Multi-omics reveals 2-bromo-4,6-dinitroaniline (BDNA)-induced hepatotoxicity and the role of the gut-liver axis in rats.

2-Bromo-4, 6-dinitroaniline (BDNA) is a widespread azo-dye-related hazardous pollutant. However, its reported adverse effects are limited to mutagenicity, genotoxicity, endocrine disruption, and reproductive toxicity. We systematically assessed the hepatotoxicity of BDNA exposure via pathological and biochemical examinations and explored the underlying mechanisms via integrative multi-omics analyses of the transcriptome, metabolome, and microbiome in rats. After 28 days of oral administration, compared with the control group, 100 mg/kg BDNA significantly triggered hepatotoxicity, upregulated toxicity indicators (e.g., HSI, ALT, and ARG1), and induced systemic inflammation (e.g., G-CSF, MIP-2, RANTES, and VEGF), dyslipidemia (e.g., TC and TG), and bile acid (BA) synthesis (e.g., CA, GCA, and GDCA). Transcriptomic and metabolomic analyses revealed broad perturbations in gene transcripts and metabolites involved in the representative pathways of liver inflammation (e.g., Hmox1, Spi1, L-methionine, valproic acid, and choline), steatosis (e.g., Nr0b2, Cyp1a1, Cyp1a2, Dusp1, Plin3, arachidonic acid, linoleic acid, and palmitic acid), and cholestasis (e.g., FXR/Nr1h4, Cdkn1a, Cyp7a1, and bilirubin). Microbiome analysis revealed reduced relative abundances of beneficial gut microbial taxa (e.g., Ruminococcaceae and Akkermansia muciniphila), which further contributed to the inflammatory response, lipid accumulation, and BA synthesis in the enterohepatic circulation. The observed effect concentrations here were comparable to the highly contaminated wastewaters, showcasing BDNA's hepatotoxic effects at environmentally relevant concentrations. These results shed light on the biomolecular mechanism and important role of the gut-liver axis underpinning BDNA-induced cholestatic liver disorders in vivo.

Short-term toxicity assessment of combined use of zidovudine, lamivudine and lopinavir/ritonavir in vitro and in vivo.

INTRODUCTION: A combination of zidovudine (AZT), lamivudine (3TC) and lopinavir/ritonavir (LPV/r) is one of the most effective drugs for preventing mother-to-child transmission (PMTCT) of HIV. However, limited information is available regarding its systemic toxicity. This study aimed to investigate its potential toxicity. METHOD: An acute oral toxicity test was conducted to assess the potential acute toxicity of AZT + 3TC + LPV/r. Bacterial reverse mutation, mammalian erythrocyte micronucleus and mouse spermatogonia chromosomal aberration tests were conducted to assess its potential genotoxicity. A 28-day feeding test was conducted to assess the potential subacute toxicity. RESULTS: In mice, the LD50 of the AZT + 3TC + LPV/r mixture was greater than 2000 mg/kg body weight (BW). The rate of micronucleated polychromatic erythrocytes (PCEs) increased in a dose-dependent manner in mice (P < 0.01). After treatment with AZT + 3TC + LPV/r for 28 days, the BW gain of male and female rats in the high-dose group was lower than that in the control group (P < 0.05); the relative weights of the liver, kidney, spleen and brain increased (P < 0.05); and pathological abnormalities appeared in the thyroid and spleen of male and female rats in the high-dose group. The haemoglobin (HGB) and red blood cells (RBCs) count in male and female rats decreased, but the white blood cells (WBCs) and lymphocyte apoptosis rates in male and female rats in the high-dose group increased (P < 0.05). The total protein, albumin, cholesterol and blood glucose levels of male and female rats in the high-dose group were significantly decreased (P < 0.05). The alanine aminotransferase (ALT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), creatinine (Cr) and blood urea nitrogen (BUN) levels of male and female rats in the medium- and high-dose groups increased significantly (P < 0.05). CONCLUSION: The results suggest that AZT + 3TC + LPV/r may exhibit genotoxicity and subacute toxicity under experimental conditions.

Toxicological evaluation of water-extract sericin from silkworm (Bombyx mori) in pregnant rats and their fetus during pregnancy.

