Aryl phosphorus-containing flame retardants (aryl-PFRs) have been frequently detected with increasingly used worldwide as one of alternatives for brominated flame retardants. However, information on ...their adverse effects on human health and ecosystem is insufficient, with limited study on their molecular mode of action in vitro. In this study, the cytotoxicity, DNA damage, mitochondrial impairment and the involved molecular mechanisms of certain frequently detectable aryl-PFRs, including 2-ethylhexyldiphenyl phosphate (EHDPP), methyl diphenyl phosphate (MDPP), bisphenol-A bis (diphenyl phosphate) (BDP), isodecyl diphenyl phosphate (IDPP), cresyl diphenyl phosphate (CDP) and the structurally similar and widely used organophosphorus pesticide chlorpyrifos (CPF), were evaluated in A549 cells using high-content screening (HCS) system. Aryl-PFRs showed different lethal concentration 50 (LC50) values ranging from 97.94 to 546.85 μM in A549 cells using CCK-8 assay. EHDPP, IDPP, CDP, MDPP and CPF demonstrated an ability to induce DNA damage, evidenced by increased DNA content and S phase-reducing cell cycle arrest effect using fluorophore dye cocktail assay. Additionally, the selected aryl-PFRs induced mitochondrial impairment by the increasing mitochondrial mass and decreasing mitochondrial membrane potential. Moreover, BDP, MDPP, and CDP, which contain short alkyl chains showed their potential oxidative stress with intracellular ROS and mitochondrial superoxide overproduction from an initially relatively low concentration. Additionally, based on the promotion of firefly luminescence in p53-transfected A549 cells, p53 activation was found to be involved in aryl-PFRs-induced DNA damage. Further real-time PCR results showed that all selected aryl-PFRs triggered p53/p21/gadd45β-, and p53/p21/mdm2-mediated cell cycle pathways, and the p53/bax mediated apoptosis pathway to induce DNA damage and cytotoxic effects. These results suggest that aryl-PFRs (e.g., BDP, MDPP, CDP) cause oxidative stress-mediated DNA damage and mitochondrial impairment, and p53-dependent pathway was involved in the aryl-PFRs-induced DNA damage and cell cycle arrest. In conclusion, this study improves the understanding of PFRs-induced adverse outcomes and the involved molecular mechanism.
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•The cytotoxicity, DNA damage, and mitochondrial impairment induced by aryl-PFRs were evaluated.•Aryl-PFRs induced oxidative stress-mediated DNA and mitochondrial damage.•EHDP, IDPP, CDP, MDPP and CPF exerted S phase cell ratio-reducing effects.•p53 pathway activation is an underlying mechanism of the induced DNA damage.•Cytotoxicity was evidenced by the upregulation of the p53/bax-mediated apoptosis pathway.
Organophosphorus flame retardants (OPFRs) have been frequently detected with relatively high concentrations in various environmental media and are considered emerging environmental pollutants. ...However, their biological effect and underlying mechanism is still unclear, and whether chlorinated OPFRs (Cl-OPFRs) cause adverse outcomes with the same molecular initial events or share the same key events (KEs) remains unknown. In this study, in vitro bioassays were conducted to analyze the cytotoxicity, mitochondrial impairment, DNA damage and molecular mechanisms of two Cl-OPFRs. The results showed that these two Cl-OPFRs, which have similar structures, induced severe cellular and molecular damages via different underlying mechanisms. Both tris(2-chloroethyl) phosphate (TCEP) and tris(1-chloro-2-propyl) (TCPP) induced oxidative stress-mediated mitochondrial impairment and DNA damage, as shown by the overproduction of intracellular reactive oxygen species (ROS) and mitochondrial superoxide. Furthermore, the DNA damage caused by TCPP resulted in p53/p21-mediated cell cycle arrest, as evidenced by flow cytometry and real-time PCR. At the cellular and molecular levels, TCPP increased the sub-G1 apoptotic peak and upregulated the p53/Bax apoptosis pathway, possibly resulted in apoptosis associated with its stronger cytotoxicity. Although structurally similar to TCPP, TCEP did not induce mitochondrial impairment and DNA damage by the same KEs. These results provide insight into the toxicity of Cl-OPFRs with similar structures but different mechanisms, which is of great significance for constructing adverse outcome pathways or determining intermediate KEs.
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•TCEP was less cytotoxic than TCPP in A549 cells.•Both TCEP and TCPP induced oxidative stress and mitochondrial damage.•TCPP resulted in S phase cell decreased, which was greater than that of TCEP.•TCPP induces intact p53/p21/Gadd45β-mediated cell cycle-related signaling pathways.•TCPP activated the p53/Bax-mediated apoptosis pathway with stronger toxicity.
