Non-lethal Concentrations Research Articles

Tebuconazole induced oxidative stress related hepatotoxicity in adult and larval zebrafish (Danio rerio).

Tebuconazole is widely used as fungicide and has frequently been detected at elevated concentrations in environmental media. To characterize the potential toxicity of tebuconazole on vertebrate and humans. Using zebrafish as a vertebrate model, the toxic effects in liver that produced by low-toxic concentrations of tebuconazole were assessed in adult zebrafish. We further focused on tebuconazole-induced toxicity and its possible mechanism in larval zebrafish using a hepatotoxicity assay. The induction of oxidative stress in adult fish was evaluated by superoxide dismutase (T-SOD), catalase (CAT), peroxidase (POD), glutathione S-transferase (GST) activity, and the increased aspartate aminotransferase (AST)/alanine aminotransferase (ALT) ratio. Significantly increased enzyme activity was observed in the liver of male and female fish at both exposure and depuration stage. Exposure to maximum non-lethal (MNLC) concentration of tebuconazole from 72 to 120 h post-fertilization (hpf) affected the liver size and yolk retention in larval zebrafish. Decreased fluorescence intensity was observed in larval Tg(Apo14:GFP) zebrafish, indicating liver degeneration after tebuconazole treated. Histopathological examination confirmed the alterations in liver histoarchitecture in exposed zebrafish. Significant 1.28-fold and 1.65-fold increases in reactive oxygen species levels were observed in juveniles exposed to MNLC and lethal concentration 10 (LC10) group, respectively. The acridine orange staining assay showed that apoptotic cells occurred in the liver regions. These results indicated that tebuconazole exposure resulted in impacts on the ecological risk in fish and vertebrate. Overall, the present study suggested further research in needed to better understand the tebuconazole-induced toxicity mechanism that associated with oxidative stress.

Evaluation of the sensitivity of Microhyla fissipes tadpoles to aqueous cadmium.

Cadmium (Cd) exposure is harmful to amphibians in natural environments and the Cd concentration is a key parameter in water monitoring. Cd pollution has been a severe issue in the Yangtze River and its southern reaches in recent years. Acute toxicity assays were employed to determine the tolerance limits of Cd for Microhyla fissipes tadpoles and five different concentrations of Cd (0, 50, 100, 200 and 300 μg/L) were involved to detect its chronic effects on metamorphosis, growth, locomotion, genotoxicity and enzymatic activities of M. fissipes tadpoles. The results showed that the 24-h and 48-h LC50 values of Cd on M. fissipes tadpoles were 2591.3 μg/L and 1567.9 μg/L, respectively, and the presumable non-lethal concentration obtained was 172.2 μg/L. During the 70-day chronic toxicity assays, Cd showed negative impacts on survival, growth, metamorphosis and the frequency of erythrocytes nuclear abnormality of M. fissipes tadpoles. However, the Cd exposure caused the increased body size and condition of tadpoles at complete metamorphosis (GS46). The tadpoles exposed to 200 μg/L of Cd exhibited degraded locomotor performance at GS46. Weight increments of tadpoles were inhibited at Day 14 and massive deaths were observed over the next 14 days. The enzymatic activities of tadpoles experienced a shock response stage (GS30-GS35) and a complete recovery stage (GS36-GS41) in all treatments. However, the enzymatic activities (except alkaline phosphatase) of tadpoles at GS46 increased after Cd exposure, especially at high concentrations. In summary, Cd is a threat to M. fissipes tadpoles as that causes reduced fitness.

Ultralong AgNWs-induced toxicity in A549 cells and the important roles of ROS and autophagy

Safety concerns have been raised with regard to silver nanowires (AgNWs) because of their extensive applications. Recently, ultralong AgNWs have shown physical properties superior to those of short AgNWs. However, little is known about their toxicity and potential risks. In this study, we demonstrated a series of ultralong AgNWs-induced biological effects in human lung cancer epithelial cells (A549). Ultralong AgNWs treatments induced ROS generation, mitochondria-mediated apoptosis, and self-protective autophagy at nonlethal concentrations. In contrast to some previous reports, apoptosis was found not to correlate with the reduction of intracellular ROS. Measuring the processing of ROS generation, apoptosis and autophagy, we demonstrated that ROS not only enhance mitochondrial damage, but also raise protective autophagic flux in ultralong AgNW-treated cells. Moreover, ultralong AgNWs were found to be internalized into the cytoplasm of the epithelial cells. This study not only investigates ultralong AgNWs-induced cytotoxicity but also pinpoints ROS as a key signal in mechanisms of their toxicity.

