Chronic Exposure to Environmentally Relevant Concentrations of Tetracycline Perturbs Gut Homeostasis in Zebrafish

The prevalence of residual antibiotics is a global threat to human health.Less is known about the potential health effects of residual antibiotics in freshwater systems.Here,zebrafish were used to explore chronic effects of environmentally relevant concentrations(ERCs)of tetracycline(TC).Although chronic exposure to TC did not significantly alter the body weight of adult zebrafish,the exposed zebrafish parents exhibited substantial changes in gut microbiota composition and a reduced gut-weight ratio.Notably,male fish exposed to TC showed a significant decline of critical intestinal function-related metabolites(i.e.,triglycerides,glucose,and free fat acid),while this was not observed in females,resulting in sex-dependency.The gut microbial composition of chronically exposed zebrafish parents changed substantially,but the disruption was not transferred to their respective offspring without exposure.However,the perturbation of insulinrelated signaling pathways caused by TC exposure was not attenuated in the zebrafish offspring after removal of TC exposure.Taken together,our findings suggest that chronic exposure to TC disturbs gut microbial communities and metabolism and exerts chronic effects on the insulin/IGF-1 mediated signaling cascades,implying that exposure to antibiotics not only leads to the selection of resistant microbes but also poses long-term deleterious health risks for the next generation.

Genomic Instability Is a Mechanism for Diminished Male Fertility Following Chronic Dichlorvos Exposure

Background and Objectives: Chronic low-dose exposure to dichlorvos occurs in communities in Africa where the substance is used indiscriminately for a variety of purposes. This experiment used an animal model to evaluate genomic instability induced by this pattern of chronic exposure and its relationship with some measures of fertility in males. Methods: Seventy-five male Rattus norvegicus rats obtained for this experiment, were randomly allotted into five groups. Dichlorvos was given by oral gavage at doses of 0.28 mg/kg, 0.56 mg/kg and, 1.68 mg/kg, respectively, to three of the groups, on alternate days for 50 weeks. The remaining two groups received plain drinking water and cyclophosphamide as negative and positive controls, respectively. Samples were collected at 17, 34, and 50 weeks. Sperm count, sperm morphology and serum levels of follicle-stimulating hormone, luteinizing hormone, dihydrotestosterone, oestrogen and progesterone were determined. Furthermore, the frequency of micronucleated polychromatic erythrocytes was determined in bone marrow cells obtained from the femur. Results: The mean ranks of micronuclei frequency had an increasing trend. The frequency of micronucleated polychromatic erythrocytes (MnPCE) had a significant negative correlation with oestrogen (rs = -0.47, p = 0.00, n = 50), follicle-stimulating hormone (rs = -0.41, p = 0.00, n = 50) and progesterone (rs = -0.37, p = 0.01, n = 50) serum levels. A positive monotonic relationship also existed between micronuclei frequency and those of tubular necrosis, tubular vacuolation, and residual bodies. A positive significant moderate correlation was found between MnPCE and the proportion of immotile sperms (rs = 0.41, p = 0.00, n = 50). Conclusion: The nature of the correlations between micronuclei frequency and the proportion of immotile sperms, adverse histological changes and serum hormone levels found in this study suggest genomic instability as the possible mechanism for diminished fertility in males chronically exposed to dichlorvos.

Oxidative damage of chronic PM2.5 exposure to rat lung tissue

Objective:To investigate the oxidative damage caused by chronic exposure to PM2.5 in rat lung tissues according to the PM2.5 concentration in the atmospheric environment.Methods:Forty healthy SD rats were randomly divided into 4 groups,the control group and the 5 mg/kg,10 mg/kg,20 mg/kg PM2.5 exposure groups.The rats in each group were administered by intratracheal instillation with 1ml/kg,2 times per week.After 8 weeks of exposure,the changes of lung histopathology,superoxide dismutase(SOD),malondialdehyde(MDA),catalase(CAT)and reduced glutathione/oxidized glutathione(GSH/GSSG)were detected.Results:Compared with the control group,the histopathological changes in the lung tissue of the three dose groups exposed to PM2.5 were significant.Masson staining showed obvious collagen fibrosis.The activities of SOD and CAT in the lung tissue of the exposed group were decreased,and the contents of MDA were increased with statistically significant(P<0.05),showing that the difference was the most obvious in the middle-dose group.The GSH/GSSG ratio in the lung tissue decreased,indicating that the degree of oxidative damage was aggravated.Conclusion:Chronic exposure to PM2.5 could induce oxidative stress in the lung tissue of rats and caused different degrees of oxidative damage.

