| Silver?nanoparticles(Ag NPs)has been widely used in biotechnology and life sciences due to their excellent bactericidal properties.It has been found that Ag NPs can cause deformities in vertebrate embryos,but few studies have been conducted on the underlying molecular characteristics and mechanisms.Our studies have found that Ag NPs stressed zebrafish embryos exhibit developmental defects such as dysfunctional behaviors.In order to reveal the detailed molecular characteristics and the potential regulating mechanisms,we screened the transcriptome data for Ag NPs stressed embryos in this work firstly.In this study,we found that Ag NPs induced down-regulated expression in crystal structure-,neural circuit-and metabolism-related genes in zebrafish embryos,and their reduced expression was further verified by both in situ hybridization(whole mount in situ hybridization,WISH)and q RT-PCR(quantitative real-time polymerase chain reaction).Additionally,we unveiled that Ag NPs and their released silver ions(Ag~+)destroyed the formation of neural circuits,including motor neuron,neurotransmitter production and synaptic transmission,then,led to abnormal membrane potential of nerve cells and the resultant dysfunctional touch response and abnormal locomotor behavior.Moreover,this study unveiled that Ag NPs specifically induced the defects in the development of lens rather than retina,and analyzed the detailed molecular characteristics of Ag NPs induced metabolism abnormalities.The results of this study revealed the molecular characteristics and the potential regulatory mechanisms of the developmental deficits of the eye and nervous system of zebrafish embryos under Ag NPs stresses,as well as the down-regulation of their glycolysis and tyrosine metabolisms.The main results as following:1. Silver nanoparticles induce abnormal touch responses by damaging neural circuits in zebrafish embryosFirst,the abnormal touch response and neuronal membrane potential in Ag NPs-stressed embryos were found by touch stimulation and membrane potential detection in this study.Next,neurogenesis genes were unveiled to be down-regulated in Ag NPs stressed embryos by microarray technology and bioinformatics analysis.Additionally,the down-regulated expression of dopaminergic neurotransmitter genes otpa?sncgb?gad1b?gad2,synaptic transmission genes robo2?vim and glrbb,and motor neuron genes isl1 and isl2a was further identified in both Ag NPs and Ag+stressed embryos by q RT-PCR and whole-mount in situ hybridization(WISH).Moreover,this study unveiled that the reduced expression of gad1b?gad2 and isl1 could be recovered by adding Ag+chelating compound L-cysteine in Ag NPs stressed embryos,suggesting Ag NPs released Ag+mediated most of Ag NPs roles in inhibiting neural circuit formation.2. Silver nanoparticles affect lens rather than retina development in zebrafish embryosIn this study,we found that clefts and vacuoles were observed in lens of embryos from Ag NPs stressed group by microscopic observations and hematoxylin-eosin staining.Additionally,the down-regulated expressions of different lens crystallin isoform genes and the normal expression of retinal genes were observed in Ag NPs stressed embryos were unveiled by microarray technology,q RT-PCR and WISH.Moreover,no obvious cell apoptosis,and normal m RNA export was observed in Ag NPs stressed lens cells,suggesting Ag NPs induced lens developmental defects via other signaling rather than damaged m RNA export and induced cell apoptosis during embryogenesis.3. Metabolism responses to silver nanoparticles stresses during zebrafish embryogenesisMicroarray analysis unveiled the altered expression of metabolic genes in Ag NPs-stressed zebrafish embryos at 24 hpf(hour post fertilization).In this study,it was found that the down-regulated expression of the genes in glycolysis and tyrosine metabolic pathways in Ag NPs-stressed embryos at 24 hpf was followed by a compensatory increased expression in embryos at 72 hpf.Additionally,Using Elase kit,we found that decreased hexokinase&succinate de-hydrogenase activities in glycolysis pathway and high level of pyruvate were observed in the Ag NPs stressed embryos,which was similar to the metabolic molecular characters observed in progressive muscular dystrophy patients.Moreover,the altered glycolysis metabolism was assumed to influence the muscle fibrils maintenance via regulating the concentration of glycolysis metabolites rather than cell apoptosis and proliferation in trunk muscle cells.Furthermore,the altered tyrosine metabolic activities might be another contributor to touch response defects parallel to the defective neural circuit formation in Ag NPs-stressed embryos... |
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