Toxicological Effects And Underlying Molecular Mechanisms Of TBBPA In Caenorhabditis Elegans

Tetrabromobisphenol A(TBBPA)is one of the most common brominated flame retardants used world widely.Owing to its extensive application and improper operation,TBBPA has been regularly detected in various environmental matrices such as water,soil,air,sediments,and even in human body.A number of studies have shown that TBBPA can cause endocrine disruption,neurotoxicity,reproductive toxicity and developmental immunotoxicity on aquatic organisms.However,the research about the effects of TBBPA exposure on invertebrates is limited,and lack of comprehensive mechanism studies.The nematode Caenorhabditis elegans(C.elegans)is an important model organism in environmental toxicology for evaluating environmental toxicants because of its short lifespan,transparency,302 wholly identified neurons,and 60?80%homologues with human genes.The main objective of current research was to explore the toxic effects and mechanism of C.elegans exposed to TBBPA from physiological,biochemical,molecular,and population levels.Including toxic effects of C.elegans chronic exposure to TBBPA,trans-generational effect of neurotoxicity in C.elegans exposed to TBBPA,neurotoxic effect and molecular mechanism of C.elegans subacute exposure to TBBPA,and ecotoxicity of TBBPA to C.elegans in simulate environmental exposure model.The main conclusions were as follows:(1)Chronic exposure to TBBPA could significantly inhabit the growth,locomotive behavior,and ingestive behavior of C.elegans.Locomotive behavior represented by body bends is the most sensitive indicator.After 3 days and 5 days of exposure at 1 g/L,significantly negative responses to body bends of C.elegans can be caused.Exposure to 0.1 g/L TBBPA for 10 days resulted in a significant reduction in the body bends of C.elegans.After 10 days of exposure,ROS level,SOD enzyme and CAT enzyme accumulation in nematodes gradually increased with the increasing of exposure concentration.Pearson correlation analysis showed that oxidative damage was one of the main way to cause long-term toxicity effect of nematodes.After 10 days exposure,the expression levels of 15 stress-related genes were significantly up-regulated.According to the testing results of GA186 and VC754 mutational strains nematodes,it is further proved that sod-3 and ctl-2 genes regulate the oxidative stress response of C.elegans.(2)The impacts of parental generation(Generation 1,G1)nematode acute exposure to TBBPA and their progeny(Generation 2,G2),which were both investigated to study the trans-generational effects under TBBPA-free conditions.TBBPA exposure at 1000 g/L would have significantly negative effects on the growth,locomotive behavior,and crawling movement behavior of nematodes in both generations.Among them,the most sensitive indicator was body bends,and 1 g/L and 100 g/L TBBPA can cause significantly negative effects on the body bends of G1 and G2 C.elegans respectively.Dopaminergic,y-aminobutyric acid and glutamatergic neurons,labeled by green fluorescent protein,revealed damages related to neurobehavioral endpoints in G1 worms with no hereditary effects in the progeny.The expression responses of oxidative stress-related indicators(ROS level,MDA content,SOD and CAT enzyme)and 20 stress-related genes were more significantly enhanced in G2 nematodes compared to G1 C.elegans.Combined with Pearson's correlation analysis,it was found that oxidative stress and stress-related genes were related to the neurotoxic effect and had intergenerational genetic effects.(3)Subacute exposure to TBBPA could significantly inhabit the growth,locomotive behavior,crawling movement behavior and ingestive behavior of exposed C.elegans.Among them,in contrast to what had been found from chronic exposure and trans-generational effects,the locomotive behavior represented by head thrashes was the most sensitive indicator of neural behavior and could cause significantly negative reactions at 1 g/L concentration.ROS content,CAT enzyme and SOD enzyme all accumulated significantly above 0.1 g/L dose.157 differentially expressed genes(Fold change>1.5,p-value<0.05)were found by using Illumina HiSeqTM 2000 sequencer.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway database analysis demonstrated that dorso-ventral axis formation pathway was related to neurotoxicity.Xenobiotics by Cytochrome P450 pathway and glutathione-S-transferase pathway were found to be the vital metabolic mechanisms.Gene cyp-13A7,cyp-13A10,cyp-13A12,cyp-13C1,gst-21,gst-33 and gst-44 were involved in above two metabolic pathways confirmed by quantitative real-time polymerase chain reaction.The VC40 mutational strain C.elegans demonstrated that the gene cyp-13a7 was responsible for the detoxification mechanism under subacute exposure to TBBPA.(4)The simulate environmental exposure model of TBBPA can cause significantly negative effects on physiological,biochemical,molecular and population levels of C.elegans.With the increase of TBBPA concentration,negative effects of growth,locomotive behavior,and ingestive behavior gradually intensified.Compared with the 5 days exposure,the locomotive behavior and ingestive behavior of nematodes after 10 days exposure recovered at low dose concentrations,showing an adaptive response.After 10 days exposure,ROS level and CAT enzymes in nematodes were significantly accumulated in the treatment group of 10 ? 100?g/L,and the expression levels of 20 stress-related genes were up-regulated to a certain extent.At the population level,10 ?g/L TBBPA exposure could resulted in significantly decrease in nematode populations.Combined with Pearson's correlation analysis,oxidative stress was an important cause to the toxic effects of C.elegans in the simulate environmental exposure model.In summary,the study found that TBBPA can cause reproductive and developmental toxicity,neurotoxicity and ecological toxicity effects on C.elegans.The locomotive behavior ability represented by head thrashes and body bends is the most sensitive indicator to TBBPA,and oxidative stress is an important cause to the physiological effect of C.elegans.The dorsal ventral axis formation pathway was mainly involved in the regulation of neural behavior in C.elegans,and the cytochrome P450 pathway and GST metabolic pathway were mainly responsible for the regulation of the detoxification mechanism.