Stress injuries and autophagy in mouse hippocampus after chronic cold exposure

Cold exposure is an external stress factor that causes skin frostbite as well as a variety of diseases.Estrogen might participate in neuroprotection after cold exposure,but its precise mechanism remains unclear.In this study,mice were exposed to 10°C for 7 days and 0–4°C for 30 days to induce a model of chronic cold exposure.Results showed that oxidative stress-related c-fos and cyclooxygenase 2 expressions,MAP1LC3-labeled autophagic cells,Iba1-labeled activated microglia,and interleukin-1β-positive pyramidal cells were increased in the hippocampal CA1 area.Chronic cold exposure markedly elevated the levels of estrogen in the blood and the estrogen receptor,G protein-coupled receptor 30.These results indicate that neuroimmunoreactivity is involved in chronic cold exposure-induced pathological alterations,including oxidative stress,neuronal autophagy,and neuroimmunoreactivity.Moreover,estrogen exerts a neuroprotective effect on cold exposure.

Chronic corticosterone disrupts the circadian rhythm of CRH expression and m^(6)A RNA methylation in the chicken hypothalamus

Background:Corticotropin-releasing hormone(CRH),the major secretagogue of the hypothalamic-pituitary-adrenal(HPA)axis,is intricately intertwined with the clock genes to regulate the circadian rhythm of various body functions.N6-methyladenosine(m^(6)A)RNA methylation is involved in the regulation of circadian rhythm,yet it remains unknown whether CRH expression and m^(6)A modification oscillate with the clock genes in chicken hypothalamus and how the circadian rhythms change under chronic stress.Results:Chronic exposure to corticosterone(CORT)eliminated the diurnal patterns of plasma CORT and melatonin levels in the chicken.The circadian rhythms of clock genes in hippocampus,hypothalamus and pituitary are all disturbed to different extent in CORT-treated chickens.The most striking changes occur in hypothalamus in which the diurnal fluctuation of CRH mRNA is flattened,together with mRNA of other feeding-related neuropeptides.Interestingly,hypothalamic m^(6)A level oscillates in an opposite pattern to CRH mRNA,with lowestm^(6)A level after midnight(ZT18)corresponding to the peak of CRH mRNA before dawn(ZT22).CORT diminished the circadian rhythm of m^(6)A methylation with significantly increased level at night.Further site-specific m^(6)A analysis on 3’UTR of CRH mRNA indicates that higher m^(6)A on 3’UTR of CRH mRNA coincides with lower CRH mRNA at night(ZT18 and ZT22).Conclusions:Our results indicate that chronic stress disrupts the circadian rhythms of CRH expression in hypothalamus,leading to dysfunction of HPA axis in the chicken.RNA m^(6)A modification is involved in the regulation of circadian rhythms in chicken hypothalamus under both basal and chronic stress conditions.

Glucose supplementation improves intestinal amino acid transport and muscle amino acid pool in pigs during chronic cold exposure

Mammals in northern regions chronically suffer from low temperatures during autumn-winter seasons.The aim of this study was to investigate the response of intestinal amino acid transport and the amino acid pool in muscle to chronic cold exposure via Min pig models(cold adaptation)and Yorkshire pig models(non-cold adaptation).Furthermore,this study explored the beneficial effects of glucose supplementation on small intestinal amino acid transport and amino acid pool in muscle of cold-exposed Yorkshire pigs.Min pigs(Exp.1)and Yorkshire pigs(Exp.2)were divided into a control group(17℃,n=6)and chronic cold exposure group(7℃,n=6),respectively.Twelve Yorkshire pigs(Exp.3)were divided into a cold control group and cold glucose supplementation group(8℃).The results showed that chronic cold exposure inhibited peptide transporter protein 1(PepT1)and excitatory amino acid transporter 3(EAAT3)expression in ileal mucosa and cationic amino acid transporter-1(CAT-1)in the jejunal mucosa of Yorkshire pigs(P<0.05).In contrast,CAT-1,PepT1 and EAAT3 expression was enhanced in the duodenal mucosa of Min pigs(P<0.05).Branched amino acids(BCAA)in the muscle of Yorkshire pigs were consumed by chronic cold exposure,accompanied by increased muscle RING-finger protein-1(MuRF1)and muscle atrophy F-box(atrogin-1)expression(P<0.05).More importantly,reduced concentrations of dystrophin were detected in the muscle of Yorkshire pigs(P<0.05).However,glycine concentration in the muscle of Min pigs was raised(P<0.05).In the absence of interaction between chronic cold exposure and glucose supplementation,glucose supplementation improved CAT-1 expression in the jejunal mucosa and PepT1 expression in the ileal mucosa of cold-exposed Yorkshire pigs(P<0.05).It also improved BCAA and inhibited MuRF1 and atrogin-1 expression in muscle(P<0.05).Moreover,dystrophin concentration was improved by glucose supplementation(P<0.05).In summary,chronic cold exposure inhibits amino acid absorption in the small intestine,depletes BCAA and promotes protein degradation in muscle.Glucose supplementation ameliorates the negative effects of chronic cold exposure on amino acid transport and the amino acid pool in muscle.