Sericin is a natural protein produced by the silkworm Bombyx mori, which has a wide range of biological activities and has a broad application prospect in multiple areas. However, systemic toxicity and safety assessment of sericin is still rare. This study was aimed to evaluate the toxic effects of water-extract sericin from cocoons of Bombyx mori in pregnant rats and their fetuses during pregnancy. Eighty pregnant rats were randomly divided into three treatment groups, one negative and one positive control group. The treatment groups were administered water-extract sericin solutions at doses of 1,000, 500, and 250 mg/kg, while the negative and positive control groups were administered pure water and 300 mg/kg aspirin, respectively. Rats were exposed daily by oral gavage from the seventh day of gestation for 10 consecutive days and sacrificed on the 20th day of gestation. The results showed that water-extract sericin did not induce any treatment-related changes on pregnant rats (clinical signs, body weights, food consumption, ovarian and uterine weights) and fetuses (body weights, body lengths, tail lengths, visceral, and skeletal development). The no-observed-adverse-effect-level (NOAEL) of sericin was determined to be 1,000 mg/kg body weight in rats. These results indicated that water-extract sericin is of low teratogenic potential under the experimental conditions of this study.

Evaluation of the genotoxicity and teratogenicity of xylan using different model approaches.

Xylan is the second most abundant polysaccharide group in plants and has a wide variety of food and pharmaceutical applications. However, little information on the safety assessment of extracted xylan as dietary supplement is available. As part of a comprehensive toxicological assessment, this study examined the potential toxicity of xylan extracted from sugarcane bagasse by three genotoxicity studies (Ames test, in vivo mice bone marrow micronucleus test, and mice sperm abnormality test) and a teratogenicity study in rats. In the Ames test, xylan showed no mutagenic activity on histidine dependent strains of Salmonella typhimurium at concentrations up to 5000 µg/plate; results of the in vivo mice bone marrow micronucleus test and mice sperm abnormality test indicated no significant effect on sperm morphology and micronucleus rate of polychromatic erythrocytes in mice at doses up to 5 g/kg body weight. In the teratogenicity study, a total of 60 pregnant rats were exposed to 10, 5, and 2.5% xylan in diet, from gestation days 7 to 16, and the no-observed-adverse-effect levels (NOAEL) of xylan was determined to be 9.8 g/kg body weight. The safe dose of xylan for human was estimated to be 98 mg/kg/day (i.e., 6.86 g/day for a 70-kg person), using a 100-fold safety factor. Taken together, results of this study indicated that xylan is practically nontoxic in terms of potential dietary consumption by humans in food or as a dietary supplement.

Teratogenicity of 30 nm Aluminum Oxide Nanoparticles (Al2O3NPs) in Rats by Gavage.

Aluminum oxide nanoparticles (Al2O3NPs) are one class of widely used nanomaterials. However, the teratogenicity toxicity of Al2O3NPs in mammal remains poorly understood. This study was aimed to evaluate the teratogenicity of Al2O3NPs in Sprague Dawley (SD) rats by gavage and to compare the effects of Al2O3NPs to those of equivalent dose of microscale aluminum oxide (bulk Al2O3). Sixty pregnant rats were randomly divided into 5 groups and treated with 100 and 200 mg/kg body weight (bw) Al2O3NPs (30 nm), 200 mg/kg bulk Al2O3, deionized water (as the negative control), and 300 mg/kg aspirin (as the positive control). Rats were exposed daily by oral gavage from the 7th day of gestation for 10 consecutive days and sacrificed on the 20th day of gestation. Results of the study showed that there were no significant effects of Al2O3NPs on pregnant rats (clinical signs, body weight, food consumption, ovary and uterus weight, number of corpora lutea) and fetuses (body weight, sex, body length, tail length, skeletal and visceral development). Under the experimental conditions of the present study, 10 consecutive days of repeated oral administration of Al2O3NPs at doses of up to 200 mg/kg/day did not induce any treatment-related teratogenicity in SD rats. Accordingly, the NOAEL was determined to be 200 mg/kg Al2O3NPs (106 mg Al/kg bw/day) in rats. The teratogenic effects of Al2O3NPs in rats were comparable to those of the bulk Al2O3 of same doses (200 mg/kg).

Safety Assessment of Water-Extract Sericin from Silkworm (Bombyx mori) Cocoons Using Different Model Approaches.

