纳米二氧化硅对16HBE细胞存活率及PARP-1表达的影响
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摘要
目的探讨纳米二氧化硅(SiO_2)对人支气管上皮细胞(16HBE细胞)存活率和聚二磷酸腺苷核糖聚合酶-1(PARP-1)表达的影响。方法 (1)以质量浓度为0~100 mg/L纳米SiO_2处理16HBE细胞24.0 h,以CCK-8法检测细胞存活率。(2)将16HBE细胞分为6组:溶剂对照组(予等体积溶剂处理)、微米SiO_2对照组(予终质量浓度为20 mg/L微米SiO_2处理),5、10、20 mg/L纳米SiO_2组(予相应终质量浓度的纳米SiO_2处理),姜黄素组(先予终浓度为10μmol/L的姜黄素处理2.0 h,再予终质量浓度为20 mg/L的纳米SiO_2处理)。各组细胞经处理后,分别于培养4.0、12.0和24.0 h时间点收获细胞。采用荧光实时定量聚合酶链式反应检测细胞中PARP-1 mRNA的相对表达水平,以免疫印迹法检测PARP-1蛋白的相对表达水平。结果 (1)随着纳米SiO_2处理剂量的增加,细胞存活率下降,呈剂量-效应关系(P<0.01)。(2)在12.0和24.0 h时间点,纳米SiO_2刺激后,16HBE细胞的PARP-1 mRNA和蛋白相对表达水平均出现剂量依赖性下降(P<0.01);与同时间点溶剂对照组比较,5、10、20 mg/L纳米SiO_2组16HBE细胞在该2个时间点的PARP-1 mRNA和蛋白相对表达水平均下降(P<0.05)。在12.0和24.0 h时间点,20 mg/L纳米SiO_2组16HBE细胞的PARP-1 mRNA和蛋白相对表达水平均低于同时间点的微米SiO_2对照组(P<0.05);姜黄素组16HBE细胞的12.0 h时间点上述2个指标均高于20 mg/L纳米SiO_2组(P<0.05)。结论纳米SiO_2刺激可导致16HBE细胞存活率下降并呈剂量依赖性;PARP-1表达的下调可能是纳米SiO_2致16HBE增殖抑制的机制之一。姜黄素对纳米SiO_2诱导的16HBE的细胞损伤具有一定的保护作用。
Objective To explore the effects of nano-silicon dioxide( SiO_2) on the survival and poly( ADP-ribose)polymerase-1( PARP-1) expression in human bronchial epithelial cells( 16 HBE cells). Methods i) The 16 HBE cells were treated with nano-SiO_2 at concentrations ranging from 0 to 100 mg/L for 24. 0 hours,and CCK-8 assay was used to examine cell viability. ii) The 16 HBE cells were divided into 6 groups: solvent control group( equal volume solvent treatment),micro-SiO_2 control group( treated with 20 mg/L micro-SiO_2),5,10,and 20 mg/L nano-SiO_2 groups( treated with the corresponding final dose of nano-SiO_2),and curcumin group. The curcumin group was given pretreatment with curcumin at a final concentration of 10 μmol/L for 2. 0 hours followed by treatment with a final concentration of 20 mg/L of nano-SiO_2. Cells in each group were harvested at time points of 4. 0,12. 0 and 24. 0 hours after treatment. The relative expression of PARP-1 mRNA and protein in 16 HBE cells was detected by quantitative real-time polymerase chain reaction and Western blotting respectively. Results i) The survival of 16 HBE cells decreased with increasing nano-SiO_2 treatment dose,showing a dose-effect relationship( P < 0. 01). ii) The expression of PARP-1 mRNA and protein in 16 HBE cells were dose-dependently decreased after nano-SiO_2 stimulation at the 12. 0 and 24. 0 hours time points( P < 0. 01). The expression of PARP-1 mRNA and protein in 5,10,and 20 mg/L nano-SiO_2 groups decreased at the above mentioned time points( P < 0. 05),compared with the solvent control group at the same time points. The expression of PARP-1 mRNA and protein in 20 mg/L nano-SiO_2 group was lower than that in the micro-SiO_2 control group at the same 12. 0 and 24. 0 hours time point( P < 0. 05). The above two indexes of cells were higher in curcumin group than that of 20 mg/L nano-SiO_2 group at the 12. 0 hours time point( P < 0. 05). Conclusion Nano-SiO_2 stimulation can lead to decrease survival of 16 HBE cells in a dose-dependent manner and down-regulation of PARP-1 expression may be one of the mechanisms of proliferation and inhibition of 16 HBE cells induced by nano-SiO_2. Curcumin has certain protective effect on nano-SiO_2-induced 16 HBE cell injury.
