Cytotoxicity of emerging halophenylacetamide disinfection byproducts in drinking water: Mechanism and prediction.

Halophenylacetamides (HPAcAms) have been identified as a new group of nitrogenous aromatic disinfection byproducts (DBPs) in drinking water, but the toxicity mechanisms associated with HPAcAms remain almost completely unknown. In this work, the cytotoxicity of HPAcAms in human hepatoma (HepG2) cells was evaluated, intracellular oxidative stress/damage levels were analyzed, their binding interactions with antioxidative enzyme were explored, and a quantitative structure-activity relationship (QSAR) model was established. Results indicated that the EC50 values of HPAcAms ranged from 2353 µM to 9780 µM, and the isomeric structure as well as the type and number of halogen substitutions could obviously induce the change in the cytotoxicity of HPAcAms. Upon exposure to 2-(3,4-dichlorophenyl)acetamide (3,4-DCPAcAm), various important biomarkers linked to oxidative stress and damage, such as reactive oxygen species, 8­hydroxy-2-deoxyguanosine, and cell apoptosis, exhibited a significant increase in a dose-dependent manner. Moreover, 3,4-DCPAcAm could directly bind with Cu/Zn-superoxide dismutase and induce the alterations in the structure and activity, and the formation of complexes was predominantly influenced by the van der Waals force and hydrogen bonding. The QSAR model supported that the nucleophilic reactivity as well as the molecular compactness might be highly important in their cytotoxicity mechanisms in HepG2 cells, and 2-(2,4-dibromophenyl)acetamide and 2-(3,4-dibromophenyl)acetamide deserved particular attention in future studies due to the relatively higher predicted cytotoxicity. This study provided the first comprehensive investigation on the cytotoxicity mechanisms of HPAcAm DBPs.

Regulation mechanisms of ferric ions release from iron-loaded transferrin protein caused by nano-sized polystyrene plastics-induced conformational and structural changes.

Currently, the binding of iron-binding protein transferrin (TF) with NPs and their interaction mechanisms have not been completely elucidated yet. Here, we probed the conformation-dependent release of Fe ions from TF induced by nano-sized polystyrene plastics (PS-NPs) using dialysis, ICP-MS, multi-spectroscopic techniques, and computational simulation. The results showed that the release of free Fe ions from TF was activated after PS-NPs binding, which displayed a clear dose-effect correlation. PS-NPs binding can induce the unfolding and loosening of polypeptide chain and backbone of TF. Alongside this we found that the TF secondary structure was destroyed, thereby causing TF protein misfolding and denaturation. In parallel, PS-NPs interacted with the chromophores, resulting in the occurrence of fluorescence sensitization effects and the disruption of the surrounding micro-environment of aromatic amino acids. Also, the binding of PS-NPs induced the formation of new aggregates in the PS-NPs-TF system. Further simulations indicated that PS-NPs exhibited a preference for binding to the hinge region that connects the C-lobe and N-lobe, which is responsible for the Fe ions release and structural alterations of TF. This finding provides a new understanding about the regulation of the release of Fe ions of iron-loaded TF through NPs-induced conformational and structural changes.

Molecular mechanisms of polystyrene nanoplastics and alpha-amylase interactions and their binding model: A multidimensional analysis.

Plastic fragments are widely distributed in different environmental media and has recently drawn special attention due to its difficulty in degradation and serious health and environmental problems. Among, nanoplastics (NPs) are smaller in size, larger in surface/volume ratio, and more likely to easily adsorb ambient pollutants than macro plastic particles. Moreover, NPs can be easily absorbed by wide variety of organisms and accumulate in multiple tissues/organs and cells, thus posing a more serious threat to living organisms. Alpha-amylase (α-amylase) is a hydrolase, which can be derived from various sources such as animals, plants, and microorganisms. Currently, no studies have concentrated on the binding of NPs with α-amylase and their interaction mechanisms by employing a multidimensional strategy. Hence, we explored the interaction mechanisms of polystyrene nanoplastics (PS-NPs) with α-amylase by means of multispectral analysis, in vitro enzymatic activity analysis, and molecular simulation techniques under in vitro conditions. The findings showed that PS-NPs had the capability to bind with the intrinsic fluorescence chromophores, leading to fluorescence changes of these specific amino acids. This interaction also caused the alterations in the micro-environment of the fluorophore residues mainly tryptophan (TRP) and tyrosine (TYR) residues of α-amylase. PS-NPs interaction promoted the unfolding and partial expansion of polypeptide chains and the loosening of protein skeletons, and destroyed the secondary structure (increased random coil contents and decreased α-helical contents) of this protein, forming a larger particle size of the PS-NPs-α-amylase complex. Moreover, the enzymatic activity of α-amylase in vitro was found to be inhibited in a concentration dependent manner, thereby impairing its physiological functions. Further molecular simulation found that PS-NPs had a higher tendency to bind to the active site of α-amylase, which is the cause for its structural and functional changes. Additionally, the hydrophobic force played a major role in mediating the binding interactions between PS-NPs and α-amylase. Taken together, our study indicated that PS-NPs interaction can initiate the abnormal physiological functions of α-amylase through PS-NPs-induced structural and conformational alternations.