The environmental risks of environmental estrogens (EEs) are often assessed via the same mode of action in the concentration addition (CA) model, neglecting the complex combined mechanisms at the ...genetic level. In this study, the cell proliferation effects of estrone, 17α-ethinylestradiol, 17β-estradiol, estriol, diethylstilbestrol, estradiol valerate, bisphenol A, 4-tert-octylphenol and 4-nonylphenol were determined individually using the CCK-8 method, and the proliferation effects of a multicomponent mixture of estrogenic chemicals mixed at equipotent concentrations using a fixed-ratio design were studied using estrogen-sensitive MCF-7 cells. Furthermore, transcription factors related to cell proliferation were analyzed using RT-PCR assays to explore the potential molecular mechanisms related to the estrogenic proliferative effects. The results showed that the estrogenic chemicals act together in an additive mode, and the combined proliferative effects could be predicted more accurately by the response addition model than the CA model with regard to their adverse outcomes. Furthermore, different signaling pathways were involved depending on the different mixtures. The RT-PCR analyses showed that different estrogens have distinct avidities and preferences for different estrogen receptors at the gene level. Furthermore, the results indicated that estrogenic mixtures increased ERα, PIK3CA, GPER, and PTEN levels and reduced Akt1 level to display combined estrogenicity. These findings indicated that the potential combined environmental risks were greater than those found in some specific assessment procedures based on a similar mode of action due to the diversity of environmental pollutions and their multiple unknown modes of action. Thus, more efforts are needed for mode-of-action-driven analyses at the molecular level. Furthermore, to more accurately predict and assess the individual responses in vivo from the cellular effects in vitro, more parameters and correction factors should be taken into consideration in the addition model.
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•Dose-response concentration of single and combined effects of nine selected environmental estrogens (EEs) were evaluated.•RA model were more suitable than CA model for predicting the environmental risks of EEs at environmentally relevant concentrations.•Different estrogens have distinct avidities and preferences for different estrogen receptors.•EEs showed combined effects by increasing PIK3CA, GPER, and PTEN levels and reducing Akt1 level in the PI3K/AKT pathway.
An increasing number of inorganic ultraviolet filters (UVFs), such as nanosized zinc oxide (nZnO) and titanium dioxide (nTiO
), are formulated in sunscreens because of their broad UV spectrum ...sunlight protection and because they limit skin damage. However, sunscreen-derived inorganic UVFs are considered to be emerging contaminants; in particular, nZnO and nTiO
UVFs have been shown to undergo absorption and bioaccumulation, release metal ions, and generate reactive oxygen species, which cause negative effects on aquatic organisms. We comprehensively reviewed the current study status of the environmental sources, occurrences, behaviors, and impacts of sunscreen-derived inorganic UVFs in aquatic environments. We find that the associated primary nanoparticle characteristics and coating materials significantly affect the environmental behavior and fate of inorganic UVFs. The consequential ecotoxicological risks and underlying mechanisms are discussed at the individual and trophic transfer levels. Due to their persistence and bioaccumulation, more attention and efforts should be redirected to investigating the sources, fate, and trophic transfer of inorganic UVFs in ecosystems.
Surfactants such as alkylphenol polyethoxylates (APEOs) and nonylphenol ethoxylates (NPEOs) are commonly used worldwide, but the majority of these compounds, together with their metabolites, have ...been reported to induce severe biological toxicity. Here, we evaluated for the first time the cytotoxicity, genotoxicity and mitochondrial damage in Chinese hamster ovary (CHO-K1) cells caused by a novel non-ionic surfactant, vanillin ethoxylates (VAEOs), an alternative to APEOs. In parallel, the same in vitro bioassays were conducted on NPEOs along with their metabolic byproducts 4-nonylphenol (4-NP) and vanillin. The results showed that the cytotoxic potency order was NPEOs > 4-NP > VAEOs>vanillin using CCK-8 assays. Also, 4-NP showed potential direct DNA damage in SOS/umu tests, whereas NPEOs, VAEOs and vanillin showed no positive result with and without S9 addition. In addition, none of the test compounds showed obvious genotoxic effects with low olive tail moment value using comet assays. However, all test compounds were shown to cause mitochondrial impairment by increasing mitochondrial mass and decreasing mitochondrial membrane potential in a concentration-dependent manner. And further analysis of reactive oxygen species (ROS) and mitochondrial superoxide (MNSOD) measurement showed that mitochondrial impairment was induced by oxidative stress with intracellular ROS and MNSOD overproduction. It's worth noting that VAEOs and vanillin cause relative lower cytotoxic, genotoxic and mitochondrial damage effects than NPEOs and 4-NP, indicating that VAEOs have the potential to substitute NPEOs as suitable surfactants. Take together, this study elucidates the toxicity profiles of VAEOs and NPEOs relatively comprehensively, and further toxicity analyses are suggested in the population, community and ecosystem.