Sericin is a natural protein component of silks of silkworm and has potential utility in multiple areas such as pharmacological, cosmetics, and biotechnological industries. However, the understanding of its toxicological safety is still limited. This study evaluated the safety of water-extract sericin from silkworm (Bombyx mori) cocoons using different model approaches, including three genotoxicity studies (the bacterial reverse mutation test, the mammalian erythrocyte micronucleus test, and the mouse spermatogonia chromosomal aberration test) and a 90-day subchronic toxicity study in Sprague-Dawley (SD) rats. The results of this study showed that water-extract sericin was nonmutagenic and nongenotoxic both in vitro and in vivo. Sericin did not induce significant changes in the body and organ weight, food intake, blood hematology and serum biochemistry, urine index, and histopathology in rats. The NOAEL of sericin was determined to be 1 g/kg/day for male and female rats. These results indicated that water-extract sericin was of low toxicity in the experimental conditions of the current study and had the potential for application in food-related products.

Genotoxicity and teratogenicity of seabuckthorn (Hippophae rhamnoides L.) berry oil.

As botanicals and dietary supplements are used increasingly in many countries, the issue of safety is particularly critical for regulation of food products containing these substances. Seabuckthorn (Hippophae rhamnoides L.) has been used for centuries as a medicine and nutritional supplement in Asia and Europe. However, data regarding to the safety assessment of the plant and its extracts is still rare. This study was to evaluate the potential toxicity of seabuckthorn berry (SB) oil conducted in three genotoxicity studies and a teratogenicity study. Results of the genotoxicity studies indicated that SB oil has no genotoxicity under the experimental conditions of this study. Specifically, SB oil did not display any mutagenic activity on histidine dependent strains of Salmonella typhimurium under exposure concentrations of 8, 40, 200, 1000, and 5000 µg/plate; SB oil did not have significant effect on sperm morphology and have no influence on micronucleus rate of polychromatic erythrocytes in mice at doses of 9.36, 4.68, and 2.34 g/kg body weight. In the teratogenicity study, pregnant rats were treated with 4.68, 2.34, and 1.17 g/kg SB oil from gestation day 7 to 16 and no treatment-related maternal toxicity or embryo toxicity was observed. Taken together, these results support the safe use of seabuckthorn berry oil for potential dietary consumption in food or as a dietary supplement.

Subchronic Study of a White Kidney Bean (Phaseolus vulgaris) Extract with α-Amylase Inhibitory Activity.

Common bean extract as a dietary supplement has received increased attention globally owing to its α-amylase inhibitory activity. The objective of this study was to evaluate the toxicity of a white kidney bean (Phaseolus vulgaris) extract by a repeated-dose 90-day subchronic oral toxicity study in Sprague-Dawley rats. In the subchronic toxicity study, 80 rats were orally administrated with white kidney bean extract at doses of 4, 2, and 1 g/kg body weight daily for 90 days. The results showed that the white kidney bean extract at doses up to 4 g/kg/day did not induce significant changes in body weight, organ weight, food consumption, hematology, serum biochemistry, and histopathology in rats, as compared to the control. The no-observed-adverse-effect level (NOAEL) of white kidney bean extract was determined to be >4 g/kg/day for both male and female rats, under the experimental conditions of this study.

Long-term exposure to low level of fluoride induces apoptosis via p53 pathway in lymphocytes of aluminum smelter workers.

Long-term occupational exposure to low level of fluoride can induce oxidative stress and apoptosis in many cells, including lymphocyte. However, the underlying mechanism remains unclear. Hence, this study was designed to explore the potential oxidative stress and apoptosis of long-term occupational exposure to low level of fluoride in aluminum smelter workers. A total of 120 aluminum smelter workers were recruited in control, low-, middle-, and high-fluoride exposure groups with 30 workers for each group. The peripheral blood samples were collected, centrifuged, and isolated to obtain serum and lymphocyte suspensions. The air and serum fluoride concentrations were detected by fluoride ion-selective electrode method. The lymphocytic apoptosis rate, DNA damage, oxidative stress, and mRNA levels of p53, Bcl-2, and Bax were assessed by Annexin V/PI staining, comet assay, attenuated total reflectance Fourier transform infrared spectroscopy and real-time polymerase chain reaction, respectively. Results showed that the air and serum fluoride concentrations of fluoride-exposed groups were higher than those of the control group (p < 0.05). Fluoride exposure might induce apoptosis, DNA damage and oxidative stress in a dose-dependent manner in lymphocytes (p < 0.05). The expression levels of p53 and Bax were increased with fluoride exposure in lymphocytes (p < 0.05), whereas the Bcl-2 expression was decreased but not significantly. Taken together, these observations indicate that long-term occupational exposure to low level of fluoride may lead to oxidative stress and induce apoptosis through the p53-dependent pathway in peripheral blood lymphocytes of aluminum smelter workers. Serum fluoride level may be the potential biomarker of fluoride exposure.