Doxorubicin induces trans-differentiation and MMP1 expression in cardiac fibroblasts via cell death-independent pathways.

Although doxorubicin (DOX)-induced cardiomyopathy causes lethal heart failure (HF), no early detection or effective treatment methods are available. The principal mechanisms of cardiotoxicity are considered to involve oxidative stress and apoptosis of cardiomyocytes. However, the effect of DOX on cardiac fibroblasts at non-lethal concentrations remains unknown. The aim of this study was to investigate the direct effect of doxorubicin on the activation of cardiac fibroblasts independent of cell death pathways. We first found that DOX induced α-SMA expression (marker of trans-differentiation) at a low concentration range, which did not inhibit cell viability. DOX also increased MMP1, IL-6, TGF-β and collagen expression in human cardiac fibroblasts (HCFs). In addition, DOX promoted Akt and Smad phosphorylation. A Smad inhibitor prevented DOX-induced α-SMA and IL-6 protein expression. An PI3K inhibitor also prevented MMP1 mRNA expression in HCFs. These findings suggest that DOX directly induces fibrotic changes in HCFs via cell death-independent pathways. Furthermore, we confirmed that these responses are organ- and species-specific for HCFs based on experiments using different types of human and murine fibroblast cell lines. These results suggest potentially new mechanisms of DOX-induced cardiotoxicity from the viewpoint of fibrotic changes in cardiac fibroblasts.

Effect of fingolimod on oligodendrocyte maturation under prolonged cerebral hypoperfusion

Oligodendrocytes (OLGs) support neuronal system and have crucial roles for brain homeostasis. As the renewal and regeneration of OLGs derived from oligodendrocyte precursor cells (OPCs) are inhibited by various pathological conditions, the restoration of impaired oligodendrogenesis is a therapeutic strategy for OLG-related diseases such as subcortical ischemic vascular dementia (SIVD). Fingolimod (FTY720), a drug for multiple sclerosis, is reported to elicit a cytoprotective effect on OPCs in vitro. However, the effects of fingolimod against ischemia-induced suppression of OPC differentiation remain unknown. Hence, the purpose of this study was to investigate the effectiveness of fingolimod against ischemia-induced suppression of oligodendrogenesis. For the in vitro experiments, primary rat cultured OPCs were incubated with a non-lethal concentration of CoCl2 to induce chemical hypoxic conditions and were treated with or without fingolimod-phosphate. We found that low concentration fingolimod-phosphate directly rescued ischemia-induced suppression of OPC differentiation via the phosphoinositide 3-kinase-Akt pathway. For the in vivo experiments, we used a mouse model of SIVD generated by bilateral common carotid artery stenosis. On day 28 after surgery, fingolimod ameliorated ischemia-induced demyelination and promoted oligodendrogenesis under prolonged cerebral hypoperfusion. The present study demonstrates that fingolimod can promote oligodendrogenesis under ischemic conditions and may be a therapeutic candidate for SIVD.

Bioaccumulation, biotransformation and toxic effect of fipronil in Escherichia coli

Fipronil is a highly effective, broad-spectrum insecticide used to control pests, globally. The increased usage has led to contamination of soil, water, fruits, and vegetables. The wide and frequent usage of fipronil across the globe calls for attention regarding risk assessment of undesirable effects on non-target microorganisms. In this context, the present study was carried to understand the impact of fipronil on non-pathogenic Escherichia coli. The non-pathogenic E. coli are important commensal of the intestinal tract of humans and animals and are also indicator organisms in the environment. Our study indicates that exposure of E. coli to fipronil (100 μM concentration) leads to significant reactive oxygen species production, loss of membrane potential and viability. Further, we have witnessed the bioaccumulation and biotransformation of fipronil by E. coli at non-lethal concentrations. The bio-transformed products (fipronil sulfone and fipronil sulfide) are also the major metabolites (along with amide) reported in the feces of the mammals when exposed to fipronil. Thus, there is a possibility that the gut E. coli might play a role in the degradation and thereby removal of fipronil. In addition, the bioaccumulation of fipronil in bacteria is of concern and need to be further explored because it can lead to biomagnification in the higher trophic level and can disturb the ecological balance. In our knowledge, this is the first report on the determination of fipronil and its metabolites in bacteria through GC-MS/MS.