Objective To explore the effects of nano-silicon dioxide( SiO_2) on the survival and poly( ADP-ribose)polymerase-1( PARP-1) expression in human bronchial epithelial cells( 16 HBE cells). Methods i) The 16 HBE cells were treated with nano-SiO_2 at concentrations ranging from 0 to 100 mg/L for 24. 0 hours,and CCK-8 assay was used to examine cell viability. ii) The 16 HBE cells were divided into 6 groups: solvent control group( equal volume solvent treatment),micro-SiO_2 control group( treated with 20 mg/L micro-SiO_2),5,10,and 20 mg/L nano-SiO_2 groups( treated with the corresponding final dose of nano-SiO_2),and curcumin group. The curcumin group was given pretreatment with curcumin at a final concentration of 10 μmol/L for 2. 0 hours followed by treatment with a final concentration of 20 mg/L of nano-SiO_2. Cells in each group were harvested at time points of 4. 0,12. 0 and 24. 0 hours after treatment. The relative expression of PARP-1 mRNA and protein in 16 HBE cells was detected by quantitative real-time polymerase chain reaction and Western blotting respectively. Results i) The survival of 16 HBE cells decreased with increasing nano-SiO_2 treatment dose,showing a dose-effect relationship( P < 0. 01). ii) The expression of PARP-1 mRNA and protein in 16 HBE cells were dose-dependently decreased after nano-SiO_2 stimulation at the 12. 0 and 24. 0 hours time points( P < 0. 01). The expression of PARP-1 mRNA and protein in 5,10,and 20 mg/L nano-SiO_2 groups decreased at the above mentioned time points( P < 0. 05),compared with the solvent control group at the same time points. The expression of PARP-1 mRNA and protein in 20 mg/L nano-SiO_2 group was lower than that in the micro-SiO_2 control group at the same 12. 0 and 24. 0 hours time point( P < 0. 05). The above two indexes of cells were higher in curcumin group than that of 20 mg/L nano-SiO_2 group at the 12. 0 hours time point( P < 0. 05). Conclusion Nano-SiO_2 stimulation can lead to decrease survival of 16 HBE cells in a dose-dependent manner and down-regulation of PARP-1 expression may be one of the mechanisms of proliferation and inhibition of 16 HBE cells induced by nano-SiO_2. Curcumin has certain protective effect on nano-SiO_2-induced 16 HBE cell injury.
引文
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[18]焦常平,叶亚婧,张渊,等.纳米二氧化硅诱导A549细胞损伤及其氧化应激机制[J].中国药理学与毒理学杂志,2013,27(6):995-999.
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[20]THOMAS C,KOTOVA E,TULIN A V.PARP-1 Interaction with and Activation by Histones and Nucleosomes[J].Methods Mol Biol,2017,1608:255-267.
[21]MANGERICH A,BURKLE A.Pleiotropic cellular functions of PARP1 in longevity and aging:genome maintenance meets inflammation[J].Oxid Med Cell Longev,2012:321653.
[22]LI H,LI Q,LI W,et al.The Role of PARP-1 in Host-Pathogen Interaction and Cellular Stress Responses[J].Crit Rev Eukaryot Gene Expr,2015,25(2):175-190.