The combined effects of polystyrene nanoplastics with nickel on oxidative stress and related toxic effects to earthworms from individual and cellular perspectives.

Nanoplastics may adsorb other pollutants in the environment due to their high specific surface area and small size. We used earthworms as experimental organisms to evaluate the ecotoxicity of NPs and Ni combined pollution at the individual and cellular levels. The results showed that when only 20 mg/L Ni2+ was added to the combined pollution system, the antioxidant system of earthworm coelomocytes was destroyed to a certain extent, the ROS level increased, the cell viability decreased significantly, and the redox balance was destroyed. With the introduction of PS-NPs and the increase of concentration, the oxidative damage in the coelomocytes of earthworms gradually increased, and finally tended to be stable when the maximum concentration of 50 mg/L PS-NPs and Ni were exposed together. At the animal level, the activities of CAT and SOD decreased within 28 days of exposure, and the combined pollution showed a synergistic effect. At the same time, it promoted the synthesis of GST in earthworms, improved their detoxification ability and reduced oxidative damage. The changes of T-AOC and MDA showed that the combined pollution caused the accumulation of ROS and caused more serious toxicological effects. With the increase of exposure time, the antioxidant system of earthworms was continuously destroyed, and the oxidative damage was serious, which induced more serious lipid peroxidation and caused the damage of earthworm body wall structure.

New mechanistic insights into halogen-dependent cytotoxic pattern of monohaloacetamide disinfection byproducts.

As highly toxic nitrogenous disinfection byproducts (DBPs), monohaloacetamides (monoHAcAms) generally exhibited a cytotoxic rank order of iodoacetamide ˃ bromoacetamide ˃ chloroacetamide. However, the mechanisms underlying the halogen-dependent cytotoxic pattern remain largely veiled as yet. In this work, oxidative stress/damage levels in monoHAcAm-treated Chinese hamster ovary cells were thoroughly analyzed, and binding interactions between monoHAcAms and antioxidative enzyme Cu/Zn-superoxide dismutase (Cu/Zn-SOD) were investigated by multiple spectroscopic techniques and molecular docking. Upon exposure to monoHAcAms, the intracellular levels of key biomarkers associated with oxidative stress/damage, including reactive oxygen species, malondialdehyde, lactate dehydrogenase, 8-hydroxy-2-deoxyguanosine, cell apoptosis, and G1 cell cycle arrest, were all significantly increased in a dose-response manner with the same halogen-dependent rank order as their cytotoxicity. Moreover, this rank order was also determined to be applicable to the monoHAcAm-induced alterations in the conformation, secondary structure, and activity of Cu/Zn-SOD, the microenvironment surrounding aromatic amino acid residues in Cu/Zn-SOD, as well as the predicted binding energy of SOD-monoHAcAm interactions. Our results revealed that the halogen-dependent cytotoxic pattern of monoHAcAms was attributed to their differential capacity to induce oxidative stress/damage and their interaction with antioxidative enzyme, which contribute to a better understanding of the halogenated DBP-induced toxicological mechanisms.

Molecular mechanisms of nano-sized polystyrene plastics induced cytotoxicity and immunotoxicity in Eisenia fetida.

