Nanoplastics (NPs) represent a growing concern for global environmental health, particularly in marine ecosystems where they predominantly accumulate. The impact of NPs on marine benthic organisms, such as bivalves, raises critical questions regarding ecological integrity and food safety. Traditional methods for assessing NP toxicity are often limited by their time-intensive nature and ethical considerations. Herein, we explore the toxicological effects of NPs on the marine bivalve Ruditapes philippinarum, employing a combination of in vitro cellular assays and advanced modeling techniques. Results indicate a range of adverse effects at the organismal level, including growth inhibition (69.5-108%), oxidative stress, lipid peroxidation, and DNA damage in bivalves, following exposure to NPs at concentrations in the range of 1.6 × 109-1.6 × 1011 particles/mL (p/mL). Interestingly, the growth inhibition predicted by models (54.7-104%), based on in vitro cellular proliferation assays, shows strong agreement with the in vivo outcomes of NP exposure. Furthermore, we establish a clear correlation between cytotoxicity observed in vitro and the toxicological responses at the organismal level. Taken together, this work suggests that the integration of computational modeling with in vitro toxicity assays can predict the detrimental effects of NPs on bivalves, offering insightful references for assessing the environmental risk assessment of NPs in marine benthic ecosystems.

Oxidative/nitrosative damage takes part in chronic disease development, which generates an urgent need for intervention and better therapies to manage them. The scientific community has demanded easy-to-run, cheap, and reliable methods for cellular antioxidant activity assays. This work standardised and validated an erythrocyte cellular antioxidant activity and membrane protection/injury (HERYCA-P) protocol to study food-derive extracts. The method measures intracellular reactive oxygen species (ROS) generation, lipoperoxidation, and haemolysis induced by 2,2\'-azobis(2-amidinopropane) dihydrochloride. Quercetin decreased ROS generation by 50.4% and haemolysis by 2.2%, while ascorbic acid inhibited lipid peroxidation by 40.1%. Total phenolic contents of teas were correlated with decreased ROS generation (r = -0.924), lipoperoxidation (r = -0.951), and haemolysis (r = -0.869). The erythrocyte ROS generation and lipoperoxidation were also associated with CUPRAC (r = -0.925; r = -0.951) and hydroxyl radical scavenging activity (r = -0.936; r = -0.949). The precision rates of antioxidant standards and tea samples were below 15%. HERYCA-P is feasible as a complementary antioxidant assay for food matrices.

The potential health risk caused by long-term exposure to heavy metals in household dust is not only depended on their total content, but also bioaccessibility. In this study, twenty-one dust samples were collected from residential buildings, schools, and laboratories in 14 provincial-capital/industrial cities of China, aiming to evaluate the total contents, fractionation, bioaccessibility and health risks of nine heavy metals (As, Cd, Cr, Ni, Pb, Mn, Zn, Fe, and Cu). Results showed that the highest levels of Cd, Cr, Ni and Zn were found in laboratory dust, As, Pb and Mn in school dust, and Fe and Cu in residential dust, indicating different source profiles of the heavy metals. The mean bioaccessibility of the heavy metals across all samples as evaluated using SBRC (Solubility Bioavailability Research Consortium), IVG (In Vitro Gastrointestinal), and PBET (Physiologically Based Extraction Test) assays was 58.4%, 32.4% and 17.2% in gastric phase (GP), and 24.9%, 21.9% and 9.39% in intestinal phase (IP), respectively. Cadmium had the highest content in the fractions of E1+C2 (43.7%), as determined by sequential extraction, and Pb, Mn, and Zn had a higher content in E1+C2+F3 (64.2%, 67.2%, 78.8%), resulting in a higher bioaccessibility of these heavy metals than others. Moreover, the bioaccessibility of most heavy metals was inversely related to dust pH (R = -0.18 in GP; -0.18 in IP; P < 0.01) and particle size, while a positive correlation was observed with total organic carbon (R = 0.40 in GP; 0.38 in IP; P < 0.01). The exposure risk calculated by the highest bioaccessibility was generally lower than that calculated by the total content. However, Pb in one school dust sample had an unacceptable carcinogenic risk (adult risk = 1.19 × 10-4; child risk = 1.08 × 10-4). This study suggests that bioaccessibility of heavy metals in household dust is likely related to geochemical fractions and physical/chemical properties. Further research is needed to explore the sources of bioaccessible heavy metals in household dust.

