Biological Mechanism Of EDCs On Fish Lipid Metabolism And Risk Assessment Of EDCs In Fish On Human Health

Environmental endocrine disrupting chemicals(EDCs)which have the hormone activities spread over environmental water bodies,and disturbing immune,endocrine and nervous systems.Fish act as the aquatic organism which are directly exposed to EDCs in water,and show greater adverse impacts than terrestrial animals.EDCs could damage the reproductive development,and even cause the production of sex reversal in vertebrates.At present,the feminization of reproductive characteristics has been found in male fish in some areas.Moreover,EDCs could also disturb energy metabolism in fish.Actually,fish metabolic diseases are very common in current aquaculture,especially excessive fat deposition including fatty liver and abdominal fat deposition.The metabolic diseases not only affect development and decrease tolerance in farmed fish,but also reduce the quality of aquatic products and increase the risk of food safety for consumers.However,studies which explore the effects of EDCs on lipid metabolism between fish sexual dimorphism are blank.Furthermore,EDCs could also induce cancer and malformation in human beings.Nevertheless,there is no study to expound the health risk of EDCs on consumer human via farmed fish.Therefore,this study aimed to explore the effects and molecule mechanism of EDCs on lipid metabolism between fish sexual dimorphism,and also assess human health risk of EDCs on via farmed fish.The biotechnologies,including lipid composition analysis,real-time RT-PCR,western bloting,transcriptomic,solid phase extraction(SPE),pressurized liquid extraction(PLE)and histology,were applied in this study.Firstly,zebrafish(Danio rerio)was chosen as the experimental animal,for exploring the effects and mechanism of EDCs which included 17β-estradiol(E2)and bisphenol A(BPA)on lipid metabolism between sexual dimorphism.Whereafter,the farmed fish large yellow croaker(Pseudosciaena crocea),Nile tilapia(Oreochromis niloticus)and yellow catfish(Pelteobagrus fulvidraco)were chosen as experimental animals,for assessing human health risk and tracking the major pathway of EDCs come into farmed fish,through detecting and analyzing EDCs contents in fish.The main results and conclusions in the present study were listed in the following: 1.The effects and biological mechanism of EDCs on lipid metabolism between fish sexual dimorphism.Before we explored the effects of E2 and BPA on lipid metabolism between fish sexual dimorphism,we should determine whether the differences of lipid metabolism between female and male fish were exist.Firstly,we fed female and male zebrafish with the diets contained 1%,7% and 13% lipid for 6 weeks,then we compared the lipid composition and metabolic regulation between female and male zebrafish.The results indicated that,contents of fish total fat and liver triglyceride(TG)in female zebrafish were significantly higher than those in male fed with the same diet.Moreover,the contents of fish total fat and liver TG in male fish among the treatments of three kinds of diets were compared,whereas the contents of fish total fat and liver TG in female fish among three treatments were dramatic differences,especially in the high-fat diet treatment.The results of hepatic transcriptomic analysis revealed that,lipid metabolic pathways between female and male fish were notably different.Genes related lipid synthesis were upregulated in female liver as compared with those in male fish,including acetyl-coenzyme A carboxylase(acc),sterol regulatory element binding transcription factor 1c(srebp-1c),and fatty acid synthase(fasn).These findings systematically testified the notable differences on lipid composition and lipid metabolic patterns between female and male fish.Basing on the differences of lipid metabolism between female and male fish,male zebrafish were chosen to carry out the concentration screening of E2 and BPA exposure.We fed male zebrafish with normal-fat diet,and exposed fish to gradient concentrations of E2 and BPA for 6 weeks.The treatments of E2 and BPA(200 ng/L,100 μg/L)were treated as the treatments of low concentrations,and E2 and BPA(2000 ng/L,2000μg/L)were treated as the treatments of high concentrations for exploring the effects of different concentrations of E2 and BPA on fish lipid metabolism.The E2 and BPA in all exposure treatments improved zebrafish growth,damaged the testis structure,inhibited sperm development,and induce the reproductive system in male fish to females.At the same time,the lipid composition and lipid metabolic pattern could also be altered in the treatments of low concentrations,which induce the contents of total fat,TG and free fatty acid(FFA)in fish liver as compared with the control and the treatments of high concentrations.These results indicated that,the effects of E2 and BPA on lipid metabolism were concentration-dependent in fish.The preliminary experiment revealed that E2(200ng/L)and BPA(100μg/L)markedly affected lipid metabolism in fish.Thus,we exposed female and male zebrafish to E2(200 ng/L)and BPA(100 μg/L)for 6 weeks for exploring the effects and biological mechanism of E2 and BPA on lipid metabolism between fish sexual dimorphism.At the same time,the treatments of estrogen receptor inhibitor tamoxifen(TAM)were added.The results indicated that,E2 and BPA mainly upregulated expression of genes related with lipogenesis in male fish,and improved the contents of fish total fat and liver TG,and then made the lipid contents in male close those in female.At the same time,the patterns of lipid metabolic