Growth, Physiological and Biochemical Responses of two Greek Cotton Cultivars to Salt Stress and their Impact as Selection Indices for Salt Tolerance

Soil salinity is a major constrain of crop productivity. Upland cotton (Gossypium hirsutum L.) is an important fiber crop worldwide and a major agricultural product in Greece. Two commercial cotton cultivars (‘Hersi’ and ‘ST 318’) were studied to compare their response under non-saline and saline conditions in a greenhouse experiment. Salt stress on plants was imposed by two different approaches: a gradual and an initial acclimatization to a non-lethal NaCl concentration (150 mM). To explore salt stress responses, growth (height of plants, roots, shoots and leaves dry weight, reproductive shoots, Salinity Sensitivity Index), gas exchange (Photosynthetic rate, Stomatal conductance, Transpiration rate and Water Use Efficiency) and biochemical parameters (proline, H2O2 and MDA content), were examined as well as ion homeostasis. ‘Hersi’ had significantly higher dry weight of roots, shoots and leaves, lower salinity sensitivity index of roots compared to ‘ST 318’. In this regard, it appears that ‘Hersi’ cultivar performed better than ‘ST 318’ to increased salinity conditions, due to better control of gas exchange parameters and K+/Na+ homeostasis as well as better membrane integrity. Furthermore, the gradual acclimatization to the 150 mM NaCl concentration had a milder effect on both cultivars compared to the initial acclimatization.
 
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 In press - Online First. Article has been peer reviewed, accepted for publication and published online without pagination. It will receive pagination when the issue will be ready for publishing as a complete number (Volume 47, Issue 3, 2019). The article is searchable and citable by Digital Object Identifier (DOI). DOI link will become active after the article will be included in the complete issue.
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Integrating multi-omics and regular analyses identifies the molecular responses of zebrafish brains to graphene oxide: Perspectives in environmental criteria

With the broad application of nanoparticles, nanotoxicology has attracted substantial attention in environmental science. However, the methods for detecting few and targeted genes or proteins, even single omics approaches, may miss other responses, including the major responses induced by nanoparticles. To determine the actual toxicological mechanisms of zebrafish brains induced by graphene oxide (GO, a popular carbon-based nanomaterial applied in various fields) at nonlethal concentrations, multi-omics and regular analyses were combined. The biomolecule responses were remarkable, although GO was not obviously observed in brain tissues. The trends for gene and protein changes were the same and accounted for 3.53% and 5.36% of all changes in the genome and proteome, respectively, suggesting a limitation of single omics analysis. Transcriptomics and proteomics analyses indicated that GO affected the functions or pathways of the troponin complex, actin cytoskeleton, monosaccharide transmembrane transporter activity, oxidoreductase activity and focal adhesion. Both metabolomics and proteomics revealed mitochondrial dysfunction and disruption of the citric acid cycle. The integrated analysis of omics, transmission electron microscopy and immunostaining confirmed that GO induced energy disruptions and mitochondrial damage by downregulating tubulin. The combination of multi-omics and regular analyses provides insights into the actual and highly influential mechanisms underlying nanotoxicity.

Profiling of VOCs released from different salivary bacteria treated with non-lethal concentrations of silver nitrate

Considering the shortcomings related to antibiotics usage, the introduction of other bacteriostatic and bactericidal agents that present synergetic effects or standalone properties is urgently needed. AgNO3 is an important bactericidal agent, which imparts various functions on bacteria dependent on its concentration. Therefore, an understanding of its mechanisms of action in infinitesimal concentrations plays an important role which can ultimately lead to AgNO3 involvement in the pharmaceutical industry. The monitoring of VOC (volatile organic compound) profiles emitted by bacteria is a simple method to assess changes occurring in bacterial metabolism. In this study, VOCs of Hafnia alvei, Pseudomonas luteola and Staphylococcus warneri cultures were analyzed both in the absence and in the presence of three concentrations of AgNO3. Headspace solid-phase microextraction gas chromatography/mass spectrometry (HS-SPME-GC/MS) was employed for extraction and analysis. After supplementation with AgNO3, changes in the emitted fingerprints were investigated. Odorants associated with mouth-related and systemic diseases, like dimethyl trisulfide, indole (halitosis) and 2-hexanone (celiac disease), were also affected by addition of AgNO3. Statistical tests proved discrimination between obtained profiles with more that 90% variability. Moreover, physiological states of bacteria after dosage with various concentration of stressing agent were investigated and explained by the mechanisms of action.