Role of nitric oxide in the genotoxic response to chronic microcystin-LR exposure in human–hamster hybrid cells

Microcystin-LR （MC-LR） is the most abundant and toxic microcystin congener and has been classified as a potential human carcinogen （Group 2B） by the International Agency for Research on Cancer. However, the mechanisms underlying the genotoxic effects of MC-LR during chronic exposure are still poorly understood. In the present study, human-hamster hybrid （AL） cells were exposed to MC-LR for varying lengths of time to investigate the role of nitrogen radicals in MC-LR-induced genotoxicity. The mutagenic potential at the CD59 locus was more than 2-fold higher （p 〈 0.01） in AL ceUs exposed to a cytotoxic concentration （1 μmol/L） of MC-LR for 30 days than in untreated control ceils, which was consistent with the formation of micronucleus. MC-LR caused a dose-dependent increase in nitric oxide （NO） production in treated cells. Moreover, this was blocked by concurrent treatment with the NO synthase inhibitor NC-methyl-L-arginine （L-NMMA）, which suppressed MC-LR- induced mutations as well. The survival of mitochondrial DNA-depleted （pO） AL ceils was markedly decreased by MC-LR treatment compared to that in AL cells, while the CD59 mutant fraction was unaltered. These results provided clear evidence that the genotoxicity associated with chronic MC-LR exposure in mammalian cells was mediated by NO and might be considered as a basis for the development of therapeutics that prevent carcinogenesis.

Accumulation and elimination of iron oxide nanomaterials in zebrafish(Danio rerio) upon chronic aqueous exposure

A 52-day continuous semi-static waterborne exposure（test media renewed daily） regimen was employed to investigate the accumulation and elimination profiles of two iron oxide nanomaterials（nano-Fe2O3 and nano-Fe3O4） in zebrafish（Danio rerio）. Adult zebrafish were exposed to nanomaterial suspensions with initial concentrations of 4.0 and 10.0 mg/L for28 days and then were moved to clean water for 24 days to perform the elimination experiment. Fe content was measured in fish body and feces to provide data on accumulation and elimination of the two iron oxide nanomaterials in zebrafish. The experiment revealed that：（1） high accumulation of nano-Fe2O3 and nano-Fe3O4 were found in zebrafish, with maximum Fe contents, respectively, of 1.32 and 1.25 mg/g for 4.0 mg/L treatment groups and 1.15 and 0.90 mg/g for 10.0 mg/L treatment groups;（2） accumulated nanoparticles in zebrafish can be eliminated efficiently（the decrease of body burden of Fe conforms to a first-order decay equation） when fish were moved to nanoparticle-free water,and the elimination rates ranged from 86% to 100% by 24 days post-exposure; and（3）according to analysis of Fe content in fish excrement in the elimination phase, iron oxide nanomaterials may be adsorbed via the gastrointestinal tract, and stored for more than12 days.

