2种烷基多环芳烃对仿刺参CYP450和p53基因表达的影响研究
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摘要
海洋环境中多环芳烃类(PAHs)主要来源于海洋溢油事故以及沿海石油化工企业的废水排放,国内外大量研究发现海洋中的多环芳烃对海洋生物造成了潜在的生态风险。为了揭示不同浓度多环芳烃类污染物对海参的生态毒理效应,将仿刺参(Apostichopus japonicus)分别暴露于不同浓度的2种烷基多环芳烃3-甲基菲(5、10、100μg·L~(-1))和2-甲基蒽(5、10、50μg·L~(-1))中,检测暴露3 d、7 d和14 d后,3-甲基菲和2-甲基蒽胁迫下仿刺参CYP450和p53基因的相对表达量。结果表明,3-甲基菲和2-甲基蒽胁迫下,仿刺参CYP450和p53基因的表达均对毒物产生了不同程度的响应。与对照组相比,3-甲基菲各处理组对仿刺参CYP450和p53基因的表达均产生显著的抑制作用(P<0.05); 2-甲基蒽各处理组对仿刺参CYP450和p53基因的表达影响作用不同,暴露7 d后,2-甲基蒽各处理组对仿刺参CYP450基因的表达表现出抑制作用,对p53基因的表达表现出诱导作用。相同浓度与时间胁迫下,2-甲基蒽对仿刺参CYP450和p53基因表达的影响比3-甲基菲的影响大。上述研究结果表明,3-甲基菲和2-甲基蒽均可不同程度影响仿刺参CYP450和p53基因的表达,且与3-甲基菲相比,2-甲基蒽对仿刺参CYP450和p53基因表达的影响较明显。上述结果为多环芳烃类污染物对仿刺参的生物毒性评价提供了基础数据。
Marine organisms are facing a risk of exposure to a wide variety of toxic chemicals, such as polycyclic aromatic hydrocarbons(PAHs) from oil spill incidents and discharge of petrochemical wastewater. Besides, this kind of environmental pollutants pose a threat to the sea farming. As such, Apostichopus japonicus was exposed to 3-methylphenanthrene(5, 10 and 100 μg·L~(-1)) and 2-methylanthracene(5, 10 and 50 μg·L~(-1)) to study the toxic effects of alkyl-PAHs on the marine animals. Relative expression levels of CYP450 and p53 genes of Apostichopus japonicus under the stress of 3-methylphenanthrene and 2-methylanthracene were measured after 3 d, 7 d and 14 d. Results showed that 3-methylphenanthrene had significant inhibitory effect(P < 0. 05) on the expression of both CYP450 and p53 genes, while 2-methylanthracene had a different effect on the expression of CYP450 and p53 genes. From 7 thday onwards, CYP450 gene expression was inhibited by 2-methylanthracene, but p53 gene expression was induced. Meanwhile, Apostichopus japonicus were more sensitive to 2-methylanthracene than 3-methylphenanthrene within the same concentration range and exposure duration. These results indicated that 3-methylphenanthrene and 2-methylanthracene could affect the expression of CYP450 and p53 genes of Apostichopus japonicus, and compared with 3-methylphenanthrene, 2-methylanthracene has significant effect on the expression of CYP450 and p53 genes. These results provide the basic data for the biotoxicity evaluation of PAHs pollutants on Apostichopus japonicus.
Marine organisms are facing a risk of exposure to a wide variety of toxic chemicals, such as polycyclic aromatic hydrocarbons(PAHs) from oil spill incidents and discharge of petrochemical wastewater. Besides, this kind of environmental pollutants pose a threat to the sea farming. As such, Apostichopus japonicus was exposed to 3-methylphenanthrene(5, 10 and 100 μg·L~(-1)) and 2-methylanthracene(5, 10 and 50 μg·L~(-1)) to study the toxic effects of alkyl-PAHs on the marine animals. Relative expression levels of CYP450 and p53 genes of Apostichopus japonicus under the stress of 3-methylphenanthrene and 2-methylanthracene were measured after 3 d, 7 d and 14 d. Results showed that 3-methylphenanthrene had significant inhibitory effect(P < 0. 05) on the expression of both CYP450 and p53 genes, while 2-methylanthracene had a different effect on the expression of CYP450 and p53 genes. From 7 thday onwards, CYP450 gene expression was inhibited by 2-methylanthracene, but p53 gene expression was induced. Meanwhile, Apostichopus japonicus were more sensitive to 2-methylanthracene than 3-methylphenanthrene within the same concentration range and exposure duration. These results indicated that 3-methylphenanthrene and 2-methylanthracene could affect the expression of CYP450 and p53 genes of Apostichopus japonicus, and compared with 3-methylphenanthrene, 2-methylanthracene has significant effect on the expression of CYP450 and p53 genes. These results provide the basic data for the biotoxicity evaluation of PAHs pollutants on Apostichopus japonicus.