[23]ZHANG N,ZHANG Y,ZHAO S,et al.Septin4 as a novel binding partner of PARP1 contributes to oxidative stress induced human umbilical vein endothelial cells injure[J].Biochem Biophys Res Commun,2018,496(2):621-627.
[24]GONG C,TAO G,YANG L,et al.Methylation of PARP-1 promoter involved in the regulation of nano-Si O2-induced decrease of PARP-1mRNA expression[J].Toxicol Lett,2012,209(3):264-269.
[25]WANG F,JIAO C,LIU J,et al.Oxidative mechanisms contribute to nanosize silican dioxide-induced developmental neurotoxicity in PC12cells[J].Toxicol In Vitro,2011,25(8):1548-1556.
[26]CALKINS A S,IGLEHART J D,LAZARO J B.DNA damageinduced inhibition of rRNA synthesis by DNA-PK and PARP-1[J].Nucleic Acids Res,2013,41(15):7378-7386.
[27]赵光强,黄云超,李光剑,等.纳米二氧化硅在人支气管上皮细胞内的亚细胞分布和遗传毒性[J].中国肺癌杂志,2013,16(3):117-124.
[28]洪文旭,陈晓丹,彭圣明,等.姜黄素对纳米二氧化硅诱导Ha Ca T细胞氧化应激损伤保护作用研究[J].热带医学杂志,2013,13(9):1061-1064.
[29]洪文旭,周桂凤,胡韬,等.姜黄素对纳米二氧化硅诱导Ha Ca T细胞氧化损伤的保护作用[J].毒理学杂志,2013,27(6):412-415.
[30]GONG C,YANG L,ZHOU J,et al.Possible role of PAPR-1 in protecting human Ha Ca T cells against cytotoxicity of Si O2nanoparticles[J].Toxicol Lett,2017,280:213-221.
[2]MEBERT A M,BAGLOLE C J,DESIMONE M F,et al.Nanoengineered silica:Properties,applications and toxicity[J].Food Chem Toxicol,2017,109(Pt 1):753-770.
[3]BAI P.Biology of poly(ADP-Ribose)polymerases:the factotums of cell maintenance[J].Mol Cell,2015,58(6):947-958.
[4]JUBIN T,KADAM A,JARIWALA M,et al.The PARP family:insights into functional aspects of poly(ADP-ribose)polymerase-1 in cell growth and survival[J].Cell Prolif,2016,49(4):421-437.
[5]NAVARRO J,GOZALBO-LOPEZ B,MéNDEZ A C,et al.PARP-1/PARP-2 double deficiency in mouse T cells results in faulty immune responses and T lymphomas[J].Sci Rep,2017,7:41962.doi:10.1038/srep41962.
[6]李杰,胡道荣,徐世田.姜黄素对脂多糖诱导的HK2细胞炎症反应和生长抑制的保护作用[J].热带医学杂志,2017,17(5):612-615.
[7]余俊青,姚广裕,胡晓磊,等.姜黄素通过降低MT1-MMP表达抑制乳腺癌细胞的增殖与侵袭[J].实用医学杂志,2017,33(9):1394-1396.
[8]HUANG Y,CAO S,ZHANG Q,et al.Biological and pharmacological effects of hexahydrocurcumin,a metabolite of curcumin[J].Arch Biochem Biophys,201,646:31-37.
[9]耿维佳,李阳,于永波,等.N-乙酰半胱氨酸对纳米二氧化硅所致细胞毒性的抑制作用[J].吉林大学学报(医学版),2015,41(3):486-490.
[10]GONG C,TAO G,YANG L,et al.The role of reactive oxygen species in silicon dioxide nanoparticle-induced cytotoxicity and DNA damage in Ha Ca T cells[J].Mol Biol Rep,2012,39(4):4915-4925.
[11]戴春,黄云超,周永春.纳米二氧化硅的肺毒性的研究进展[J].中国肺癌杂志,2014,(10):760-764.