Nanoplastics (NPs) are currently everywhere and environmental pollution by NPs is a pressing global problem. Nevertheless, until now, few studies have concentrated on the mechanisms and pathways of cytotoxic effects and immune dysfunction of NPs on soil organisms employing a multidimensional strategy. Hence, earthworm immune cells and immunity protein lysozyme (LZM) were selected as specific receptors to uncover the underlying mechanisms of cytotoxicity, genotoxicity, and immunotoxicity resulting from exposure to polystyrene nanoplastics (PS-NPs), and the binding mechanisms of PS-NPs-LZM interaction. Results on cells indicated that when earthworm immune cells were exposed to high-dose PS-NPs, it caused a notable rise in the release of reactive oxygen species (ROS), resulting in oxidative stress. PS-NPs exposure significantly decreased the cell viability of earthworm immune cells, inducing cytotoxicity through ROS-mediated oxidative stress pathway, and oxidative injury effects, including reduced antioxidant defenses, lipid peroxidation, DNA damage, and protein oxidation. Moreover, PS-NPs stress inhibited the intracellular LZM activity in immune cells, resulting in impaired immune function and immunotoxicity by activating the oxidative stress pathway mediated by ROS. The results from molecular studies revealed that PS-NPs binding destroyed the LZM structure and conformation, including secondary structure changes, protein skeleton unfolding/loosening, fluorescence sensitization, microenvironment changes, and particle size changes. Molecular docking suggested that PS-NPs combined with active center of LZM easier and inhibited the protein function more, and formed a hydrophobic interaction with TRP 62, a crucial amino acid residue closely associated with the function and conformation of LZM. This is also responsible for LZM conformational changes and functional inhibition /inactivation. These results of this research offer a fresh outlook on evaluating the detriment of NPs to the immune function of soil organisms using cellular and molecular strategies.

Novel evidence on iodoacetic acid-induced immune protein functional and conformational changes: Focusing on cellular and molecular aspects.

Elevated levels of iodide occur in raw water in certain regions, where iodination disinfection byproducts are formed during chloramine-assisted disinfection of naturally iodide-containing water. Iodoacetic acid (IAA) is one of the typical harmful products. The mechanisms underlying IAA-induced immunotoxicity and its direct effects on biomolecules remained unclear in the past. Cellular, biochemical, and molecular methods were used to investigate the mechanism of IAA-induced immunotoxicity and its binding to lysozyme. In the presence of IAA, the cell viability of coelomocytes was significantly reduced to 70.8 %, as was the intracellular lysozyme activity. Upon binding to IAA, lysozyme underwent structural and conformational changes, causing elongation and unfolding of the protein due to loosening of the backbone and polypeptide chains. IAA effectively quenched the fluorescence of lysozyme and induced a reduction in particle sizes. Molecular docking revealed that the catalytic residue, Glu 35, which is crucial for lysozyme activity, resided within the docking range, suggesting the preferential binding of IAA to the active site of lysozyme. Moreover, electrostatic interaction emerged as the primary driving force behind the interaction between IAA and lysozyme. In conclusion, the structural and conformational changes induced by IAA in lysozyme resulted in impaired immune protein function in coelomocytes, leading to cellular dysfunction.

Decoding the biological toxicity of phenanthrene on intestinal cells of Eisenia fetida: Effects, toxicity pathways and corresponding mechanisms.