引文
[1]Mu J L,Wang J Y,Jin F,et al.Comparative embryotoxicity of phenanthrene and alkyl-phenanthrene to marine medaka(Oryzias melastigma)[J].Marine Pollution Bulletin,2014,85(2):505-515
[2]靳非,穆景利,王冬婉,等.菲、3-甲基菲和菲醌对河鲀(Takifugu rubripes)幼鱼肝组织损伤的比较研究[J].生态毒理学报,2016,11(1):248-253Jin F,Mu J L,Wang D W,et al.The comparative research of phenanthrene,3-methyl phenanthrene and phenanthraquinone on the liver tissue injury of pufferfish(Takifugu rubripes)larvae[J].Asian Journal of Ecotoxicology,2016,11(1):248-253(in Chinese)
[3]关晓燕,王摆,董颖,等.二甲苯、蒽、苯并[α]芘对虾夷扇贝幼贝血清SOD、CAT活性的影响[J].生态毒理学报,2016,11(1):289-294Guan X Y,Wang B,Dong Y,et al.Effects of xylene,anthracene,and benzo[α]pyrene on enzymatic activities of SOD and CAT in the Japanese scallop(Mizuhopecten yessoensis)serums[J].Asian Journal of Ecotoxicology,2016,11(1):289-294(in Chinese)
[4]张薇,宋玉芳,孙铁珩.多环芳烃菲污染土壤的蚯蚓细胞色素P450生物标记诊断[J].毒理学杂志,2005(S1):311-312Zhang W,Song Y F,Sun T H.Diagnosis of sputum cytochrome P450 biomarkers in polycyclic aromatic hydrocarbon phenanthrene contaminated soil[J].Journal of Toxicology,2005(S1):311-312(in Chinese)
[5]张倩倩,乔敏.五氯酚对土壤跳虫代谢转化酶基因和蜕皮相关基因表达的影响[J].生态毒理学报,2017,12(5):72-78Zhang Q Q,Qiao M.Effects of pentachlorophenol on expression of metabolic enzyme genes and molt-related genes of soil Collembolan[J].Asian Journal of Ecotoxicology,2017,12(5):72-78(in Chinese)
[6]顾颖,李延,陈蔚丰,等.斑马鱼p53基因结构比对分析及其在生态毒理学中的应用[J].生态毒理学报,2006,1(1):45-51Gu Y,Li Y,Chen W F,et al.Structure alignments of zebrafish p53 gene and its application in ecotoxicology[J].Asian Journal of Ecotoxicology,2006,1(1):45-51(in Chinese)
[7]李春杰,赵建,张世勇,等.细胞色素P450 CYP3A65斑马鱼模型建立及对环境污染物的生物响应[J].中国药理学与毒理学杂志,2014,28(6):870-877Li C J,Zhao J,Zhang S Y,et al.Generation of cytochrome P-450 CYP3A65 labeled fluorescence transgenic zebrafish and its biological response to environmental pollutants[J].Chinese Journal of Pharmacology and Toxicology,2014,28(6):870-877(in Chinese)
[8]赵春,蒋美琳,周新初,等.蚯蚓细胞色素P450基因的克隆及序列分析[J].上海交通大学学报:农业科学版,2015,33(4):33-37Zhao C,Jiang M L,Zhou X C,et al.Clone and sequence analysis of cytochrome P450 Gene in earthworm[J].Journal of Shanghai Jiaotong University:Agricultural Science,2015,33(4):33-37(in Chinese)
[9]Rasmussen K,Klausen L,Ekstrand B.Regulation of cytochrome P450 m RNA expression in primary porcine hepatocytes by selected secondary plant metabolites from chicory(Cichorium intybus L)[J].Food Chemistry,2014,146:255-263
[10]刘超,吴富村,林思恒,等.高温刺激导致虾夷扇贝死亡因素的探究[J].海洋科学,2016,40(11):91-98Liu C,Wu F C,Lin S H,et al.Pilot study on reasons for yesso scallop’s(Patinopecten yessoensis)survival after heat shock[J].Marine Sciences,2016,40(11):91-98(in Chinese)
[11]刘晓威,姜森颢,周一兵,等.大连地区仿刺参养殖池塘底栖硅藻生产状况的周年变化研究[J].水产科学,2012,31(11):679-682Liu X W,Jiang S H,Zhou Y B,et al.Annual changes in production of benthic diatoms in sea cucumber Apostichopus japonicus culture ponds in Dalian[J].Fisheries Science,2012,31(11):679-682(in Chinese)