The goal of this study is to look into the pharmacological mechanism of Bruceae Fructus in conjunction with GEO, network pharmacology, and in vitro assays for the treatment of laryngeal cancer to provide theoretical support for its therapeutic use. The active components and matching targets of Bruceae Fructus were retrieved from the TCMSP database, while genes linked with laryngeal cancer were obtained from the GEO, GeneCards, DisGeNET, and DrugBank databases. Besides, the components and targets were supplemented by literatures in PubMed database. Cytoscape software was used to create the active ingredients-target network diagram. The String database was used to build the PPI network. Following that, the core targets were subjected to GO enrichment and KEGG pathway analysis using the DAVID database. Finally, AutoDock was used to perform molecular docking between the core components and the core targets. To investigate the biological effects of beta-sitosterol, the viability of laryngeal cancer cells was assessed after beta-sitosterol therapy using the MTS technique. Following that, how beta-sitosterol affected colony formation after 14 days of culture of treated cells was researched. Flow cytometry was utilized to detect apoptosis to examine the influence of beta-sitosterol on laryngeal cancer cell apoptosis, and then detected mRNA and protein expression levels of 10 key genes by RT-qPCR and Western Blot assay. There were 1258 laryngeal cancer-related genes and 15 Bruceae Fructus components, with beta-sitosterol and luteolin serving as key components. Bruceae Fructus\' primary targets against laryngeal cancer were IL6, JUN, TNF, IL2, IL4, IFNG, RELA, TP53, CDKN1A, and AKT1. GO enrichment yielded 41 CC, 78 MF, and 383 BP. Platinum drug resistance, the PI3K-Akt signaling pathway, the p53 signaling pathway, apoptosis, the HIF-1 signaling pathway, and 147 additional pathways have been added to KEGG. The results of molecular docking revealed that the core components had a high affinity for the core target. The results of the cell experiment indicate that beta-sitosterol suppressed Hep-2 cell activity in a concentration-dependent manner. Besides, beta-sitosterol has powerful antiproliferative properties in Hep-2 cells. Flow cytometry results showed that beta-sitosterol promoted laryngeal cancer cell apoptosis in a concentration-dependent manner. The results of RT-qPCR and Western Blot assay showed that the mRNA and protein expression levels of TP53, JUN, TNF-α, CDKN1A, and IL-2 were significantly up-regulated after beta-sitosterol treatment, while the mRNA and protein expression levels of RELA, AKT1, IL-6, IFNG, and IL-4 were significantly down-regulated. This study integrating GEO, network pharmacology, and in vitro assays investigated the probable mechanism of Bruceae Fructus\' anti-laryngeal cancer activity, which can give a theoretical foundation for additional future animal experiments.

Dietary calcium (Ca) intake can alleviate fluoride (F) induced fluorosis to maintain bone health. However, it is unclear whether calcium supplements can reduce the oral bioavailability of F present in contaminated soils. Here we evaluated the effects of Ca supplements on F bioavailability in three soils using an in vitro method (Physiologically Based Extraction Test) and an in vivo mouse model. Seven Ca salts, commonly used in calcium supplements, significantly reduced the F bioaccessibility in the gastric and small intestinal phases. Particularly for Ca phosphate at 150 mg Ca supplementation, F bioaccessibility in the small intestinal phase was reduced from 35.1-38.8% to 0.7-1.9% where soluble F concentrations were less than 1 mg/L. Overall, the eight Ca tablets tested in this study showed greater efficiency at decreasing F solubility. The in vitro bioaccessibility after Ca supplementation was consistent with the relative bioavailability of F. As supported by X-ray photoelectron spectroscopy, a possible mechanism is that freed F can be bound by Ca to form insoluble CaF2 and exchanged with OH groups from Al/Fe hydroxide to strongly adsorb F. These findings provide evidence of Ca supplementation in reducing health risks associated soil F exposure.