Histone acetylation of the htr3a gene in the prefrontal cortex of Wistar rats regulates ethanol-seeking behavior

Previous reports showed that decreased histone deacetylase activity significantly potentiated the rewarding effects of psychostimulants, and that encoding of the 5-HT3 receptor by the htr3a gene was related to ethanol-seeking behavior. However, the effects of a histone deacetylase inhibitor on ethanol-seeking behavior and epigenetic regulation of htr3a mRNA expression after chronic ethanol exposure are not fully understood. Using quantitative reverse transcription-polymerase chain reaction and chromatin immunoprecipitation analysis, we investigated the effects of chronic ethanol exposure and its interaction with a histone deacetylase inhibitor on histone-acetylation-mediated changes in htr3a mRNA expression in the htr3a promoter region. The conditioned place preference procedure was used to evaluate ethanol-seeking behavior. Chronic exposure to ethanol effectively elicited place conditioning. In the prefrontal cortex, the acetylation of H3K9 and htr3a mRNA expression in the htr3a promoter region were significantly higher in the ethanol group than in the saline group. The histone deacetylase inhibitor sodium butyrate potentiated the effects of ethanol on htr3a mRNA expression and enhanced ethanol-induced conditioned place preferences. These results suggest that ethanol upregulates htr3a levels through mechanisms involving H3K9 acetylation, and that histone acetylation may be a therapeutic target for treating ethanol abuse.

Risk of cataract of different morphological types in Urals population chronically exposed at low doses

Objective To assess the risk of lens opacity depending on the dose in the population exposed to external and internal radiation for a long time,based on a long-term clinical follow-up of the cohorts of people exposed to radiation as a result of two radiation accidents in the South Urals.Methods A layer-by-layer morphological study of the lens of 1,377 exposed individuals was conducted according to a special program in the Clinical Department of the Urals Research Center for Radiation Medicine(URCRM)68 years after the onset of exposure.Lens changes were classified in accordance with the lens opacities classification(LOCS)III and included images of lens.To calculate the doses to lens,the data were taken into account,including the person's residence history in the radioactively contaminated territory,age and sex that affected their lifestyle and diet,radionuclide distribution in organs and tissues.Individualized lens doses were calculated using Techa River Dosimetry System(TRDS)-2016.Case-control method was used for statistical analysis.Results Individual values of absorbed dose to lens ranged from 0 to 600mGy.A dose-dependent increased risk of posterior subcapsular cataract(PSC)(OR1.54,95%CI:1.04–2.27)and nuclear cataract(OR1.84,95%CI:1.14–2.95)was found among chronically exposed individuals by a case-control method.No evidence of dose effect was found for cortical cataracts.Conclusions The results showed that population exposed to long-term chronic low-dose radiation was subjected to an increased risk of PSC and nuclear cataract development.

Reproductive toxicity of enrofloxacin in Caenorhabditis elegans involves oxidative stress-induced cell apoptosis

Fluoroquinolone antibiotics(FQs)that persist and bioaccumulate in the environment have aroused people’s great concern.Here,we studied the adverse effects of FQs in soil animals of Caenorhabditis elegans via food-chronically exposure.The result shows C.elegans exposed to FQs exhibited reproductive toxicity with small-brood size and low-egg hatchability.To study the underlying mechanism,we conduct a deep investigation of enrofloxacin(ENR),one of the most frequently detected FQs,on nematodes which is one of commonly used animal indicator of soil sustainability.The concentration-effect curves simulated by the Hill model showed that the half effect concentrations(EC50)of ENR were(494.3±272.9)μmol/kg and(107.4±30.9)μmol/kg for the brood size and the hatchability,respectively.Differential gene expression between the control and the ENR-exposure group enriched with the oxidative stress and cell apoptosis pathways.The results together with the enzyme activity in oxidative stress and the cell corpses suggested that ENR-induced reproductive toxicity was related to germ cell apoptosis under oxidative stress.The risk quotients of some soil and livestock samples were calculated based on the threshold value of EC10 for the egg hatchability(2.65μmol/kg).The results indicated that there was possible reproductive toxicity on the nematodes in certain agricultural soils for the FQs.This study suggested that chronic exposure to FQs at certain levels in environment would induce reproductive toxicity to the nematodes and might reduce the soil sustainability,alarming the environment risks of antibiotics abuse.

Genotoxic effects of microcystins mediated by nitric oxide and mitochondria

Microcystins are potent toxins,produced naturally by cyanobacteria（blue green algae）,and they present significant threats to human and animal health（WHO,1999;Chorus,2001;Carmichael et al.,2001;Falconer,2005;IARC,2010;Ma et al.,2015）.