[12]高士博,王摆,董颖,等.苯系物对仿刺参catalase基因表达及酶活性的影响[J].生态毒理学报,2015,10(2):297-305Gao S B,Wang B,Dong Y,et al.The effects of BTEXs on the catalase gene expression and catalase activities in respiratory tree and intestine of sea cucumber Apostichopus japonicus[J].Asian Journal of Ecotoxicology,2015,10(2):297-305(in Chinese)
[13]林芳,毛楷林,江秀,等.Ba P和DDT暴露对翡翠贻贝胚胎重要酶活性影响的比较研究[J].生态毒理学报,2018,13(3):129-137Lin F,Mao K L,Jiang X,et al.Comparative studies of the critical enzymes activity of Perna viridis embryo exposed to Ba P and DDT[J].Asian Journal of Ecotoxicology,2018,13(3):129-137(in Chinese)
[14]Jennings P,Limonciel A,Felice L,et al.An overview of transcriptional regulation in response to toxicological insult[J].Archives of Toxicology,2012,87(1):49-72
[15]欧瑞康,武小燕,库培佳,等.食蚊鱼(Gambusia afinis)cat、gapdh和gst基因的克隆及其在生态毒理学中的应用[J].生态毒理学报,2015,10(3):83-92Ou R K,Wu X Y,Ku P J,et al.Cloning of cat,gapdh and gst genes of Gambusia affinis and its application in ecotoxicology[J].Asian Journal of Ecotoxicology,2015,10(3):83-92(in Chinese)
[16]Livingstone D R.Oxidative stress in aquatic organisms in relation to pollution and aquaculture[J].Revue de Medecine Veterinaire,2003,154(6):427-430
[17]Richardson B J,Mak E,De Luca-Abbott S B,et al.Antioxidant responses to polycyclic aromatic hydrocarbons and organochlorine pesticides in green-lipped mussels(Perna viridis):Do mussels“integrate”biomarker responses?[J].Marine Pollution Bulletin,2008,57(6):503-514
[18]Gravato C,Almeida J R,Silva C,et al.Using a multibiomarker approach and behavioural responses to assess the effects of anthracene in Palaemon serratus[J].Aquatic Toxicology,2014,149(8):94-102
[19]Apel K,Hirt H.Reactive oxygen species:Metabolism,oxidative stress,and signal transduction[J].Annual Review of Plant Biology,2014,55(1):728-749
[20]宋玉芳,周启星,许华夏,等.菲、芘、1,2,4-三氯苯对蚯蚓的急性毒性效应[J].农村生态环境,2003(1):36-39Song Y F,Zhou Q X,Xu H X,et al.Effect of acute toxicity of phenanthrene,pyrene and 1,2,4-trichlorobenzene on earthworms in soils[J].Rural Eco-Environment,2003(1):36-39(in Chinese)
[21]谢丹君,徐晶莹,诸葛淳淳,等.多环芳烃类物质对细胞色素P450 1A1的诱导表达[J].同济大学学报:医学版,2013,34(2):1-5Xie D J,Xu J Y,Zhuge C C,et al.Induction of cytochrome P450 1A1 expression in A549 cells by polycyclic aromatic hydrocarbons[J].Journal of Tongji University:Medical Science,2013,34(2):1-5(in Chinese)
[22]Eggen R I L,Behra R,Burkhardt-holm P,et al.Challenges in ecotoxicology[J].Environmental Science&Technology,2004,38(3):58A-64A
[23]刘林,赵群芬,朱帅旗,等.纳米氧化锌对斑马鱼肠组织的氧化损伤[J].水产学报,2015,39(11):1702-1711Liu L,Zhao Q F,Zhu S Q,et al.Oxidative damage of zinc oxide nanoparticles to zebrafish intestine[J].Journal of Fisheries of China,2015,39(11):1702-1711(in Chinese)
