Proteomic And Metabolomic Studies On The Toxicological Effects Of Cadmium And Arsenic In Juvenile Rockfish Sebastes Schlegelii

With the rapid development of the economy in the coastal areas and the establishment of a large number of heavy metal smelting,processing and electroplating enterprises,a large amount of industrial wastewater rich in heavy metals has been discharged into the ocean,resulting in the increasing pollution of trace metals in the coastal areas of China.Trace metals are nondegradable and can be easily accumulated in the environment for a long time,which in turn poses remarkable adverse effects to marine organisms and ecosystem.Based on the current status of trace metal pollution in offshore and coastal areas in China,we selected two typical heavy metal pollutants,cadmium(Cd)and arsenic(As(?)),for further exposure experiments.The juvenile rockfish Sebastes schlegelii was chosen as experimental animal for both Cd and As(?)exposures.In this study,two “-omic” approaches including nuclear magnetic resonance-based metabolomics and isobaric tags for relative and absolute quantitation-based proteomics were used to elucidate the toxicologicl effects of Cd and As(?)to juvenile rockfish S.schlegelii.Results could provide the theoretic basis for environmental risk assessment of trace metals.The major results were summarized as following:(1)Toxicological effects of Cd to rockfish S.schlegeliiAfter exposure to different concentrations of cadmium(5 and 50 ?g/L)for 14 days,the concentrations of Cd in juvenile rockfish showed a linear pattern with the exposure concentrations.The concentration in the juvenile rockfish of 5 ?g/L Cd-treated group was 2.75 times higher than that of the control group without statistical significance(P > 0.05),while the concentration in 50 ?g/L Cd-treated group was 9.5 times(P < 0.01)higher than that of the control group.Metabolomic analysis indicated that the metabolites,including lactate,phosphocholine,ATP,alanine and inosine were notably altered in Cd-treated groups.A total of 168 proteins showed significantly differential expressions in juvenile rockfish in Cd-treated groups.Ninety one differentially expressed proteins were indentified in 5 ?g/L Cd-treated group consisting of 52 increased and 39 decreased proteins.In 50 ?g/L Cd-treated group,there were 77 differentially expressed proteins,including 32 up-regulated and 45 down-regulated proteins.Forty eight proteins were commonly detected in both Cd-treated groups.On the whole,Cd could stimulate numerous molecular events in juvenile rockfish such as inhibiting glycolysis process and TCA cycle,for example,the expressions of glycolytic aldolase,glyceraldehyde 3-phosphate dehydrogenase,fumarate hydratase and malic enzyme were inhibited,promoting lipid,amino acid and protein metabolism,inducing immune and oxidative stress,disturbing the stability of the cytoskeleton,influencing the signal transduction,inducing the neurotoxicity,influencing gene expression and protein transport.In addition,peroxisome bifunctional enzymes and glyceraldehyde-3-phosphate dehydrogenase were preliminarily identified as key proteins in 5 and 50 ?g/L Cd-treated groups,respectively.These findings demonstrated that proteomic and metabolomic approaches could partially validate one another and their combination could provide a perceptive view into the influences of environmental metal pollution in marine fish at molecular levels.(2)Toxicological effects of As(?)to rockfish S.schlegeliiAfter exposure to different concentrations of arsenic(5 and 50 ?g/L)for 14 days,the concentrations of As(?)in juvenile rockfish showed a linear pattern with the exposure concentrations.The juvenile rockfish in 5 and 50 ?g/L As(?)-treated group exhibited significant(P < 0.05)As accumulations compared with control group.The total As contents in 5 and 50 ?g/L As(?)-treated group significantly increased by 16.1% and 19.5% compared with the control group,respectively(P < 0.05).The total As content in the high concentration of As(?)-exposed group was only 2.9% higher than that in the low concentration of As(?)-exposed group,without significant difference(P > 0.05).Metabolomic analysis demonstated that numerous metabolites were prominently altered in As(?)-treated groups,including lactate,alanine,ATP,inosine and phosphocholine.In total,one hundred and sixty three proteins showed significantly differential expressions in juvenile rockfish in As-treated groups.A total of 137 differentially expressed proteins were indentified in 5 ?g/L As-treated group consisting of 83 increased and 54 decreased proteins.There were 39 proteins differentially expressed in 50 ?g/L Cd-treated group,including 22 up-regulated and 17 down-regulated proteins.Only 13 proteins were commonly altered in both As treatments.Overall,As(V)inhibited the glycolysis process,promoted lipid metabolism,disturbed energy metabolism,promoted the oxidative phosphorylation activity,such as up regulating the expressions of cytochrome c oxidase subunit 4 isoform 1,NADH dehydrogenase(ubiquinone)flavoprotein 2 and cytochrome b-c1 complex subunit 6,induced oxidative stress and immune response,influenced gene expression and protein transport.Furthermore,expressions of anti-apoptotic protein and pro-apoptotic protein were also up regulated in 5 and 50 ?g/L As(?)-treated group,respectively.Additionally,glyceraldehyde-3-phosphate dehydrogenase,inosine 5'-phosphate dehydrogenase 1 and 40 S ribosomal protein SA were preliminarily identified as key proteins in 5 ?g/L As(?)-treated groups.