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Lake Baiyangdian is one of China's largest macrophyte - derived lakes, facing severe challenges related to water quality maintenance and eutrophication prevention. Dissolved organic matter (DOM) was ...a huge carbon pool and its abundance, property, and transformation played important roles in the biogeochemical cycle and energy flow in lake ecosystems. In this study, Lake Baiyangdian was divided into four distinct areas: Unartificial Area (UA), Village Area (VA), Tourism Area (TA), and Breeding Area (BA). We examined the diversity of DOM properties and sources across these functional areas. Our findings reveal that DOM in this lake is predominantly composed of protein - like substances, as determined by excitation - emission matrix and parallel factor analysis (EEM - PARAFAC). Notably, the exogenous tyrosine-like component C1 showed a stronger presence in VA and BA compared to UA and TA. Ultrahigh - resolution mass spectrometry (FT - ICR MS) unveiled a similar DOM molecular composition pattern across different functional areas due to the high relative abundances of lignan compounds, suggesting that macrophytes significantly influence the material structure of DOM. DOM properties exhibited specific associations with water quality indicators in various functional areas, as indicated by the Mantel test. The connections between DOM properties and NO3N and NH3N were more pronounced in VA and BA than in UA and TA. Our results underscore the viability of using DOM as an indicator for more precise and scientific water quality management.
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Limited toxic and ecological studies were focused on physical sunscreen that is considered to have “safer performance”, in which nanosize zinc oxide (nZnO) and nanosize titanium dioxide (nTiO2) ...generally are added as ultraviolet filters. Herein, the common button coral Zoanthus sp. was newly used to assess the toxic effects and underlying mechanisms of physical sunscreen. Results showed that physical sunscreen induced severe growth inhibition effects and largely compelled the symbiotic zooxanthellae, indicating that their symbiotic systems were threatened and, also, that neural and photosynthesis functions were influenced. Zn2+ toxicity and bioaccumulation were identified as the main toxic mechanisms, and nTiO2 particles released from physical sunscreen also displayed limited bioattachment and toxicity. Oxidative stress, determined by increased reactive oxygen species, superoxide dismutase, and malondialdehyde content, was indicated as another important toxic mechanism. Furthermore, when Zoanthus sp. was restored, the inhibited individual coral could be largely recovered after a short (3 d) exposure time; however, a longer exposure time damaged the coral irretrievably, which revealed the latent environmental risks of physical sunscreen. This study investigated the toxic effect of physical sunscreen on Zoanthus sp. in a relatively comprehensive manner, thus providing new insights into the toxic response of sunscreen on marine organisms.
Phosphorus-containing flame retardants (PFRs) have been frequently detected in various environmental samples at relatively high concentrations and are considered emerging environmental pollutants. ...However, their biological effects and the underlying mechanism remain unclear, especially alkyl-PFRs. In this study, a battery of in vitro bioassays was conducted to analyze the cytotoxicity, oxidative stress, mitochondrial impairment, DNA damage and the involved molecular mechanisms of several selected alkyl-PFRs. Results showed that alkyl-PFRs induced structural related toxicity, where alkyl-PFRs with higher logKow values induced higher cytotoxicity. Long-chain alkyl-PFRs caused mitochondrial and DNA damage, resulting from intracellular reactive oxygen species (ROS) and mitochondrial superoxide overproduction; while short-chain alkyl-PFRs displayed adverse outcomes by significantly impairing mitochondria without obvious ROS generation. In addition, alkyl-PFRs caused DNA damage-induced cell cycle arrest, as determined by flow cytometry, and transcriptionally upregulated key transcription factors in p53/p21-mediated cell cycle pathways. Moreover, compared to the control condition, triisobutyl phosphate and trimethyl phosphate exposure increased the sub-G1 apoptotic peak and upregulated the p53/bax apoptosis pathway, indicating potential cell apoptosis at the cellular and molecular levels. These results provide insight into PFR toxicity and the involved mode of action and indicate the mitochondria is an important target for some alkyl-PFRs.