Phenanthrene is frequently detected and exists extensively in the soil environment, and its residues inevitably impose a significant threat to soil organisms. Exposure to and toxicity of phenanthrene on earthworms has been extensively studied before, however, the possible mechanisms and related pathways associated with phenanthrene-triggered toxicity at the intestinal cell level remain unclear. Herein, primary intestinal cells isolated from Eisenia fetida (Annelida, Oligochaeta) intestine were used as targeted receptors to probe the molecular mechanisms involved in ROS-mediated damaging effects and the potential pathways of phenanthrene-induced toxicity at cellular and sub-cellular levels. Results indicated that phenanthrene exposure induced oxidative stress by activating intracellular ROS (elevated O2-, H2O2, and OH- content) bursts in E. fetida intestinal cells, causing various oxidative damage effects, including lipid peroxidation (increased MDA content), protein oxidation (enhanced PCO levels), and DNA damage (enhanced 8-OHdG levels). The enzymatic and non-enzymatic strategies in earthworm cells were activated to mitigate these detrimental effects by regulating ROS-mediated pathways involving defense regulation. Also, phenanthrene stress destroyed the cell membrane of E. fetida intestinal cells, resulting in cellular calcium homeostasis disruption and cellular energetic alteration, ultimately causing cytotoxicity and cell apoptosis/death. More importantly, the mitochondrial dysfunction in E. fetida cells was induced by phenanthrene-caused mitochondrial membrane depolarization, which in turn caused un-controlled ROS burst and induced apoptosis through mitochondria-mediated caspase-3 activation and ROS-mediated mitochondrial-dependent pathway. Furthermore, exposure to phenanthrene activated an abnormal mRNA expression profile associated with defense regulation (e.g., Hsp70, MT, CRT, SOD, CAT, and GST genes) in E. fetida intestinal cells, resulting in various cellular dysfunctions and pathological conditions, eventually, apoptotic cell death. Taken together, this study offers valuable insights for probing the toxic effects and underlying mechanisms posed by phenanthrene at the intestinal cell level, and is of great significance to estimate the detrimental side effects of phenanthrene on soil ecological health.

Compound effect and mechanism of oxidative damage induced by nanoplastics and benzo [a] pyrene.

Nanoplastics and polycyclic aromatic hydrocarbons (PAHs) are ubiquitous in soil environments. In order to objectively evaluate the toxic interaction between polystyrene nanoplastics (PS NPs) and benzo [a] pyrene (BaP), oxidative damage at the level of earthworm cells and biomacromolecules was investigated by experiments combined with molecular dynamics simulation. Studies on cells reveal that PS NPs and BaP had synergistic toxicity when it came to causing oxidative stress. Cellular reactive oxygen species (ROS) levels under combined pollutant exposure were 24% and 19% higher, respectively than when PS NPs and BaP were exposed alone (compared to the blank group). In addition, BaP and PS NPs inhibited the ability of CAT to decompose H2O2 by affecting the structure of the proximal amino acid Tyr 357 in the active center of CAT, which exacerbated oxidative stress to a certain extent. Therefore, the synergistic toxic effect of BaP and PS NPs is due to the mutual complement of the two to the induction of protein structural looseness, and the strengthening of the stability of the conjugate (CAT-BaP-PS) under the weak interaction. This work provides a new perspective and approach on how to talk about the toxicity of combined pollutants.

Environmental relevant concentrations of polystyrene nanoplastics and lead co-exposure triggered cellular cytotoxicity responses and underlying mechanisms in Eisenia fetida.

Heavy metal pollution of soils and the widespread use of plastics have caused environmental problems worldwide. Nanoplastics (NPs) contaminants in water and soil environments can adsorb heavy metals, thereby affecting the bioavailability and toxicity of heavy metals. In this paper, the effect of co-exposure of polystyrene microspheres with 100 nm particle size and lead acetate (Pb) on the Eisenia fetida coelomocytes was investigated. The environmental concentration of NPs used was 0.01 mg/L and the concentration of Pb ranged from 0.01 to 1 mg/L, and the exposed cells were incubated at 298 k for 24 h. Our study demonstrated that exposure of cells to environmental relevant concentrations of NPs did not significantly affect the cytotoxicity of Pb exposure. It was shown that co-exposure induced cellular production of reactive oxygen species (ROS, increased to 134.4 %) disrupted the antioxidant system of earthworm body cavity cells, activated superoxide dismutase and catalase (CAT), produced reduced glutathione, and inhibited glutathione-dependent enzyme (GST) activity (Reduced to 64 %). Total antioxidant capacity (T-AOC) is first enhanced against ROS due to the stress of NPs and Pb. When the antioxidant reserves of cells are exhausted, the antioxidant capacity will decrease. The level of malondialdehyde, a biomarker of eventual lipid peroxidation, increased to 231.7 %. At the molecular level, due to co-exposure to NPs and Pb, CAT was loosely structured and the secondary structure is misfolded, which was responsible for exacerbating oxidative damage in E. fetida coelomocytes. The findings of this study have significant implications for the toxicological interaction and future risk assessment of co-contamination of NPs and Pb in the environment.

Molecular interaction mechanisms and cellular response of superoxide dismutase and catalase to fluoranthene.