Acute and subchronic toxicity studies of seabuckthorn (Hippophae rhamnoides L.) oil in rodents.

Seabuckthorn (Hippophae rhamnoides L.) has been traditionally used as medicine and nutritional supplement for a long period of time. However, information on the systemic toxicity and safety evaluation of seabuckthorn and its extracts is still scarce. The purpose of this study was to evaluate the potential toxicity of seabuckthorn oil by an acute oral toxicity study in mice and a 90-day repeated oral toxicity study in rats. No mortality or signs of toxicity was observed in mice treated with 20 mL/kg body weight seabuckthorn oil in the acute toxicity study. In the subchronic toxicity study, 80 Sprague-Dawley rats (10 animals per sex per treatment group) were administrated with 10, 5, 2.5 and 0 (control) mL/kg body weight of seabuckthorn oil daily for 90 days by gavage. There were no signs of toxicity and treatment-related changes in rats treated with seabuckthorn oil on mortality, body and organ weights, food consumption, blood biochemistry and hematology, gross necropsy and histopathological examinations. Based on the finding of this study, the maximum tolerated dose of seabuckthorn oil was >20 mL/kg for mice for acute toxicity study, and the no-observed-adverse-effect level was 10 mL/kg body weight for both male and female rats for 90-day toxicity study.

Safety assessment of xylan by a 90-day feeding study in rats.

Xylans are present naturally in various plants and have important uses in nutrition, food, novel material and biotechnology; however, to date, data regarding their systemic toxicity and safety evaluation is still limited. This study investigated the potential toxicity of xylan from sugarcane bagasse by a subchronic toxicity study in rats. A total of 80 male and female rats were fed with diets containing 10%, 5%, 2.5% and 0% (control) xylan for 90 days. A toxicological assessment was performed including mortality, body and organ weights, food consumption, blood biochemistry, hematology, urinalysis, gross necropsy and histopathological examinations. There were no signs of toxicity and treatment-related changes in rats treated with xylan. The no-observed-adverse-effect levels (NOAEL) of xylan were 9.0 g kg-1 bw for males and 10.6 g kg-1 bw for females of rats under this experimental condition, respectively.

Toxicological evaluation of silver nanoparticles and silver nitrate in rats following 28 days of repeated oral exposure.

The increasing application of silver nanoparticles (AgNPs) has been raising concerns about their potential adverse effects to human and the environment. However, the knowledge on the systemic toxicity of AgNPs in mammalian systems is still limited. The present study investigated the toxicity of PVP-coated AgNPs in rats treated with repeated oral administration, and compared that with equivalent dose of AgNO3 . Specifically, one hundred male and female rats were orally administrated with particulate or ionic forms of silver (Ag) separately at doses of 0.5 and 1 mg kg-1 body weight daily for 28 days. The results reveal no significant toxic effects of AgNPs and AgNO3 up to 1 mg kg-1 body weight, with respect to the body weight, organ weight, food intake, and histopathological examination. Ag distribution pattern in organs of rats treated with AgNPs was similar to that of AgNO3 treated rats, showing liver and kidneys are the main target organs followed by testis and spleen. The total Ag contents in organs were significantly lower in the AgNPs treated rats than those in the AgNO3 treated rats. However, the comparisons between AgNPs and AgNO3 treatments further indicated more potent of AgNPs in biochemical and hematological parameters in rats, including red blood cell count (RBC), platelet count (PLT), white blood cell count (WBC) and aspartate transaminase (AST). Results of this study suggested that particulate Ag at least partially contributed to the observed toxicity of AgNPs, and both ionic and particulate Ag should be taken into consideration in toxicological evaluation of AgNPs. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 609-618, 2017.