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•Alkyl-PFRs showed dose- and logKow value-dependent cytotoxicity on A549 cells.•Long-chain alkyl-PFRs caused more severe oxidative stress and mitochondrial damage.•Alkyl-PFRs caused DNA damage-induced G1 phase arrest via ROS generation.•p53/p21-mediated cell cycle pathway was activated to regulate the DNA damage.
Alkylated organophosphate esters (alkyl-OPEs) are widely used and extensively detected in aquatic organisms. This work investigated the tissue-specific toxicokinetics of two common alkyl-OPEs, ...tri(2‑butoxyethyl) phosphate (TBOEP) and tri‑n‑butyl phosphate (TNBP) in Chinese rare minnow (Gobiocypris rarus) through a 50 day uptake and depuration experiment. The tissue-specific bioconcentration factor (BCF) values for the two alkyl-OPEs ranged from 1 to 30 L/kg wet weight (ww), with the kidney and ovary as the tissues with the highest accumulation. The tissue BCFs only exhibited a significant correlation with lipid contents only in storage tissues (i.e., muscle, brain, ovary and testis), indicating that lipids might not be the major contributor to tissue distribution of TBOEP and TNBP. However, the contribution of blood perfusion and active transport to tissue-specific OPE accumulation needs to be further investigated. Lower accumulation of metabolites than parent chemicals was observed, with metabolite parent concentration factors (MPCFs) <1. Di-alkyl phosphate (DAP), bis(2‑butoxyethyl) phosphate (BBOEP) and di(n-butyl) phosphate (DNBP) were the most abundantly formed metabolites of TBOEP and TNBP in various tissues, followed by the monohydroxylated OPEs (OH-OPEs). However, bis(2‑butoxyethyl) hydroxyethyl phosphate (BBOEHEP), was detected at much lower levels in the tissues. All the investigated metabolites showed high production rates (kprod,metabolites) in the fish liver, followed by the GI tract and the kidney, indicating the importance of the hepatobiliary and urinary systems in eliminating the metabolites. Our study suggested that metabolism plays an important role in eliminating these two alkyl-OPEs in rare minnow and results in different tissue distribution mechanisms for metabolites and their compounds.
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•TBOEP and TNBP highly accumulated most favorably in Kidney, followed by the ovary and liver.•Lipid content and tissue BCFs were significantly correlated only in storage tissues.•DAPs and OH-OPEs were both major metabolites, but showed low MPCFs.•Metabolites mainly concentrated in liver, followed by GI tract and kidney.•Metabolism plays a major role in overall elimination of TBOEP and TNBP in rare minnow
Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) and tris(2-butoxyethyl) phosphate (TBOEP), two of the most widely applied phosphorus-containing flame retardants (PFRs), have been shown to have adverse ...effects in different organisms, but their underlying toxicity mechanism remains to be elucidated. In the present study, the toxicological effects of TDCPP and TBOEP on human lung carcinoma (A549) cells and their associated mechanisms were investigated at the cellular, organelle and transcriptional levels. Cell Counting Kit-8 assay results showed that TDCPP and TBOEP reduced cell viability in a concentration-dependent manner. High-content screening assays showed that both PFRs caused mitochondrial impairment at the organelle level, decreasing the mitochondrial membrane potential and increasing the mitochondrial mass, and leading to increased cell cycle arrest-associated DNA damage that decreased the ratio of cells in S phase and increased the DNA content. In addition, oxidative stress was identified as a possible initial mechanistic process underlying PFR-induced cytotoxicity and mitochondrial impairment, as evidenced by intracellular reactive oxygen species and mitochondrial superoxide overproduction from environmentally relevant concentrations. Furthermore, p53 activation was shown to be involved in regulating DNA damage after A549 cell exposure to TDCPP and TBOEP. Quantitative PCR analyses showed that TDCPP and TBOEP triggered p53/p21-mediated cell cycle arrest pathways, indicating that these PFRs can activate p53, and promote associated functional repair. However, the p53/bax-mediated apoptosis pathway was not activated following exposure to PFRs at specific concentrations. Our results showed that TDCPP and TBOEP can cause cytotoxicity, mitochondrial impairment and cell cycle arrest-associated DNA damage in A549 cells, suggesting that oxidative stress is the initial molecular event by which these adverse outcomes are induced.
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•TDCPP and TBOEP caused cellular, mitochondrial and DNA damage in A549 cells.•Oxidative stress was found to be an underlying mechanism after exposure.•The p53/p21-mediated cell cycle pathway was activated to repair the damage.•Neither PFRs induced apoptosis at environmently relevant concentrations.
