氯乙烯亚慢性染毒对大鼠肝细胞周期及mir-21和mir-192表达的影响
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摘要
目的探讨氯乙烯(vinyl chloride monomer,VCM)亚慢性染毒对大鼠肝细胞周期以及microRNA21(mir-21)和microRNA192(mir-192)表达量的影响。方法将32只健康斯普拉-道来(sprague dawley,SD)大鼠随机分为三个实验组(5 mg/kg组,25 mg/kg组和125 mg/kg组)与一个对照组(25 mg/kg清洁空气组),每组8只。采用腹腔注射进行VCM染毒,每周3次(隔日染毒)。染毒12周,处死大鼠并摘取肝组织。制备肝单细胞悬液,使用流式细胞技术检测G0/G1期、S期和G2/M期肝细胞所占百分比;提取肝细胞中的小片段RNA(<200 nt),采用实时荧光定量PCR技术检测大鼠肝细胞中mir-21和mir-192的表达量。结果 VCM染毒大鼠G0/G1期、S期和G2/M期肝细胞所占百分比在各剂量组间差异均无统计学意义(均有P>0.05);125 mg/kg组S期细胞与对照组相比明显增加,差异有统计学意义(t=-4.363,P=0.024)。各剂量组之间mir-21相对表达量差异有统计学意义(H=16.064,P=0.001);与对照组和5 mg/kg组相比,25 mg/kg组及125 mg/kg组mir-21相对表达量均降低,差异均有统计学意义(均有P<0.05)。各剂量组之间mir-192相对表达量差异有统计学意义(H=15.939,P=0.001);与对照组和5 mg/kg组相比,25 mg/kg组及125 mg/kg组mir-192相对表达量均降低,差异均有统计学意义(均有P<0.05)。结论 VCM亚慢性染毒导致大鼠肝细胞S期比例增加,mir-21和mir-192表达下调,具体调节机制有待进一步研究。
Objective To explore the sub-chronic toxicity effects of vinyl chloride monomer( VCM) on cell cycle and the expression of cell cycle related microRNA 21( mir-21) and microRNA 192( mir-192) of rat liver. Methods Thirty-two healthy sprague dawley( SD) rats were randomly divided into three VCM exposure groups( 5 mg / kg,25 mg / kg and125 mg / kg) and a control group( 25 mg / kg clean air). The rats were exposed by intraperitoneal injection three times a week( every other day) for three months. The flow cytometry was used to measure the percent of each phase( G0 / G1,S,and G2 / M). The mir-21 and mir-192 was extracted and then quantified using real-time fluorescent quantitative PCR. Results The percentage of each phase of cell cycle was not significantly different among four groups( all P > 0. 05). The proportion of S-phase cells in 125 mg / kg group was higher than the control group( t =- 4. 363,P = 0. 024). Besides,the expressions of mir-21 varied significantly among four groups( H = 16. 064,P = 0. 001) and,furthermore,decreased significantly in both 25 mg / kg and 125 mg / kg group when they were compared with control and 5 mg / kg group( all P < 0. 05).Meanwhile,the expressions of mir-192 also varied significantly( H = 15. 939,P = 0. 001),and decreased significantly in both 25 mg / kg and 125 mg / kg group,compared with control and 5 mg / kg group( all P < 0. 05). Conclusions VCM subchronic exposure induced the increase of S-phase cells and decrease of the expression of mir-21 and mir-192.
Objective To explore the sub-chronic toxicity effects of vinyl chloride monomer( VCM) on cell cycle and the expression of cell cycle related microRNA 21( mir-21) and microRNA 192( mir-192) of rat liver. Methods Thirty-two healthy sprague dawley( SD) rats were randomly divided into three VCM exposure groups( 5 mg / kg,25 mg / kg and125 mg / kg) and a control group( 25 mg / kg clean air). The rats were exposed by intraperitoneal injection three times a week( every other day) for three months. The flow cytometry was used to measure the percent of each phase( G0 / G1,S,and G2 / M). The mir-21 and mir-192 was extracted and then quantified using real-time fluorescent quantitative PCR. Results The percentage of each phase of cell cycle was not significantly different among four groups( all P > 0. 05). The proportion of S-phase cells in 125 mg / kg group was higher than the control group( t =- 4. 363,P = 0. 024). Besides,the expressions of mir-21 varied significantly among four groups( H = 16. 064,P = 0. 001) and,furthermore,decreased significantly in both 25 mg / kg and 125 mg / kg group when they were compared with control and 5 mg / kg group( all P < 0. 05).Meanwhile,the expressions of mir-192 also varied significantly( H = 15. 939,P = 0. 001),and decreased significantly in both 25 mg / kg and 125 mg / kg group,compared with control and 5 mg / kg group( all P < 0. 05). Conclusions VCM subchronic exposure induced the increase of S-phase cells and decrease of the expression of mir-21 and mir-192.