Rapamycin reduces mortality in acute-stage paraquat-induced toxicity in zebrafish

Paraquat (PQ) intoxication is frequently associated with a high mortality rate. No specific treatment has been shown to reduce mortality in victims within the first 72 hours. We investigated the protective effects of rapamycin (Rapa) against PQ-induced toxicity in a zebrafish model. To determine the maximum nonlethal concentration (MNLC) and lethal concentration 50 (LC50) of Rapa, zebrafish were treated at 2-5 days post fertilisation (dpf) and their mortality was recorded every 24 hours. At 5 dpf, the zebrafish were treated with PQ 100 µg/mL or PQ+Rapa (MNLC, 1/3 MNLC or 1/9 MNLC) for 72 hours, and the rate of survival was recorded every 24 hours. Reverse transcription-polymerase chain reaction was used to test the signalling pathway of mTOR (mammalian target of rapamycin). MNLC and LC50 of Rapa were determined to be 6.7 µg/mL and 28.9 µg/mL, respectively. At 48 hours, the PQ+Rapa groups had much lower mortality than the PQ group. The rates of survival of the PQ+Rapa groups were 43.33% (MNLC), 53.89% (1/3 MNLC) and 44.45% (1/9 MLNC), as compared to 19.45% in the PQ group, with the 1/3 MNLC group showing the highest rate of survival (p < 0.001). atg1 was slightly activated in the PQ group. In the PQ+Rapa groups, the expression of atg1 was markedly increased, suggesting strengthening of the autophagy process. Rapa can increase the rate of survival of PQ-intoxicated zebrafish by inhibiting mTOR complex 1 and activating autophagy. Rapa could be an alternative first-line drug in the treatment of PQ poisoning.

Nontoxic concentrations of OTA aggravate DON-induced intestinal barrier dysfunction in IPEC-J2 cells via activation of NF-κB signaling pathway

Deoxynivalenol (DON) is well-known enteropathogenic mycotoxin which can alter intestinal barrier functions. Consistently, Ochratoxin A (OTA) ingestion has been found to induce intestinal injuries, including inflammation and diarrhea. However, little is known whether OTA aggravates DON-induced toxicity. This study is designed to explore the effects of OTA on DON-induced intestinal barrier function and involved mechanism. Our results showed either DON or OTA could disrupt intestinal barrier function in a time- and dose-dependent manner, as demonstrated by decreased transepithelial electrical resistance (TEER) and increased paracellular permeability to 4 kDa dextran. However, to eliminate the involvement of cell death, nonlethal concentrations of DON and OTA were used in following experiments. The nontoxic concentration of OTA was observed to aggravate DON-induced intestinal barrier dysfunction, accompanied with tight junction disruption (Claudin-3 and Claudin-4). Moreover, nontoxic concentrations of OTA aggravated DON-induced up-regulation of pro-inflammatory cytokines expression and activated nuclear factor-κB (NF-κB) in IPEC-J2 cells. Adding NF-κB inhibitor (PDTC) alleviated the aggravating effects of nontoxic concentrations of OTA on DON-induced intestinal barrier dysfunction and inflammation. These findings indicate that nontoxic concentrations of OTA promoted DON-induced barrier dysfunction via NF-κB signaling pathway. Our experiment suggests that exposure to nontoxic concentrations of toxins also poses potentially harmful effects.