regulations in male fish exposed to E2 and BPA,were notably different from those in male fish in the control,whereas were highly similar to those in female fish in the control.This result suggested that,lipid metabolic regulations in male exposed to E2 and BPA trended to feminization.The results of expression of proteins related to lipid metabolism revealed that,E2 and BPA inhibited 5’ AMP-activated protein kinase(AMPK)protein expression,then downregulated the expression of gene related to lipid catabolism in downstream of AMPK pathway.At the same time,inhibited AMPK also activated mammalian target of rapamycin(mTOR)and SREBP-1c pathway,then improved gene transcription and expression of fasn and acc.Moreover,E2 and BPA also could inhibit protein expression of signal transducer and activator of transcription 3(STAT3),then activate gene transcription of srebp-1c in the downstream.These results revealed that,E2 and BPA improved the lipid accumulation in male fish,and induced the lipid metabolic patterns from male fish to females,through inhibiting AMPK expression and activating the mTOR/SREBP-1c signaling pathway related to lipid synthesis.2.The assessment of EDCs in farmed fish on human health risk.Because daily fish supply mainly come from aquaculture,thus farmed fish were selected to assess human health risk of EDCs.Firstly,we assess the human health risk of E2 and BPA in natural water bodies,through farmed large yellow croaker.We fed large yellow croaker with two different energy-contained diets in the bay of Xiangshan in China for 8 weeks to establish two famed fish models with different lipid metabolic systems for exploring the effects of E2 and BPA on the growth and pollutant deposition in fish,through detecting total fat contents and E2 and BPA concentrations in fish.The results revealed that,fat contents in the whole fish,muscle and liver in the treatment of high energy-contained diet were notably higher than those in the treatment of low energy-contained diet.Concentrations of E2 in fish between the treatments of different diets were not revealed the significantly different.However,BPA concentrations in fish tended to increase in the treatment of high energy-contained diet as compared with the treatment of low energy-contained diet.Although,BPA levels in fish won’t produce harm to human health in this study.But,the human health risks of BPA in farmed fish in treatment of high energy-contained diet were higher as compared with those in the treatment of low energy-contained diet.It suggested that the composition of diets might affect the deposition of environmental pollutants in fish,and then affect human health risks.Basing on the results of above study,we aimed to investigate the effects of feed ingredients on EDCs deposition in farmed fish and the sedimentary differences of EDCs between diverse farmed fish.We captured a batch of anchovies from Bohai Sea and Yellow Sea in China,and send them to two factories which produce the crude fish oils,then send the two different kinds of crude fish oils to one refined fish oil factory to get the refined fish oil.Furthermore,three kinds of fish oils were used as the main lipid source in diets,and then produced three kinds of diets contained different fish oils: crude fish oil 1(CFO1),crude fish oil 2(CFO2)and refined fish oil(RFO).We fed the farmed fish Nile tilapia(lean fish)and yellow catfish(fatty fish)with the three kinds of diets for 8 weeks,and then detected the growth parameters,nutrient composition,the contents of EDCs,such as DDTs,PCBs and dioxin in farmed fish fillets.The results indicated that,the growth and nutrient composition were notably different between Nile tilapia and yellow catfish.The fat contents in yellow catfish were markedly greater than those in Nile tilapia,whereas the weight gain,feed efficiency and total protein contents were notably great in Nile tilapia as compared with those in yellow catfish.Moreover,the contents of DDTs,PCBs and dioxin were also greater in yellow catfish as compared with those in Nile tilapia,At the same time,EDCs in fillets of yellow catfish induced higher human health risk than those in Nile tilapia.These demonstrated that EDCs accumulation in farmed fish fillets notably depend on the fish species,and then affected human health risks.Through analyzing the patterns of EDC composition in water,feed ingredients,fish oils and farmed fish,we found that EDCs in farmed fish mainly came from fish oil.It indicated that,fish oils were the main vehicles of EDCs from ocean to farmed fish.Furthermore,the higher EDC contents were detected in the treatments of CFOs as compared with the treatments of RFO,it revealed that the processing technology of fish oils dramatically affected EDCs concentrations in farmed fish.The findings suggested that,through reducing the utilization of crude fish oils and improving the growth and feed efficiency in farmed fish,human health risk of EDCs in farmed fish could be notably decreased.Basing on the problems and challenges of aquaculture in China and the potential human health risk of fish consumers,the effects of environmental pollutants on fish metabolic diseases were further elucidated,and the basic data were provided for the accurate evaluation of current environmental health and aquatic product quality in the present study.Moreover,for the first time,the present study tracked the pathway of marine EDCs from ocean to human bodies,via fish oils in feed and farmed fish.It provided important references for guaranteeing the safety of aquatic products and guarantees for improving the health quality of fish consumers.