As an important raw material and intermediate product of the petrochemical industry, fluoranthene (Fla) can be emitted with industrial activities and has become a typical polycyclic aromatic hydrocarbon enriched in the Chinese topsoil layer, posing a significant threat to sensitive soil biota. Here, multispectral tools and molecular simulation techniques were integrated to elucidate the molecular mechanism of Fla interaction with key antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) at the molecular level. Meanwhile, we further revealed the cellular responses of SOD and CAT and the associated redox states in earthworm (Eisenia fetida) coelomocytes based on the molecular-level results. Our results showed that the exposure to Fla affected the backbone structure of SOD and CAT molecules and resulted in the formation of Fla-SOD polymers as well as an overall reduction in the size of the Fla-CAT binding system. Fla altered the microenvironment around Tyr residues in the SOD molecule and quenched the endogenous fluorescence of Tyr within the CAT molecule. In earthworm coelomocytes, Fla at 60 and 80 µM resulted in a significant elevation of CAT and SOD activities by 114% (p = 0.032) and 6.09% (p = 0.013), respectively. Molecular simulation results suggested that Fla-induced changes in the structure and conformation of SOD and CAT may be the key reason for their altered activities. The related redox homeostasis detection in earthworm coelomocytes indicated that high concentrations (80 µM) of Fla led to a significant accumulation of intracellular ROS (p = 0.018) and resulted in the development of lipid peroxidation. Our work contributes to an in-depth understanding of the biological effect of Fla to sensitive soil fauna, thus providing new ideas for Fla ecological risk prevention and control.

Microbial electron flow promotes naphthalene degradation in anaerobic digestion in the presence of nitrate electron acceptor: Focus on electron flow regulation and microbial interaction succession.

Microbial electron flow (MEF) is produced from microbial degradation of organic compounds. Regulating MEF to promote organic pollutants biodegradation such as naphthalene (Nap) is a potential way but remains a lack of theoretical basis. Here, we regulated MEF by adding electron acceptor NO3- to achieve 2.6 times increase of Nap biodegradation with cyclodextrin as co-metabolism carbon source. With the NO3- addition, the genes inhibited by Nap of electron generation significantly up-regulated. Especially, key genes ubiD and nahD for anaerobic Nap degradation significantly up-regulated respectively 3.7 times and 6.7 times. Moreover, the ability of electron transfer in MEF was also improved consistent with 7.2 times increase of electron transfer system (ETS) activity. Furthermore, total 60 metagenome-assembled genomes (MAGs) were reconstructed through the metagenomic sequencing data with assembly and binning strategies. Interestingly, it was also first found that the Klebsiella MAG. SDU (Shandong University) 14 had the ability of simultaneous Nap biodegradation and denitrification. Our results firstly offered an effective method of regulating MEF to promote polycyclic aromatic hydrocarbons (PAHs) degradation and simultaneous methanogenesis.

The "island sign" on diffusion-weighted imaging predicts early neurological deterioration in penetrating artery territory infarctions: a retrospective study.

BACKGROUND: Early neurological deterioration (END) sometimes occurs in patients with penetrating artery territory infarction (PATI) and leads to poor prognosis. In this study, we analyzed clinical and neuroimaging characteristics of PATI, and focused on the infarct patterns on diffusion-weighted imaging (DWI). We tried to investigate whether the "island sign" pattern is associated with END. METHODS: We enrolled consecutive patients admitted with acute PATI within 48 h after onset from May 2020 to July 2022. They were divided into with and without the "island sign" pattern on DWI. According to infarct location, all the patients were classified into two groups: the territories of the lenticulostriate arteries (LSA) and paramedian pontine arteries (PPA). The patients in each group were further divided into two groups according to whether they developed END or not. Through analyzing the clinical and neuroimaging characteristics of the patients, we tried to identify the factors that might associated with the "island sign" pattern and the potential predictors of END within the LSA and PPA groups. RESULTS: Out of the 113 patients enrolled in this study, END was found in 17 patients (27.9%) in the LSA group and 20 patients (38.5%) in the PPA group. The "island sign" was found in 26 (23%) patients. In the multivariate analysis, the independent predictors of END in the LSA group were the "island sign" (OR 4.88 95% CI 1.03-23.2 P = 0.045) and high initial National Institute of Health Stroke Scale (NIHSS) (OR 1.79 95% CI 1.08-2.98 P = 0.024) and in the PPA group was the presence of lesions extending to the ventral pontine surface (OR 7.53 95% CI 1.75-32.37 P = 0.007). CONCLUSIONS: The predictive factors for END were different in the LSA and PPA groups. The "island sign" was particularly associated with END in the LSA group.