An Agricultural Detergent as Co-Adjuvant for Entomopathogenic Fungi and Chlorpyrifos to Control Pseudococcus viburni (Hemiptera: Pseudococcidae)

The agricultural detergent TS-2035® was evaluated using a Potter tower in the laboratory to expose Pseudococcus viburni (Signoret) (Hemiptera: Pseudococcidae) females to a non-lethal concentration (0.001% v/v) as a co-adjuvant of formulations containing the entomopathogenic fungi Beauveria bassiana Vuillemin (Cordycipitaceae) or Metarhizium anisopliae Sorokin (Clavicipitaceae), or the organophosphate insecticide chlorpyrifos. At that concentration, TS-2035 did not significantly remove the epicuticle wax of the pseudococcids, nor affect the pH of the solution. Adding the detergent to the M. anisopliae and chlorpyrifos formulations significantly reduced the LC50 of those solutions at 24, 72, and 144 h post treatment. For B. bassiana, the detergent significantly decreased the LC50 of that product only at 72 h post treatment. Fungal solutions with detergent did not affect the conidial germination of the entomopathogenic fungi. Our results found that mixing B. bassiana, M. anisopliae, or chlorpyrifos formulations with TS-2035 at 0.001% v/v did not control P. viburni by removal of their epicuticular wax, but still contributed significantly to mortality.

The Effects of Astrocyte and Oligodendrocyte Lineage Cell Interaction on White Matter Injury under Chronic Cerebral Hypoperfusion

Oligodendrocytes (OLGs) differentiate from oligodendrocyte-precursor-cells (OPCs) for myelination in white matter. This differentiation is maintained by cell–cell interactions through trophic factors such as brain-derived-neurotrophic-factor (BDNF). However, differentiation is impaired when white matter injury occurs in a chronic cerebral hypoperfusion model. Thus, we examined the effects of the interaction between astrocyte and oligodendrocyte lineage cells on myelination regarding the mechanism of impairment. A microcoil was applied to the bilateral common carotid arteries in male C57BL/6 mice as an in vivo cerebral chronic hypoperfusion model (BCAS model). A nonlethal concentration of CoCl2 was added to the primary cell culture from the postnatal rat cortex and incubated in vitro. White matter injury progressed in the BCAS model as myelin decreased. The numbers of OPCs and astrocytes increased after the operation, whereas that of OLGs decreased at day 28. BDNF continuously decreased until day 28. Differentiation was disrupted under the stressed conditions in the cell culture, but improved after administration of astrocyte-conditioned medium containing BDNF. Astrocytes with BDNF underwent differentiation, but differentiation was impaired under the stressed conditions due to the reduction of BDNF. We examined S100B regarding the mechanism of impairment. S100B is mainly expressed by mature astrocytes, and has neuroprotective and neurotoxic effects inside and outside of cells. GFAP-positive astrocytes increased in the corpus callosum in the BCAS model, whereas the number of mature astrocytes continued to decrease, resulting in reduced BDNF. The reduction in mature astrocytes due to the discharge of S100B in ischemic conditions caused the reduction in BDNF.

Enantioselective mixture toxicity of the azole fungicide imazalil with the insecticide α-cypermethrin in Chironomus riparius: Investigating the importance of toxicokinetics and enzyme interactions

The fungicide imazalil is a chiral compound with one R- and one S-enantiomer. Enantiomers, while having the same chemical properties, can differ in their biological activity expressed as efficacy/toxicity as well as in their degradation kinetics and pathways. Azoles such as imazalil have been shown to synergize the effect of pyrethroid insecticides like α-cypermethrin through inhibition of cytochrome P450 monooxygenase responsible for pyrethroid detoxification. The aim of this study was to investigate, if the enantiomers of imazalil are selective in their synergistic potential in a mixture with a pyrethroid insecticide tested in Chironomus riparius. Potential enantioselectivity was studied on the level of uptake and elimination, inhibition of cytochrome P450 activity measured in vitro and in vivo and on synergistic potential of α-cypermethrin induced immobilization. Synergy was measured as an increase in α-cypermethrin toxicity after 144h applying a constant non-lethal imazalil concentration of 0.65 μmol/L. The R- and S-imazalil enantiomers increased α-cypermethrin toxicity from an EC50 of 1580 ± 980 pmol/L to an EC50 of 83 ± 10 pmol/L and 53 ± 8 pmol/L, respectively. The relatively small potency difference between imazalil enantiomers could not be explained by the in vitro cytochrome P450 inhibition, as the IC50 values were similar (0.11 ± 0.01 and 0.09 ± 0.01 μmol/L for R- and S-imazalil). Measuring in vivo P450 inhibition and the toxicokinetic of imazalil did not show a clear trend of selectivity towards one or the other enantiomer. The study therefore suggests that cytochrome P450 enzymes involved in detoxification in C. riparius are not enantioselective for imazalil.