Safety assessment of powdered Cistanche deserticola Y. C. Ma by a 90-day feeding test in Sprague-Dawley rats.

Cistanche deserticola (C. deserticola), a holoparasitic plant widely distributed in arid or semi-arid areas in Eurasia and North Africa, has been used as an important tonic in traditional Eastern medicine for centuries. However, little information on the systemic toxicity and safety evaluation of it is available. The purpose of this study was to investigate the potential toxicity of powdered C. deserticola as a novel food ingredient by use of a subchronic toxicity study in Sprague-Dawley (SD) rats. A total of 80 male and female rats were fed with diets containing 8, 4, 2 and 0% (control) powdered C. deserticola for 90 days. A toxicological assessment was performed including mortality, body and organ weight, food consumption, blood biochemistry, hematology, gross necropsy and histopathological examinations. There were no signs of toxicity and treatment-related changes in rats treated with powdered C. deserticola. The no-observed-adverse-effect level (NOAEL) of powdered C. deserticola was 7.8 g kg-1 body weight for males and 8.0 g kg-1 body weight for females of rats under the experimental conditions of this study.

Impact of Predator Cues on Responses to Silver Nanoparticles in Daphnia carinata.

The past decades have witnessed a boom in nanotechnology that has led to increasing production and application of silver nanoparticles (AgNPs) in the textile industry due to their antimicrobial properties. Increase in the manufacture and use of NPs inevitably has resulted in their increased release into aquatic environments resulting in the exposure of organisms living in these environments. Recently, the risk of exposure to NPs and the potential interaction with biological systems has received increasing attention. The present study investigated the potential effects of predator cues on the toxicity of environmentally relevant concentrations of AgNPs in Daphnia carinata at organismal and biochemical levels. The results of this study show that exposure to environmentally relevant concentrations of AgNPs can result in adverse effects on daphnids with 24- and 48-h LC50 values of 3.56 and 1.75 µg/L, respectively. Furthermore, significant inhibition of reproduction was observed at concentrations as low as 0.5 µg/L. Exposure to predator cues alone resulted in an increase in reproduction and inhibition of superoxide dismutase activity in daphnids. However, coexposure to predator cues interacted in an antagonistic manner with AgNPs with a 24-h LC50 value of 10.81 µg/L compared with 3.56 µg/L for AgNPs alone. In summary, AgNPs could pose risks to aquatic invertebrates at environmentally relevant concentrations. Interestingly, the presence of other factors, such as predator cues, moderated the effects of exposure to AgNPs. Therefore, there is a need to further investigate the potential interactions between NPs and biological factors that can modulate toxicity of NPs for application to the risk assessment of aquatic invertebrates.

Distinct expression profiles of stress defense and DNA repair genes in Daphnia pulex exposed to cadmium, zinc, and quantum dots.

The ever-increasing production and use of nanocrystaline semiconductors (Quantum dots; QDs) will inevitably result in increased appearance of these nanomaterials in the aquatic environment. However, the behavior and potential toxicity of heavy metal constituted nanoparticulates in aquatic invertebrates is largely unknown, especially with regard to molecular responses. The freshwater crustacean Daphnia pulex is a well-suited toxicological and ecological model to study molecular responses to environmental stressors. In this study, D. pulex were exposed for 48 h to sublethal doses of QDs (25% and 50% of LC50) with differing spectral properties (CdTe and CdSe/ZnS QDs) and Cd and Zn salts. Our data suggest that acute exposure to both CdSO4 and Cd-based QDs leads to Cd uptake in vivo, which was biologically supported by the observation of increased expression of metallothionein (MT-1). Furthermore, Cd, Zn, and CdSe/ZnS QDs induced different patterns of gene expression regarding stress defense and DNA repair, which furthers our knowledge regarding which response pathways are affected by nanoparticulate forms of metals versus ionic forms in aquatic crustaceans.

Effects of environmentally realistic daily temperature variation on pesticide toxicity to aquatic invertebrates.