In vitro strategies have widely been used to assess bioaccessibility of organic pollutants in soils. However, studies for comparing in vitro models with in vivo data are still limited. In this study, Dichlorodiphenyltrichloroethane (DDT) and its metabolites (called as DDTr) bioaccessibility in nine contaminated soils were measured using physiologically based extraction test (PBET), in vitro digestion model (IVD), and Deutsches Institut für Normung (DIN) with/without Tenax as an absorptive sink, and DDTr bioavailability was assessed using an in vivo mouse model. Whether or not Tenax was added, DDTr bioaccessibility significantly varied among three methods, suggesting that DDTr bioaccessibility depended on the in vitro method employed. Multiple linear regression analysis indicated that sink, intestinal incubation time and bile content are identified to be the dominant factors in controlling DDTr bioaccessibility. Comparison of in vitro and in vivo results demonstrated that DIN assay with Tenax (TI-DIN) provided the best prediction for DDTr bioavailability (r2 = 0.66, slope=0.78). After extending intestinal incubation time to 6 h or increasing bile content to 4.5 g/L (same to DIN assay) of the TI-PBET and TI-IVD assays, the in vivo-in vitro correlation will improved significantly, with r2 = 0.76 and slope= 1.4 for TI-PBET and r2 = 0.84 and slope= 1.9 for TI-IVD under 6 h intestinal incubation, and r2 = 0.59 and slope= 0.96 for TI-PBET and r2 = 0.51 and slope= 1.0 for TI-IVD under 4.5 g/L of bile content. The results suggest that it is essential to understand these key factors influencing bioaccessibility for the development of standardized in vitro methods, which helps to refine the risk assessment of human exposure to contaminants via soil ingestion.

Studies analyzing the in vitro bioaccessibility (BAc) of heavy metals in biochar-amended soils are currently lacking. The present study aimed to assess the metal BAc in Cd- and Pb-spiked acidic Ultisol samples treated individually with 2% (w/w) maize, rice, wheat, soybean, and pea straw-derived biochar. The results indicate that the Cd-BAc simulated in gastric phase (GP) decreased from 78.4% to 66.5-72.3% and the Pb-BAC decreased from 74.3% to 67.2-69.2%; however, the Cd-BAc in the intestinal phase (IP) decreased from 35.6% to 27.9-33.5% and the Pb-BAc decreased from 34.7% to 29.7-32.9% after 120 d of incubation with biochar application compared to the un-amended Ultisol. The Cd- and Pb-BAc in both GP and IP were significantly negatively correlated with soil pH, CEC, and organic carbon (P < 0.05), which increased after biochar application. The soybean straw-derived biochar amendment has the greatest potential to decrease the BAc of Cd and Pb in the GP and IP, owing to the highest level of CEC, SOC, TC and TN among all soil samples. Moreover, the BAc was positively correlated with the exchangeable, and exchangeable + carbonate-bound Cd and Pb fractions (P < 0.05), indicating these fractions had a dominant influence on the BAc of cationic heavy metals. Therefore, crop straw-derived biochar amendment can decrease the BAc of Cd and Pb in acidic Ultisol, and thus mitigate the health risks posed by these metals from incidental ingestion.