Augmentation of C17.2 Neural Stem Cell Differentiation via Uptake of Low Concentrations of ssDNA-Wrapped Single-Walled Carbon Nanotubes.

Nanostructured biomaterials are extensively explored in clinical imaging and in gene/drug delivery applications. However, limited studies are performed that examine the influence that nanomaterials may have on cell behavior over long time scales at nonlethal concentrations. This study is designed to investigate whether carbon nanotubes are able to augment cell behavior at low concentrations. Single-walled carbon nanotubes are introduced to neural stem cells at different stages of differentiation at concentrations as low as 5 ng mL-1 . Results demonstrate that in this particular cell model, nanotube uptake is mediated by endocytosis. Differentiation is augmented, especially when nanotubes are introduced to cells in an actively dividing state. Significant increases in neuronal cell population are observed over the control specimens. While the mechanisms behind this observation are yet unknown, this study demonstrates that low concentrations of internalized nanomaterials can significantly alter the differentiation profile of a stem cell line.

Comparisons of acute inflammatory responses of nose-only inhalation and intratracheal instillation of ammonia in rats

Objective: To establish a rat model with respiratory and pulmonary responses caused by inhalation exposure to non-lethal concentrations of ammonia (NH3) that can be used for evaluation of new medical countermeasure strategies for NH3-induced acute lung injury (ALI). This is of great value since no specific antidotes of NH3-induced injuries exist and medical management relies on supportive and symptomatically relieving efforts.Methods: Female Sprague-Dawley rats (8–9 weeks old, 213g ± 2g) were exposed to NH3 using two different exposure regimens; nose-only inhalation or intratracheal instillation. The experiment was terminated 5 h, 24 h, 14 and 28 days post-exposure.Results: Nose-only inhalation of NH3 (9000–15 000 ppm) resulted in increased salivation and labored breathing directly post-exposure. Exposure did not increase inflammatory cells in bronchoalveolar lavage fluid but exposure to 12 000 ppm NH3 during 15 min reduced body weight and induced coagulation abnormalities by increasing serum fibrinogen levels. All animals were relatively recovered by 24 h. Intratracheal instillation of NH3 (1%) caused early symptoms of ALI including airway hyperresponsiveness, neutrophilic lung inflammation and altered levels of coagulation factors (increased fibrinogen and PAI-1) and early biomarkers of ALI (IL-18, MMP-9, TGFβ) which was followed by increased deposition of newly produced collagen 14 days later. Histopathology analysis at 5 h revealed epithelial desquamation and that most lesions were healed after 14 days.Conclusions: This study demonstrates that intratracheal instillation can reproduce several early hallmarks of ALI. Our findings therefore support that the intratracheal instillation exposure regimen can be used for new medical countermeasure strategies for NH3-induced ALI.

Toxicity and chronic effects of deltamethrin exposure on zebrafish (Danio rerio) as a reference model for freshwater fish community

In this study zebrafish specimens were exposed for 15 days to 0.25, 0.5, 1 and 2 μg L−1 non-lethal concentrations of deltamethrin (DM) knowing that is the active compound in insecticides used on agricultural crops. They were investigated important issues resulted during the chronic exposure with DM: effects on aggressive behavior and swimming performances knowing that is a high neurotoxic compound; toxicity on nervous system investigated on telencephalon, optic tectum and cerebellum; activity of PCNA, p53 and TUNEL as toxicity markers in immunocytochemistry of the histological samples; changes of elements concentrations in the fish body and their role in detoxification of DM. This scenario investigated the harmful effects of this compound for freshwater fish communities. The aggressive behavior significantly increased and remained constant for the concentration 0.5 μg L−1. They were not evidences in changing of anxiety level and swimming performances. The nervous system suffered significant damage for all studied concentrations and confirmed the changes in the behavior. Selenium concentration in the body decreased and may be involved in the detoxification processes.

Ionic liquids strongly affect the interaction of bacteria with magnesium oxide and silica nanoparticles.