The toxicity of several agricultural chemicals to aquatic invertebrates has been shown to be temperature-dependent, but the role of daily temperature variation has rarely been examined. The authors simulated a natural daily temperature pattern (a fluctuating cycle of 21 °C to 31 °C over a 24-h period) based on field-collected data from Southern High Plains wetlands (TX, USA) and conducted a series of experiments comparing responses from this exposure scenario to a constant exposure at 24 ± 1 °C. Results indicate alterations in pesticide toxicity under the fluctuating temperature regime compared with that of the constant temperature exposure. There was a significant interaction of temperature regime and bifenthrin on Chironomus dilutus survival, and C. dilutus ash-free dry mass was lower in the fluctuating temperature treatment. The 10-d median lethal concentration (LC50) for Hyalella azteca exposed to chlorothalonil was lower under the fluctuating temperature regime compared with the constant temperature regime. For Daphnia magna exposed to malathion, the main effects of temperature regime and malathion were observed on cholinesterase activity. The present study demonstrates how environmentally relevant daily temperature variation influences contaminant effects on aquatic invertebrates.

Bioaccumulation of petroleum hydrocarbons in fiddler crabs (Uca minax) exposed to weathered MC-252 crude oil alone and in mixture with an oil dispersant.

The Deepwater Horizon accident in the Gulf of Mexico resulted in a sustained release of crude oil, and weathered oil was reported to have washed onto shorelines and marshes along the Gulf coast. One strategy to minimize effects of tarballs, slicks, and oil sheen, and subsequent risk to nearshore ecosystem resources was to use oil dispersants (primarily Corexit® 9500) at offshore surface and deepwater locations. Data have been generated reporting how Corexit® 9500 and other dispersants may alter the acute toxicity of crude oil (Louisiana sweet crude) to marine organisms. However, it remains unknown how oil dispersants may influence bioaccumulation of petroleum hydrocarbons in nearshore crustaceans. We compare bioaccumulation of petroleum hydrocarbons in fiddler crabs (Uca minax) from exposures to the water accommodated fraction (WAF) of weathered Mississippi Canyon 252 oil (~30 d post spill) and chemically-enhanced WAF when mixed with Corexit® EC9500A. Whole body total petroleum hydrocarbon (TPH) concentrations were greater than background for both treatments after 6h of exposure and reached steady state at 96 h. The modeled TPH uptake rate was greater for crabs in the oil only treatment (k(u)=2.51 mL/g/h vs. 0.76 mL/g/h). Furthermore, during the uptake phase TPH patterns in tissues varied between oil only and oil+dispersant treatments. Steady state bioaccumulation factors (BAFs) were 19.0 mL/g and 14.1 mL/g for the oil only and oil+Corexit treatments, respectively. These results suggest that the toxicokinetic mechanisms of oil may be dependent on oil dispersion (e.g., smaller droplet sizes). The results also indicate that multiple processes and functional roles of species should be considered for understanding how dispersants influence bioavailability of petroleum hydrocarbons.

Effects of predator cues on pesticide toxicity: toward an understanding of the mechanism of the interaction.

Pesticide toxicity may be modified by a number of co-occurring environmental and ecological stressors. Coexposure to predator cues has been shown to potentiate and/or synergize toxicity of pesticides. However, the mechanisms behind these interactions are not well understood. Here we examine the effects of fish predator (bluegill, Lepomis macrochirus) cues on toxicity of five different pesticides to the freshwater cladoceran, Ceriodaphnia dubia. The purpose for examining patterns among pesticides was to test the idea that the mechanism of the interaction could be explained by a general stress response; that is, the interaction patterns would be similar regardless of the pesticide's mechanism of action [MOA]). Acute 96-h concentration-response experiments were conducted for pesticides with and without fish cues. Predator cues influenced the toxicity of pesticides and the interaction patterns varied among pesticides. Fipronil exhibited a synergistic interaction, while predator cues interacted antagonistically for bifenthrin and thiacloprid. Other compounds previously reported to potentiate toxicity (malathion) were found to act additively. The results demonstrate that factors such as pesticide bioavailability, K(OC) , and exposure concentration may be important for predicting the occurrence of these interactions and that patterns were not consistent among pesticides varying in MOA. Predator stress is an important component for structuring communities and ecosystem processes. Fully understanding how this process may interact with organic contaminants may best be achieved by examination at toxicokinetic and toxicodynamic scales.