Probing the molecular mechanism of interaction between polystyrene nanoplastics and catalase by multispectroscopic techniques.

Nanoplastics are emerging pollutants that pose a potential threat to the environment and organisms and are widely distributed in environmental samples and food chains. The accumulation of polystyrene nanoplastics (PS-NPs) in an organism can cause oxidative stress. Currently, toxicity studies of PS-NPs mainly focus on the individual and cellular levels, whereas few studies have been conducted on the molecular mechanisms of the interaction between PS-NPs and catalase (CAT). Based on this, CAT was chosen as the target receptor for molecular toxicity research to reveal the interaction mechanism at the molecular level between PS-NPs and CAT by using various spectroscopic means and enzyme activity detection methods. The results indicated that PS-NPs destroyed the secondary structure of CAT, causing its protein skeleton to loosen and unfold, increasing the content of α-helices, decreasing the content of ß-sheets, and exposing the position of the heme group. After exposure to PS-NPs, the internal fluorophore of CAT underwent fluorescence sensitization, resulting in a micelle-like structure, which enhanced the hydrophobicity of aromatic amino acids but did not change their polarity. In addition, the aggregation state of CAT was altered upon binding to PS-NPs, and the volume was further increased. Finally, these structural changes led to a gradual decrease in CAT activity. This study presents a comprehensive assessment of the toxicity of PS-NPs at the molecular level, which can provide more experimental support for the study of the biotoxicological efficacy of PS-NPs.

Mechanistic insights into pyridine exposure induced toxicity in model Eisenia fetida species: Evidence from whole-animal, cellular, and molecular-based perspectives.

Pyridine and its derivatives are widely used in many applications and inevitably cause extreme scenarios of serious soil contamination, which pose a threat to soil organisms. Still, the eco-toxicological effects and underlying mechanisms of pyridine-caused toxicity toward soil fauna have not been well established. Thus, earthworms (Eisenia fetida), coelomocytes, and oxidative stress-related proteins were selected as targeted receptors to probe the ecotoxicity mechanism of extreme pyridine soil exposure targeted to earthworms by using a combination of in vivo animal experiments, cell-based in vitro tests, in vitro functional and conformational analyses, and in silico analyses. The results showed that pyridine caused severe toxicity to E. fetida at extreme environmental concentrations. Exposure of pyridine induced excessive ROS formation in earthworms, causing oxidative stress and various deleterious effects, including lipid damage, DNA injury, histopathological change, and decreased defense capacity. Also, pyridine destroyed the cell membrane of earthworm coelomic cells and triggered a significant cytotoxicity. Importantly, the intracellular ROS (e.g., O2-, H2O2, and OH·-) was release-activated, which eventually inducing oxidative stress effects (lipid peroxidation, inhibited defense capacity, and genotoxicity) through the ROS-mediated mitochondrial pathway. Moreover, the antioxidant defence mechanisms in coelomocytes responded quickly to reduce ROS-mediated oxidative injury. It was conformed that the abnormal expression of targeted genes associated with oxidative stress in coelomic cells was activated after pyridine exposure. Particularly, we found that the normal conformation (particle sizes, intrinsic fluorescence, and polypeptide backbone structure) of CAT/SOD was destroyed by the direct binding of pyridine. Furthermore, pyridine bound easily to the active center of CAT, but preferentially to the junction cavity of two subunits of SOD, which is considered to be a reason for impaired protein function in cells and in vitro. Based on these evidences, the ecotoxicity mechanisms of pyridine toward soil fauna are elucidated based on multi-level evaluation.

Randomly-shaped nanoplastics induced stronger biotoxicity targeted to earthworm Eisenia fetida species: Differential effects and the underlying mechanisms of realistic and commercial polystyrene nanoplastics.