[24]Yu C,Zhou Z,Wang J,et al.In depth analysis of apoptosis induced by silica coated manganese oxide nanoparticles in vitro[J].Journal of Hazardous Materials,2015,283:519-528
[2]靳非,穆景利,王冬婉,等.菲、3-甲基菲和菲醌对河鲀(Takifugu rubripes)幼鱼肝组织损伤的比较研究[J].生态毒理学报,2016,11(1):248-253Jin F,Mu J L,Wang D W,et al.The comparative research of phenanthrene,3-methyl phenanthrene and phenanthraquinone on the liver tissue injury of pufferfish(Takifugu rubripes)larvae[J].Asian Journal of Ecotoxicology,2016,11(1):248-253(in Chinese)
[3]关晓燕,王摆,董颖,等.二甲苯、蒽、苯并[α]芘对虾夷扇贝幼贝血清SOD、CAT活性的影响[J].生态毒理学报,2016,11(1):289-294Guan X Y,Wang B,Dong Y,et al.Effects of xylene,anthracene,and benzo[α]pyrene on enzymatic activities of SOD and CAT in the Japanese scallop(Mizuhopecten yessoensis)serums[J].Asian Journal of Ecotoxicology,2016,11(1):289-294(in Chinese)
[4]张薇,宋玉芳,孙铁珩.多环芳烃菲污染土壤的蚯蚓细胞色素P450生物标记诊断[J].毒理学杂志,2005(S1):311-312Zhang W,Song Y F,Sun T H.Diagnosis of sputum cytochrome P450 biomarkers in polycyclic aromatic hydrocarbon phenanthrene contaminated soil[J].Journal of Toxicology,2005(S1):311-312(in Chinese)
[5]张倩倩,乔敏.五氯酚对土壤跳虫代谢转化酶基因和蜕皮相关基因表达的影响[J].生态毒理学报,2017,12(5):72-78Zhang Q Q,Qiao M.Effects of pentachlorophenol on expression of metabolic enzyme genes and molt-related genes of soil Collembolan[J].Asian Journal of Ecotoxicology,2017,12(5):72-78(in Chinese)
[6]顾颖,李延,陈蔚丰,等.斑马鱼p53基因结构比对分析及其在生态毒理学中的应用[J].生态毒理学报,2006,1(1):45-51Gu Y,Li Y,Chen W F,et al.Structure alignments of zebrafish p53 gene and its application in ecotoxicology[J].Asian Journal of Ecotoxicology,2006,1(1):45-51(in Chinese)
[7]李春杰,赵建,张世勇,等.细胞色素P450 CYP3A65斑马鱼模型建立及对环境污染物的生物响应[J].中国药理学与毒理学杂志,2014,28(6):870-877Li C J,Zhao J,Zhang S Y,et al.Generation of cytochrome P-450 CYP3A65 labeled fluorescence transgenic zebrafish and its biological response to environmental pollutants[J].Chinese Journal of Pharmacology and Toxicology,2014,28(6):870-877(in Chinese)
[8]赵春,蒋美琳,周新初,等.蚯蚓细胞色素P450基因的克隆及序列分析[J].上海交通大学学报:农业科学版,2015,33(4):33-37Zhao C,Jiang M L,Zhou X C,et al.Clone and sequence analysis of cytochrome P450 Gene in earthworm[J].Journal of Shanghai Jiaotong University:Agricultural Science,2015,33(4):33-37(in Chinese)
[9]Rasmussen K,Klausen L,Ekstrand B.Regulation of cytochrome P450 m RNA expression in primary porcine hepatocytes by selected secondary plant metabolites from chicory(Cichorium intybus L)[J].Food Chemistry,2014,146:255-263
[10]刘超,吴富村,林思恒,等.高温刺激导致虾夷扇贝死亡因素的探究[J].海洋科学,2016,40(11):91-98Liu C,Wu F C,Lin S H,et al.Pilot study on reasons for yesso scallop’s(Patinopecten yessoensis)survival after heat shock[J].Marine Sciences,2016,40(11):91-98(in Chinese)
[11]刘晓威,姜森颢,周一兵,等.大连地区仿刺参养殖池塘底栖硅藻生产状况的周年变化研究[J].水产科学,2012,31(11):679-682Liu X W,Jiang S H,Zhou Y B,et al.Annual changes in production of benthic diatoms in sea cucumber Apostichopus japonicus culture ponds in Dalian[J].Fisheries Science,2012,31(11):679-682(in Chinese)