[12]YANG H,WU Q Y,LI M Y,et al.Pulmonary toxicity in rats caused by exposure to intratracheal instillation of Si O2nanoparticles[J].Biomed Environ Sci.2017,30(4):264-279.
[13]李明月,劳灿山,李文超,等.miR-702-3p在纳米二氧化硅致大鼠肺纤维化中作用的初步探讨[J].航天医学与医学工程,2016,29(6):403-407.
[14]彭子荷,魏艳杰,孙悦,等.气管滴注纳米二氧化硅在大鼠体内时间分布特征[J].中国职业医学,2017,44(5):562-567.
[15]陆敏,孙湖泊,李妍.纳米二氧化硅对大鼠心肺的损伤作用及机制[J].中国老年学杂志,2017,37(9):2092-2094.
[16]林本成,袭著革,张英鸽,等.三种纳米材料致大鼠肝肾损伤的初步研究[J].卫生研究,2008,37(6):651-653.
[17]龚春梅,杨淋清,陶功华,等.纳米二氧化硅对体外培养Ha Ca T细胞凋亡的影响[J].安徽农业科学,2013,41(11):4842-4845,4876.
[18]焦常平,叶亚婧,张渊,等.纳米二氧化硅诱导A549细胞损伤及其氧化应激机制[J].中国药理学与毒理学杂志,2013,27(6):995-999.
[19]ROSADO M M,BENNICI E,NOVELLI F,et al.Beyond DNA repair,the immunological role of PARP-1 and its siblings[J].Immunology,2013,139(4):428-437.
[20]THOMAS C,KOTOVA E,TULIN A V.PARP-1 Interaction with and Activation by Histones and Nucleosomes[J].Methods Mol Biol,2017,1608:255-267.
[21]MANGERICH A,BURKLE A.Pleiotropic cellular functions of PARP1 in longevity and aging:genome maintenance meets inflammation[J].Oxid Med Cell Longev,2012:321653.
[22]LI H,LI Q,LI W,et al.The Role of PARP-1 in Host-Pathogen Interaction and Cellular Stress Responses[J].Crit Rev Eukaryot Gene Expr,2015,25(2):175-190.
[23]ZHANG N,ZHANG Y,ZHAO S,et al.Septin4 as a novel binding partner of PARP1 contributes to oxidative stress induced human umbilical vein endothelial cells injure[J].Biochem Biophys Res Commun,2018,496(2):621-627.
[24]GONG C,TAO G,YANG L,et al.Methylation of PARP-1 promoter involved in the regulation of nano-Si O2-induced decrease of PARP-1mRNA expression[J].Toxicol Lett,2012,209(3):264-269.
[25]WANG F,JIAO C,LIU J,et al.Oxidative mechanisms contribute to nanosize silican dioxide-induced developmental neurotoxicity in PC12cells[J].Toxicol In Vitro,2011,25(8):1548-1556.
[26]CALKINS A S,IGLEHART J D,LAZARO J B.DNA damageinduced inhibition of rRNA synthesis by DNA-PK and PARP-1[J].Nucleic Acids Res,2013,41(15):7378-7386.
[27]赵光强,黄云超,李光剑,等.纳米二氧化硅在人支气管上皮细胞内的亚细胞分布和遗传毒性[J].中国肺癌杂志,2013,16(3):117-124.
[28]洪文旭,陈晓丹,彭圣明,等.姜黄素对纳米二氧化硅诱导Ha Ca T细胞氧化应激损伤保护作用研究[J].热带医学杂志,2013,13(9):1061-1064.
[29]洪文旭,周桂凤,胡韬,等.姜黄素对纳米二氧化硅诱导Ha Ca T细胞氧化损伤的保护作用[J].毒理学杂志,2013,27(6):412-415.
[30]GONG C,YANG L,ZHOU J,et al.Possible role of PAPR-1 in protecting human Ha Ca T cells against cytotoxicity of Si O2nanoparticles[J].Toxicol Lett,2017,280:213-221.