Nanoplastics (NPs) are widely distributed in various environments, including soil, and have been known to adversely affect soil organisms. Currently, most of the obtained studies were principally focused on the ecological risks of commercial sphere-type microbeads (SNPs), while ignoring that they might be different from randomly-shaped nanoplastics (RNPs) in a real environment. Thus, this study was undertaken to probe the shape-dependent effects of NPs on the earthworm Eisenia fetida and the corresponding poisoning mechanisms, and discriminate the toxicity differences between SNPs and RNPs at the molecule, cell, tissue, and animal levels. The results showed SNPs and RNPs exhibited lethal effects to earthworms with the LC50 determined to be 27.42 g/kg and 21.69 g/kg, respectively after a 28-day exposure. SNPs and RNPs exposure can cause ROS-induced ROS release in worm, inducing oxidative stress through mitochondria-mediated pathway, leading to lipid peroxidation, DNA damage, and histopathological changes, thereby contributing to decreased stress resistance against exogenous stressors. To reduce ROS-mediated oxidative damage, the antioxidant defense system in E. fetida can be activated, which scavenges unwanted ROS. High doses of SNPs and RNPs inhibited the AChE activity in worms, causing excess acetylcholine accumulation in the synaptic space, which finally lead to neurotoxicity. Also, two kinds of NPs can induce the abnormal expression of genes relevant to oxidative stress, reproduction, growth, and tight junction protein in E. fetida, which ultimately contribute to various detrimental effects, tissue damage and dysfunction, reproductive and developmental toxicity. The results obtained from the Integrated Biological Response (IBR) suggested that long-term exposure to high-dose SNPs and RNPs can induce the stronger toxicity effects to E. fetida worms, and RNPs-induced toxicity can be different and stronger than that of SNPs. Our results provide insights for revealing the environmental effects posed by randomly-shaped NPs-contaminated soil, and are of importance for assessing the contribution of NPs with different physical characteristics to soil eco-safety.

Mechanistic insights into phenanthrene-triggered oxidative stress-associated neurotoxicity, genotoxicity, and behavioral disturbances toward the brandling worm (Eisenia fetida) brain: The need for an ecotoxicological evaluation.

In this study, earthworm (Eisenia fetida) brain was chosen as targeted receptors to probe the mechanisms of oxidative stress-related neurotoxicity, genotoxicity, and behavioral disturbances triggered by PHE. Results showed that PHE stress can initiate significant amounts of ROS, thus triggering oxidative stress in E. fetida brain. These effects were accompanied by a significant increase of damage to macromolecules DNA and lipids, resulting in severe oxidative effects. PHE exposure can induce AChE inhibition by ROS-induced injury and the accumulation of excess ACh at the nicotinic post-synaptic membrane, thus inducing aggravated neurological dysfunction and neurotoxicity of E. fetida through an oxidative stress pathway. Moreover, the burrowing behavior of earthworms was disturbed by oxidative stress-induced neurotoxicity after exposure to PHE. Furthermore, the abnormal mRNA expression profiles of oxidative stress- and neurotoxicity-related genes in worm brain were induced by PHE stress. The IBR results suggested that E. fetida brain was suffered more serious damage caused by PHE under higher doses and long-term exposure. Taken together, PHE exposure can trigger oxidative stress-mediated neurotoxicity and genotoxicity in worm brain and behavioral disorder through ROS-induced damage. This study is of great significance to evaluate the harmful effects of PHE and its mechanisms on soil ecological health.

Probing the toxic effect of quinoline to catalase and superoxide dismutase by multispectral method.

Quinoline is a common nitrogen heterocyclic aromatic hydrocarbon with high water solubility. Studies have shown that quinoline can be teratogenic, carcinogenic and mutagenic. And Hepatocytes are the target cell of quinoline, which contain a large number of mitochondria and are related to cell function and the balance of reactive oxygen species (ROS). However, the research on the effect of quinoline on hepatocyte damage and anti-oxidation system is still unclear. Through the means of multispectral experiments, it is concluded that quinoline can affect the catalase (CAT) and superoxide dismutase (SOD), change their structure and affect their activity. The binding mode and binding site of quinoline to CAT/SOD were analyzed by isothermal calorimetric titration (ITC) and Molecular Operating Environment (MOE). In molecular docking simulation, the binding site of quinoline-CAT system is close to the active site, and affect the microenvironment of Tyr 357. This may be the reason why quinoline affects CAT activity and synchronous fluorescence (Δλ = 15 nm). This study demonstrated that quinoline has a great effect on CAT, which may affect the intracellular ROS balance and become a potential way to cause hepatocyte damage.