Quaternary ammonium theophylline-based ionic liquids and imidazolium-based ionic liquids, magnesium oxide and silica nanoparticles were used in order to investigate the interaction with Gram negative Escherichia coli and Gram positive Bacillus cereus. The changes of bacterial sensitivity to both nanoparticles (NPs) and ionic liquids (ILs) were examined. In order to assess the impact of ILs on the interaction of nanoparticles with bacteria, respirometric analysis, activity of dehydrogenases, peroxidase analyses as well as scanning and fluorescence microscopy examinations were conducted. The interactions of ILs with nanoparticles based on adsorption and sedimentation tests were also investigated in order to assess how the ILs affect the agglomeration of NPs. It was assumed, as the main hypothesis of the present studies, that the differences in sensitivity of bacteria to combined ILs and NPs can be observed, even if the concentration of both compounds are below the minimum inhibitory concentration (MIC). The results indicated that ILs strongly affected the sensitivity of bacteria to nanoparticles however, the changes of sensitivity depended on the surface characteristics of the nanoparticles. The presence of ILs at non-lethal concentrations caused an increase of bacterial sensitivity to MgO nanoparticles. Notably, the sensitivity of Gram positive bacteria increased significantly when ILs were present. This was an important observation because the toxicity of nanoparticles toward Gram positive bacteria is usually lower than their toxicity toward Gram negative bacteria. Using silica nanoparticles, the presence of ionic liquids caused the adsorption of bacteria onto the surface of nanoparticle agglomerates. In conclusion, two opposing effects have been observed. On the one hand, the toxicity of MgO NPs in the presence of ILs has increased. On the other hand, the presence of silica nanoparticles caused a decreased sensitivity of both types of bacteria toward ILs. Our studies indicate potentially useful processes in many environmantal protection technologies like water treatment where flocculation and disinfection are extremely needed.

Induction of mTOR-dependent autophagy by WS2 nanosheets from both inside and outside of human cells.

The applications of two-dimensional transition metal dichalcogenides (2D TMDCs) pose an increased risk to both the environment and human health. Due to the large surface area of 2D nanosheets, they often form multi-layered nanoclusters of various thicknesses in aqueous solution. In this work, we address the safety issue of 2D TMDCs with focus on the cellular effects of the thickness of WS2 nanosheets. At a very low and non-lethal concentration (4 cm2 mL-1 or 25 μg mL-1), 4-layered WS2 nanosheets (WS2-4) were primarily bound to the cell surface with less internalization, while 30-layered WS2 nanosheets (WS2-30) were mostly internalized by human bronchial epithelial cells. Although the cellular interactions at this low concentration caused no alterations in the cell cycle, apoptosis, necrosis and cytotoxicity, cell autophagy was induced in both cases through mTOR-dependent pathways by perturbing a number of signaling molecules, such as amyloid precursor protein (APP) and cysteine-X-cysteine chemokine receptor 4 (CXCR-4). The finding of activation of cell autophagy from both outside and inside of cells reveals a novel feature of biological perturbations by 2D nanosheets. This finding will help in the formulation of general guidelines for the safe application of 2D nanomaterials.

Environmentally relevant concentrations of bifenthrin affect the expression of estrogen and glucocorticoid receptors in brains of female western mosquitofish

In recent decades, pyrethroid pesticides have been deemed a safer alternative to previously used pesticides. While some evidence supports this assumption in mammals and birds, exposure to certain pyrethroids can affect concentrations of hormones vital to reproduction in fish. Thus, we hypothesized that pyrethroid exposure impacts fish reproductive behavior and the expression of genes associated with reproduction. We tested our hypothesis by examining effects of the widely used pyrethroid pesticide, bifenthrin, on the reproductive behaviors of the broadly distributed livebearing western mosquitofish, Gambusia affinis. We exposed sexually mature female fish to one of five environmentally relevant concentrations of bifenthrin and conducted behavioral assays to assess reproductive, social, and space use behaviors before and after exposure. We did not detect changes in behaviors measured in response to bifenthrin. However, exposure was associated with increased expression of an estrogen receptor gene (ER-α) and glucocorticoid receptor (GR) in brain tissue at bifenthrin concentrations at concentrations of 5.90 and 24.82 ng/L, and 5.90 and 12.21 ng/L, respectively. Our study supports the perspective that the use of multiple endpoints through integrative approaches is essential for understanding the cumulative impact of pollutants. Integrating physiological, morphological, and behavioral investigations of nonlethal concentrations of pollutants like bifenthrin may heighten our potential to predict their impact on individuals, populations, and communities.