Toxicity And Mechanism Of Graphene Oxide On Zebrafish Embryogenesis

Due to their unique physiochemical properties,graphene oxide(GO)has shown great potential for biological,medical,energy,electronic and environmental applications.As a consequence of these diverse applications,GO release into the ecosystem is inevitable.Embryogenesis is an important phase of life,and dysfunctions in embryogenesis are associated with various diseases and adverse effects,such as hypoxia,malformation,organ function and eccyliosis.And the embryos is more susceptible to pollutant than the adult individuals.However,the corresponding risks are largely unknown,particularly with respect to the critical period of embryogenesis.Besides,the physicochemical property of GO may be changed due to interection with environmental factors and it not clear about the variation of their toxicology.In this study,the developmental toxicity induced by GO and its mechanism were determined using a multilevel approach,and the effects of humic acid(HA),pH and irradiation were taken into account.The methods and results are presented as following.After the zebrafish embryos were incubated in E3 solution containing GO,the developmental toxicity induced by GO were investigated using tissue section,ultrathin section,microexamination,fluorescent labeling and microelectrode.The results shown that the negative charge of GO were decreased and its stability were declined in E3 solution.GO nanosheets adsorbed on the chorion and impeded the gas exchange between embryo and environment which resulted in hypoxia in the chorional space and hatching delay.GO may enter into the chorional space via the pore canal or by the mechanical puncture.Then,GO distributed in yolk sac,eyes,heart and spine of the embryos via endocytosis and induced the malformation,such as eye developmental malformation,cardiac/yolk sac edema andtail flexure.Furthermore,GO damaged the mitochondria,prevented glutathione synthesis,produced excessive ROS,resulted in oxidative stress,lipid peroxidation,DNA damage and apoptosis.The bioinformatics of the embryos treated with GO were analysised using transcriptomics,proteomics and metabonomics.The results shown that GO leaded to up regulation of the genes and proteins that relative to stress,apoptosis,inflammation,immue,antioxidation,endocytosis and transcription,but down regulation of that relative to embryogenesis and metabolism.Furthemore,GO changed signaling parthways such as ECM-receptor interaction,Toll-like receptor signaling parthway and Fox O signaling parthwy,and disturbed the TCA,ornithine cycle,amino acid metabolism,nucleotide metabolism,glycometabolism and fatty acid metabolism.At the presence of HA,the physicochemical property and toxicity of GO were investigated.HA adsorbed GO via π-π interection which altered the shape,thinkness and stability of GO,reduced the interaction of GO with chorion,recovered the oxygen in the chorional space.Furthermore,HA reduced the mechanical puncture of GO to the chorions and decreased uptake by embryos.HA also altered the uptake and deposition of GO and decreased the aggregation of GO in embryonic yolk cells and deep layer cells.Moreover,HA mitigated the mitochondrial damage and oxidative stress induced by GO.GO exposed to irradiation for 28 days and the the physicochemical property and toxicity of the irradiated-GO were investigated.The roughness,sp2 C structure and sharp edge of GO were increased due to irradiation,but its oxygen-containing groups,surface negative charge and stability were decreased.These transformation prevented glutathione synthesis,produced excessive ROS,induced oxidative stress,lipid peroxidation,disturbed the TCA and ornithine cycle,and leaded to increase of its toxicity to zebrafish embryogenesis.GO exposed to acidic,neutral and basic environment for 20 days,respectively.Then the the physicochemical property and toxicity of the acid-treated GO were investigated.The sp2 C structure and disordered structure of GO increased with pH viriation from 3.0 to 11.0.After treated with acidic environment,the oxygen-containing groups,surface negative charge and stability decreased and prevented glutathione synthesis,produced excessive ROS,induced oxidative stress,lipid peroxidation,disturbed the TCA and ornithine cycle,and leaded to increase of its toxicity to zebrafish embryogenesis.