[12]高士博,王摆,董颖,等.苯系物对仿刺参catalase基因表达及酶活性的影响[J].生态毒理学报,2015,10(2):297-305Gao S B,Wang B,Dong Y,et al.The effects of BTEXs on the catalase gene expression and catalase activities in respiratory tree and intestine of sea cucumber Apostichopus japonicus[J].Asian Journal of Ecotoxicology,2015,10(2):297-305(in Chinese)
[13]林芳,毛楷林,江秀,等.Ba P和DDT暴露对翡翠贻贝胚胎重要酶活性影响的比较研究[J].生态毒理学报,2018,13(3):129-137Lin F,Mao K L,Jiang X,et al.Comparative studies of the critical enzymes activity of Perna viridis embryo exposed to Ba P and DDT[J].Asian Journal of Ecotoxicology,2018,13(3):129-137(in Chinese)
[14]Jennings P,Limonciel A,Felice L,et al.An overview of transcriptional regulation in response to toxicological insult[J].Archives of Toxicology,2012,87(1):49-72
[15]欧瑞康,武小燕,库培佳,等.食蚊鱼(Gambusia afinis)cat、gapdh和gst基因的克隆及其在生态毒理学中的应用[J].生态毒理学报,2015,10(3):83-92Ou R K,Wu X Y,Ku P J,et al.Cloning of cat,gapdh and gst genes of Gambusia affinis and its application in ecotoxicology[J].Asian Journal of Ecotoxicology,2015,10(3):83-92(in Chinese)
[16]Livingstone D R.Oxidative stress in aquatic organisms in relation to pollution and aquaculture[J].Revue de Medecine Veterinaire,2003,154(6):427-430
[17]Richardson B J,Mak E,De Luca-Abbott S B,et al.Antioxidant responses to polycyclic aromatic hydrocarbons and organochlorine pesticides in green-lipped mussels(Perna viridis):Do mussels“integrate”biomarker responses?[J].Marine Pollution Bulletin,2008,57(6):503-514
[18]Gravato C,Almeida J R,Silva C,et al.Using a multibiomarker approach and behavioural responses to assess the effects of anthracene in Palaemon serratus[J].Aquatic Toxicology,2014,149(8):94-102
[19]Apel K,Hirt H.Reactive oxygen species:Metabolism,oxidative stress,and signal transduction[J].Annual Review of Plant Biology,2014,55(1):728-749
[20]宋玉芳,周启星,许华夏,等.菲、芘、1,2,4-三氯苯对蚯蚓的急性毒性效应[J].农村生态环境,2003(1):36-39Song Y F,Zhou Q X,Xu H X,et al.Effect of acute toxicity of phenanthrene,pyrene and 1,2,4-trichlorobenzene on earthworms in soils[J].Rural Eco-Environment,2003(1):36-39(in Chinese)
[21]谢丹君,徐晶莹,诸葛淳淳,等.多环芳烃类物质对细胞色素P450 1A1的诱导表达[J].同济大学学报:医学版,2013,34(2):1-5Xie D J,Xu J Y,Zhuge C C,et al.Induction of cytochrome P450 1A1 expression in A549 cells by polycyclic aromatic hydrocarbons[J].Journal of Tongji University:Medical Science,2013,34(2):1-5(in Chinese)
[22]Eggen R I L,Behra R,Burkhardt-holm P,et al.Challenges in ecotoxicology[J].Environmental Science&Technology,2004,38(3):58A-64A
[23]刘林,赵群芬,朱帅旗,等.纳米氧化锌对斑马鱼肠组织的氧化损伤[J].水产学报,2015,39(11):1702-1711Liu L,Zhao Q F,Zhu S Q,et al.Oxidative damage of zinc oxide nanoparticles to zebrafish intestine[J].Journal of Fisheries of China,2015,39(11):1702-1711(in Chinese)
[24]Yu C,Zhou Z,Wang J,et al.In depth analysis of apoptosis induced by silica coated manganese oxide nanoparticles in vitro[J].Journal of Hazardous Materials,2015,283:519-528