Ecotoxicological evaluation of oxidative stress-mediated neurotoxic effects, genetic toxicity, behavioral disorders, and the corresponding mechanisms induced by fluorene-contaminated soil targeted to earthworm (Eisenia fetida) brain.

Fluorene is a commonly identified PAH pollutant in soil and exhibits various worrisome hazardous effects to soil organisms. Currently, the toxicity profiles of fluorene on earthworm brain are rare, and the mechanisms and their corresponding pathways involved in fluorene-triggered neurotoxicity, genotoxicity, and behavior changes have not been reported hitherto. Herein, earthworm (Eisenia fetida) brain was chosen as targeted receptor to explore the neurotoxic effects, genetic toxicity, behavioral disorders, and related mechanisms caused by fluorene-induced oxidative stress pathways. The results showed excess fluorene initiated the release of excessive quantities of ROS in earthworm brain, which have caused oxidative stress and accompanied by serious oxidative effects, including LPO (lipid peroxidation) and DNA injury. To minimize the damage effects, the antioxidant defense mechanisms (antioxidant enzymes and non-enzymatic antioxidants) were activated, and entailed a decrease of the antioxidant capacity in E. fetida brain, which, in turn, causes further ROS-induced ROS release. Exposure of fluorene induced the abnormal mRNA expression of genes relevant to oxidative stress (e.g., GST, SOD, CAT, GPx, MT, and Hsp70) and neurotoxicity (e.g., H02, C04, D06, and E08) in E. fetida brain. Specifically, fluorene can bind directly to AChE, destroying the conformation of this protein, and even affecting its physiological functions. This occurrence caused the inhibition of AChE activity and excess ACh accumulation at the nicotinic post-synaptic membrane, finally triggering neurotoxicity by activation of pathways related to oxidative stress. Moreover, the avoidance responses and burrowing behavior were obviously disturbed by oxidative stress-induced neurotoxicity after exposure to fluorene. The results form IBR suggested more severe poisoning effects to E. fetida brain initiated by high-dose and long-term exposure of fluorene. Among, oxidative stress injury and genotoxic potential are more sensitive endpoint than others. Collectively, fluorene stress can provoke potential neurotoxicity, genotoxicity, and behavioral disturbances targeted to E. fetida brain through the ROS-mediated pathways involving oxidative stress. These findings are of great significance to estimate the detrimental effects of fluorene and the corresponding mechanisms on soil eco-safety.

Cytotoxicity and Oxidative Stress Effects of Indene on Coelomocytes of Earthworm (Eisenia foetida): Combined Analysis at Cellular and Molecular Levels.

Indene (IND) is a kind of important aromatic hydrocarbon that is extracted from coal tar and has important applications in industry and biology. In the process of production and utilization, it is easy to enter the soil and produce toxic effects on the soil or organisms. The earthworm is an important organism in the soil. The toxicity of indene on earthworm coelomocytes is rarely studied, and the oxidative stress effects of IND on earthworm coelomocytes remain unclear. In this study, coelomocytes from earthworms and antioxidant enzymes were selected as the research targets. In addition, IND caused oxidative stress, and its related toxic effects and mechanisms were systematically studied and evaluated at the cellular and molecular levels. The results showed that IND destroyed the redox balance in earthworm coelomocytes, and the large accumulation of reactive oxygen species (ROS) significantly inhibited the activities of the antioxidant system, including superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH), and caused lipid peroxidation and membrane permeability changes, resulting in a decrease in cell viability to 74.5% of the control group. At the molecular level, IND was bound to SOD by the arene-H bond, and the binding constant was 4.95 × 103. IND changed the secondary structure of the SOD and led to a loosening of the structure of the SOD peptide chain. Meanwhile, IND caused SOD fluorescence sensitization, and molecular simulation showed that IND was mainly bound to the junction of SOD subunits. We hypothesized that the changes in SOD structure led to the increase in SOD activity. This research can provide a scientific basis for IND toxicity evaluation.