification， nitrogen blowing concentration and redissolution were carried out successively. Separation was performed using a CAPCELL PAK ADME chromatography column （100 mm×2.1 mm， 2 μm）， and gradient elution was carried out with 0.05 mmol/L ammonium fluoride aqueous solution and 0.05 mmol/L ammonium fluoride methanol solution as the mobile phases. MS detection was carried out using the electrospray ionization （ESI） negative ion scanning mode under the multi-reaction monitoring （MRM） mode. Qualitative analysis was conducted based on retention time and ion abundance ratio， and quantitative analysis was performed using the internal standard method. Under the optimized conditions， 17 BPs can be effectively separated. The linear relationships of the 17 BPs within the corresponding mass concentration ranges were good， and the correlation coefficients （r） were ≥0.998 6， the limits of detection （LODs） and quantification （LOQ） were 0.002-0.489 μg/L and 0.005-0.986 μg/L， respectively. Children’s mixed urine samples with low background content were selected as the matrix， and then spiked recovery tests were conducted at three spiked levels （low， medium and high）. The results showed that the recoveries of 17 BPs were 61.1%-121.7%， the intra-day RSDs were 1.3%-11.2%， and the inter-day RSDs were 3.7%-19.0%. This method was used to determine 50 random urine samples， and the results showed that a total of 11 BPs were detected. Among them， bisphenol S （BPS） and BPA had the highest detection rates， which were 98.0% and 86.0% respectively， and the median detection levels were 0.075 μg/L and 0.829 μg/L respectively. This method is simple to operate， sensitive and reliable. It is suitable for the rapid quantitative analysis of 17 BPs in human urine and can provide effective technical support for the risk assessment of BPs exposure in the population.

Objective: Neodymium nitrate (Nd(NO₃)₃) is widely used globally, raising concerns about its occupational and environmental safety. It enters the human body via the digestive system, accumulates in organs, and causes toxicity, including potential hepatotoxicity. However, the role of non-coding RNAs (ncRNAs) in Nd(NO₃)₃-induced liver injury remains unclear. This study aimed to identify key genes and regulatory pathways involved in Nd(NO₃)₃-induced hepatic injury using RNA sequencing (RNA-seq) and differential gene expression analysis. Methods: Mouse hepatocytes (AML12 cells) were exposed to Nd(NO₃)₃, and RNA-seq was performed to analyze the expression profiles of miRNA, lncRNA, circRNA, and mRNA. qPCR was used to validate the RNA-seq results. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were conducted to explore the functions and pathways associated with differentially expressed genes (DEGs). Results: Nd(NO₃)₃ exposure altered the expression of ferroptosis-related genes and induced significant changes in mRNA, miRNA, circRNA, and lncRNA expression levels. GO and KEGG analyses revealed that DEGs were closely related to cellular ferroptosis pathways. Specific miRNAs, lncRNAs, and circRNAs were significantly upregulated, suggesting their potential as biomarkers for Nd(NO₃)₃-induced ferroptosis and liver injury. Conclusion: This study provides the first comprehensive transcriptome database for Nd(NO₃)₃-induced liver injury, highlighting the involvement of ncRNAs in hepatotoxicity. These findings offer valuable insights for developing biomarkers and understanding the mechanisms underlying Nd(NO₃)₃-induced hepatic injury.

Bisphenol AF (BPAF) is widely utilized as an analog of bisphenol A (BPA) in the plastics industry. However, there is limited evidence on its neurodevelopmental toxicity. Existing studies suggest that BPAF has greater accumulation in vivo than other bisphenol analogs, and could pass through the placental barrier and the blood–brain barrier. In this study, we used the human neural progenitor cells line ReNcell CX, which was derived from 14-week human cortical brain tissue, as an in vitro model to investigate the neurodevelopmental toxicity effects of BPAF and BPA on ReNcell CX cells, and explored the possible mechanism by which BPAF induced neurodevelopmental toxicity on ReNcell CX cells. The results showed that BPAF reduced the proliferation of neural progenitor cells and changed the differentiation towards neurons after exposure for 24 h. Compared with BPA, ReNcell CX cells are more susceptible to BPAF exposure. In a 3D neurospheres model, BPAF affected the distance that neurons migrated outwards at the concentration of 2 μM. Furthermore, BPAF increased ROS levels in cells and reduced the expression of key proteins in the Nrf2/HO-1 pathway and its downstream molecules, such as SOD, GSH, and CAT. In conclusion, BPAF induces damage to critical nodes in neural progenitor cell development through the Nrf2/HO-1 pathway. Therefore, clarifying its neurodevelopmental toxicity and elaborating on the neurodevelopmental toxicity effects and mechanisms of bisphenol AF will help identify intervention targets for neurodevelopmental toxicity, and will have important public health significance for the safety assessment and risk prediction of bisphenol-related chemicals.

Ferroptosis, a form of regulated cell death driven by lipid peroxidation, has been implicated in the pathogenesis of liver diseases. This study investigates the role of circRNA_1156 in neodymium nitrate (Nd(NO₃)₃)-induced hepatocyte ferroptosis. Our in vitro experiments revealed that exposure to Nd(NO₃)₃ (1.2 µM) significantly reduced the viability of AML12 hepatocytes (p < 0.01), increased levels of reactive oxygen species (ROS) and malondialdehyde (MDA) (p < 0.001), and depleted glutathione (GSH) (p < 0.001). However, overexpression of circRNA_1156 effectively reversed these effects and suppressed the expression of ACSL4 and PKCβII (p < 0.01). In our in vivo experiments, chronic exposure to Nd(NO₃)₃ (7–55 mg/kg for 180 days) induced hepatic iron deposition, mitochondrial damage, and activation of the ACSL4/PKCβII pathway (p < 0.01). These adverse effects were significantly ameliorated by circRNA_1156 overexpression (p < 0.05). Our findings identify circRNA_1156 as a novel inhibitor of Nd(NO₃)₃-induced ferroptosis via downregulation of the ACSL4/PKCβII pathway, providing valuable therapeutic insights for hepatotoxicity caused by rare earth element compounds.