Parabens are ubiquitous pollutants in the environment and humans due to their wide applications in food, pharmaceuticals, and personal care products. Although the estrogenic activity of some parabens has been confirmed, the underlying mechanisms and the structure-estrogenic activity relationship are still largely unclear. Here, we systematically used in silico and in vitro approaches to investigate the estrogenic potency of typical parabens, including methyl-, ethyl-, propyl-, iso-propyl-, butyl-, iso-butyl- and benzyl-paraben. Molecular dynamics simulations and binding free energy calculations were combined to investigate the atomic-level mechanism of paraben binding to estrogen receptors (ERs). Computational analysis showed that ER were the targets of tested parabens and kept a stable agonist conformation. The calculated total binding free energies suggested that van der Waals interactions were the major driving forces for paraben-ER interaction and correlated with the structure of paraben side chains. In in vitro assays, paraben with an aromatic side chain, benzyl-paraben, showed the strongest estrogenic activity at 0.01 μM and the EC50 at 0.796 ± 0.307 μM, on par with levels commonly detected in human organs. Among tested parabens with an alkyl side chain, the estrogenicity increased as the side chain length increased from 1 to 4, but no significant difference appeared between parabens with isomeric alkyl side chains (propyl- vs isopropyl and butyl- vs iso-butylparaben). The estrogenic activity of parabens was significantly related to the calculated binding energies (R2 = 0.94, p = 0.0012), depending on the side chains of parabens. Our findings provide a significant mechanism for parabens to disrupt estrogenic function and considerations for structural optimization from the perspective of environmental protection.

To understand how Cd in different fractions contributes to Cd bioaccessibility by in vitro assays, Cd bioaccessibility in 12 contaminated soils was determined by four assays (UBM, SBRC, IVG, and PBET) and correlated with different Cd fractions based on a sequential extraction scheme. The Cd bioaccessibility in the gastric phase (GP) was high (35-107%, averaging at 77%), implicating high risk to human health, while it decreased to 19-88% averaging at 47% in the intestinal phased (IP). From the GP to IP, the reduction of extractable Cd (0.45-48 mg kg-1) and Fe (118-3884 mg kg-1) showed significant correlation (R = 0.54-0.74) via UBM, SBRC, and IVG, suggesting co-precipitation with Fe and/or sorption onto Fe oxides maybe responsible for decrease in Cd bioaccessibility. Although Cd bioaccessibility varied among assays, their results show some consistency based on their correlation in the GP (R = 0.56-0.90) and IP (0.34-0.73, excluding UBM-IP and PBET-IP). Sequential extraction data show that Cd was primarily associated with the exchangeable fraction (E1; 7.05-72.9%, averaging 39.4%). The carbonate (C2; 6.86-44.8%, 21.9%) and Fe/Mn oxides fraction (F3; 12.5-53.6%, 28.2%) were similar, while organic (O4; 0.62-25.0%, 7.91%) and residual fraction (R5; 0.22-8.54%, 2.62%) were the lowest. Significant correlation (R = 0.59-0.88) between the first two fractions (E1+C2) and bioaccessible Cd suggest they were the main sources of bioaccessible Cd in those contaminated soils.

Single extraction procedures (SEPs) have been extensively conducted to determine Cd bioavailability (Cd-Bav) in soils. However, whether SEPs can simultaneously predict Cd accumulation in crop grains and bioaccessibility (Cd-Bac) in soils remains unclear. To assess their suitability, the Cd-Bav in 20 contaminated soils (containing 0.27-56.59 mg/kg Cd) determined by four SEPs (including DTPA, EDTA, HOAc and HCl) was compared with Cd concentrations in crop grains (wheat and rice) and Cd-Bac in soils (based on SBET and PBET assays). The results indicated that both Cd-Bav (0-103.2%) and Cd-Bac (0-110.4%) in soils varied greatly with the methods used. The Cd-Bav obtained from chelators (DTPA and EDTA) was generally greater in low-Cd soils but lower in high-Cd soils as compared to those obtained from acid solutions (HOAc and HCl). Regression analysis revealed that bioavailable Cd concentrations in soils were linearly correlated with Cd concentrations in wheat grains (R2 = 0.88-0.91); however, no significant correlation was found for rice grains. The Cd-Bac in soils was significantly correlated with Cd-Bav obtained from HOAc (R2 = 0.55-0.59) or HCl (R2 = 0.60-0.68), but not with those obtained from chelators (DTPA and EDTA). Our data suggest that SEPs, particularly the HCl method, have great potential to simultaneously predict Cd accumulation in wheat grains and Cd-Bac in contaminated soils.