引文
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[20]Sun J,Fan Z,Lu S,et al.miR-192 suppresses the tumorigenicity of prostate cancer cells by targeting and inhibiting nin one binding protein[J].Int J Mol Med,2016,37(2):485-492.
[21]Jin Y,Lu J,Wen J,et al.Regulation of growth of human bladder cancer by miR-192[J].Tumour Biol,2015,36(5):3791-3797.
[22]赵然,周鸣,王贺冉.MicroRNA参与肿瘤发生发展的调控机[J].中南大学学报(医学版),2013,(12):1282-1288.
[23]Xue XY,Zhao B,Chao LM,et al.Interaction between two timing microRNAs controls trichome distribution in Arabidopsis[J].PLo S Genet,2014,10(4):e1004266.(投稿日期:2016-04-01)
[2]Brandt-Rauf PW,Li Y,Long C,et al.Plastics and carcinogenesis:The example of vinyl chloride[J].J Carcinog,2012,11:5.
[3]Benada J,Macurek L.Targeting the Checkpoint to Kill Cancer Cells[J].Biomolecules,2015,5(3):1912-1937.
[4]李武,贺庆芝,孙诗博,等.has-miR-16靶基因预测及生物信息学分析[J].中华疾病控制杂志,2016,(2):193-197.
[5]Yang J,Han S,Huang W,et al.A meta-analysis of microRNA expression in liver cancer[J].PLo S One,2014,9(12):e114533.
[6]Zhang Y,Wang X,Fu Y,et al.Expression profiling and pathway analysis of microRNA expression in the lungs of mice exposed to long-term,low-dose benzo(a)pyrene[J].Molecular&Cellular Toxicology,2014,10(1):67-74.
[7]Izzotti A,Calin GA,Arrigo P,et al.Downregulation of microRNA expression in the lungs of rats exposed to cigarette smoke[J].FASEB J,2009,23(3):806-812.
[8]陈桢坤,单云峰,何彬,等.miR-21在大鼠肝卵圆细胞活化和增殖过程中作用研究[J].中华肝脏病杂志,2014,22(11):854-859.
[9]Huang S,Deng Q,Feng J,et al.Polycyclic Aromatic Hydrocarbons-Associated MicroRNAs and Heart Rate Variability in Coke Oven Workers[J].J Occup Environ Med,2016,58(1):e24-31.
[10]王爱红,朱守民,周元陵,等.氯乙烯染毒大鼠代谢酶活性的动态变化和肝损伤[J].卫生研究,2004,(3):258-260.
[11]Deng Q,Huang S,Zhang X,et al.Plasma microRNA expression and micronuclei frequency in workers exposed to polycyclic aromatic hydrocarbons[J].Environ Health Perspect,2014,122(7):719-725.
[12]张盼红.氯乙烯对大鼠肝细胞周期相关蛋白表达的影响[D].太原:山西医科大学,2013.
[13]Santo L,Siu KT,Raje N.Targeting Cyclin-Dependent Kinases and Cell Cycle Progression in Human Cancers[J].Semin Oncol,2015,42(6):788-800.
[14]Morinello EJ,Ham AJ,Ranasinghe A,et al.Molecular dosimetry and repair of N(2),3-ethenoguanine in rats exposed to vinyl chloride[J].Cancer Res,2002,62(18):5189-5195.
[15]王慧.氯乙烯对HL-7702细胞周期的影响及G1/S关卡相关基因mRNA的表达[D].太原:山西医科大学,2012.
[16]刘庆成,杨淋清,陶功华,等.低剂量甲醛对16HBE细胞的增殖促进及DNA损伤作用[J].中华疾病控制杂志,2011,15(11):927-930.
[17]席华星,聂继盛,牛侨.苯并芘对神经细胞DNA损伤及细胞周期的影响[J].环境与职业医学,2012,(10):620-623,628.
[18]Gao S,Tian H,Guo Y,et al.miRNA oligonucleotide and sponge for miRNA-21 inhibition mediated by PEI-PLL in breast cancer therapy[J].Acta Biomater,2015,25:184-193.
[19]Zhong Z,Dong Z,Yang L,et al.miR-21 induces cell cycle at S phase and modulates cell proliferation by down-regulating h MSH2in lung cancer[J].J Cancer Res Clin Oncol,2012,138(10):1781-1788.
[20]Sun J,Fan Z,Lu S,et al.miR-192 suppresses the tumorigenicity of prostate cancer cells by targeting and inhibiting nin one binding protein[J].Int J Mol Med,2016,37(2):485-492.
[21]Jin Y,Lu J,Wen J,et al.Regulation of growth of human bladder cancer by miR-192[J].Tumour Biol,2015,36(5):3791-3797.
[22]赵然,周鸣,王贺冉.MicroRNA参与肿瘤发生发展的调控机[J].中南大学学报(医学版),2013,(12):1282-1288.
[23]Xue XY,Zhao B,Chao LM,et al.Interaction between two timing microRNAs controls trichome distribution in Arabidopsis[J].PLo S Genet,2014,10(4):e1004266.(投稿日期:2016-04-01)