The Toxicity Of Long-term Hexavalent Chromium Exposure On Marine Medaka(Oryzias Melastigma)

Recently,due to the rapid development of industrialization and urbanization in coastal area,a lagre amount of waste contained chromium were discharged into environment,which would lead to serious Cr pollution in coastal area and pose threats to marine organisms and human being.Because of the sensibility of marine medaka(Oryzias melastigma)to environmental pollutions,it was widely used in pollutant toxicological studies as the model species of marine ecotoxicological studies.In order to explore the effects of chromium on marine organisms,we chose 5-month marine medaka as experiment animals and 2.61 mg/L Cr(?)as exposure concentration to conduct a long-term exposure experiment to study the transcriptomic toxicity,DNA damage and transgenerational effects of Cr(VI).The detail results were as follow:Firstly,we studied the effects of marine medaka after exposure to 2.61 mg/L Cr(?)for three months and depurated for one month using RNA-seq.There were 3,065 differential expressed transcripts after Cr(?)long-term exposure,and 6,662 differential expressed transcripts after depuration significantly.After Cr(?)exposure and depuration,there were 6,627 transcripts with significant difference.We selected differential expressed genes annotated successful for subsequential analyses.We found that the toxicity of Cr(?)on marine medaka was mainly involved in the carcinogenic effect,because differential expressed genes were mostly enriched on the exosome of functional components and pathways related to cancer.Besides,the expression of actin genes such as F-ACTIN and ACTIN increased significantly.This is the first time that the cytoskeleton toxicity of Cr(?)was found at transcriptomic level in fish.The effects of depuration on the long-term toxicity of Cr(?)have two sides.On the one hand,depuration could attenuate the toxicity of Cr(?).Such as the significantly down-regulation of MAPK signaling pathway,the decrease of HSP expression and the restoration to normal expression levels of most genes related to cytoskeleton,like ACTIN,FILAMIN and ?-ACTININ.On the other hand,depuration may lead to stress reactions.For example,depuration may increase the sensitivity of body to intracellular residual Cr(?),and the expression of ESR2 gene was significantly up-regulated.This is the first time that depuration of pollutants in fishes was studied at the transcriptomic level.We found that the main mechanisms of depuration were involved in up-regulated ribosome pathway and down-regulated pathways related to cell connection,signaling pathways as well as cancer.We speculated that depuration may accelerate biosynthesis process of organisms to repair damage induced by Cr(?),and reduce the activity of various signal pathways to impede the process of carcinogenesis,then restore the body to normal physiological processes.Secondly,after exposure to Cr(?)for three months,oxidative stress was induced in marine medaka liver,which showed by the up-regulation of antioxidant enzyme genes like SOD,CAT and GPX.Oxidative stress is an important cause of DNA damage induced by Cr(?).Through the studies of key genes in the HR,NHEJ and BER pathways,we found that the long-term exposure of Cr(?)could cause obvious DNA damage in the liver of mainre medaka,including DNA double strand breaks,DNA single strand breaks and base oxidation.The up-regulation of XRCC4,LIG4 and PARP1 in the NHEJ pathway indicates that NHEJ pathway was the main repair pathway in DBSs repair process induced by Cr(?).The down-regulation of TB53BP in HR pathway indicates that DBSs may be biased towards HR pathway during repair.The up-regulation of MUYTH,APEX1 and PARP1 in BER pathway indicates that DNA repair was enhanced by oxidative damage.In addition,the up-regulation of TP53,APAF-1 and CASPASE-9 indicates that the apoptotic process was activated,but a slight down-regulation of CASPASE-3 suggests that the cells with serious DNA damage may escape the apoptosis process,which may lead to cancer.Finally,we found that Cr(?)exposure would greatly decrease the reproductive capacity of female parent,but had no obvious effect on fertilization rate.The transgenerational effects of Cr(?)on marine medaka was manifested in malformation,death,hatch,heart development,oxidative stress,inflammatory reaction and methylation,etc.After Cr(?)exposure,there was a reproductive compensation mechanism.The original reproduction mode with high energy consumption showing high fertility,high mortality,high deformity rate and low hatchability was turned into the later high efficiency reproduction mode showing low fertility,low mortality,low deformity rate and high hatch rate.We studied differences and similarities between the parental effects and effects from external environment of Cr(?).There are similarities including increase of malformiation and mortality,decline of hatch rate as well as increase of oxidative stress,but the parental effects were stronger than effects from external environment.At the same time,there are many differences,such as the offspring treatment would accelerate early heart development and hatch,but the parental effects are opposite,with more serious adverse effects.It is noteworthy that there was an interaction between parental and offspring treatment;and this interaction was particularly significant on mortality,heart rate,hatch rate,hatch time and hatch delay time.This interaction may lead to attenuation of parental effects on offspring when the offspring also underwent the same treatment.Such a phenomenon was particularly obvious at the late stage of exposure.We thought this was a kind of transgenerational adaptability.In addition,environmental adaptation has also been observed in the same generation to improve the fitness in the environment,showing the decrease of mortality and malformation rate,the increase of hatch rate,the shortening of hatch time as well as the restoration of heart rate for offspring at the late stage,whether offspring was in the blank or Cr(VI)environment.In this study,we innovatively studied the effects and mechanisms of Cr(VI)toxicity and depuration for long term on marine medaka from the transcriptional level.We found that the long-term exposure of Cr(VI)could cause DNA damage,activate the DNA repair pathway,and cause the carcinogenic effect.After depuration,the long-term toxicity of Cr(VI)would be alleviated,which indicated that Cr(VI)toxicity was reversible.The main molecular mechanisms of depuration was involved in the up-regulation of ribosome and the downregulation of intercellular connections pathways,signaling pathways as well as pathways related to cancer.At the same time,the transgenerational effects of toxicant was studied for the first time in a relatively long time dimension.We found that there were transgenerational effects of Cr(VI)on marine medaka,and transgenerational effects were much stronger than effects from external environment.During the long-term exposure process,there was adaptability between the same generation and the different generation as well as reproductive compensation mechanism.