脱锰后大鼠PARK2基因表达及运动和记忆功能的变化
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摘要
[目的]探讨大鼠脱锰前后全血中PARK2基因表达水平的改变,及其与大鼠运动、记忆功能的关系。[方法]60只健康成年SPF级SD大鼠(6周龄,160~180 g)随机分为对照组(生理盐水)、低剂量锰组(1 mg/kg)、高剂量锰组(5 mg/kg),每组20只。以5 m L/kg体积大鼠腹腔注射MnCl_2染锰,隔天1次,连续4周。每组随机抽取10只大鼠进行转棒和Y迷宫实验分别检测运动和记忆功能;麻醉后取全血,石墨炉原子吸收仪检测全血中锰质量浓度(后称浓度),实时荧光定量PCR检测全血中PARK2基因表达情况。各组余下大鼠停止染锰,继续常规饲养5个月后操作同前。期间每月对大鼠进行乙醚麻醉后取内眦静脉全血动态检测血液中PARK2表达变化。[结果]染锰1个月后:与对照组[(2.10±0.15)mg/L]比较,高[(5.31±0.23)mg/L]、低[(3.13±0.16)mg/L]剂量组大鼠血锰浓度增高(P<0.05);高、低剂量组大鼠PARK2基因相对表达水平分别为6.83±1.97、13.15±3.84,均低于对照组(26.05±3.66)(P<0.05);高剂量组大鼠转棒距离[(743.56±99.66)cm]低于对照组[(1 547.09±192.53)cm](P<0.05);大鼠全血锰浓度与转棒距离(r=-0.74)、全血PARK2表达(r=-0.57)均呈负相关(均P<0.05),全血PARK2水平与转棒距离呈正相关(r=0.61,P=0.02)。脱锰5个月后:高剂量组大鼠血锰浓度[(2.54±0.20)mg/L]较前下降(P<0.05),但仍高于对照组[(1.80±0.09)mg/L](P<0.05);高剂量组大鼠转棒距离为(1 105.73±132.23)cm,较染锰1个月时增加(P<0.05)。[结论]亚急性锰染毒大鼠脱锰后,全血PARK2基因相对表达水平及运动功能可恢复,PARK2基因可能作为锰致大鼠运动功能损伤的潜在标志。
[Objective] To explore the change of PARK2 gene expression in whole blood before and after cessation of manganese exposure in rats, and the association with motor and memory functions.[Methods] Sixty healthy adult SD rats were randomly divided into control group(saline), low dose group(MnCl_2, 1 mg/kg), and high dose group(MnCl_2, 5 mg/kg), with 20 rats in each group. The animals were injected intraperitoneally with MnCl_2 at 5 m L/kg, once every other day. After continuous intraperitoneal injection for 4 weeks, 10 rats in each group were randomly selected for experiments on motor and memory functions such as rotarod performance test(RPT) and Y maze experiment. PARK2 gene expression in whole blood was detected by real-time fluorescence quantitative PCR(RT-PCR), and concentration of manganese in whole blood by graphite furnace atomic absorption spectrometry(AAS). The remaining rats in each group stopped intraperitoneal injection of manganese and received normal feeding for 5 months. They were anesthetized with ether every month to collect whole blood samples of infrarenal vein and detect changes of PARK2 expression.[Results] One month after intraperitoneal injection of MnCl_2, the blood levels of manganese of the high dose group [(5.31±0.23) mg/L] and low dose group [(3.13±0.16) mg/L] were increased compared with the control group [(2.10±0.15) mg/L](P < 0.05); the PARK2 gene expression levels of the high dose group(6.83±1.97) and the low dose group(13.15±3.84) were lower than that of the control group(26.05±3.66)(P < 0.05); the distance traveled in RPT of the high dose group [(743.56±99.66) cm] was lower than that of the control group [(1 547.09±192.53) cm](P < 0.05); the level of manganese in whole blood was negatively correlated with the RPT distance traveled(r =-0.74) and PARK2 gene expression(r =-0.57)(Ps < 0.05); PARK2 gene expression was positively correlated with the distance traveled in RPT(r = 0.61, P=0.02). After 5 months of cessation of manganese exposure, the blood level of manganese in the high dose group [(2.54±0.20) mg/L] was decreased(P < 0.05), but still higher than that in the control group [(1.80±0.09) mg/L](P < 0.05); the distance traveled of the high dose group was(1 105.73±132.23) cm, which was higher than that before cessation of manganese exposure(P < 0.05).[Conclusion] PARK2 gene expression in whole blood and motor function of rats would recover after cessation of subacute manganese poisoning, and PARK2 gene may be used to determine the damage to motor function caused by manganese.
[Objective] To explore the change of PARK2 gene expression in whole blood before and after cessation of manganese exposure in rats, and the association with motor and memory functions.[Methods] Sixty healthy adult SD rats were randomly divided into control group(saline), low dose group(MnCl_2, 1 mg/kg), and high dose group(MnCl_2, 5 mg/kg), with 20 rats in each group. The animals were injected intraperitoneally with MnCl_2 at 5 m L/kg, once every other day. After continuous intraperitoneal injection for 4 weeks, 10 rats in each group were randomly selected for experiments on motor and memory functions such as rotarod performance test(RPT) and Y maze experiment. PARK2 gene expression in whole blood was detected by real-time fluorescence quantitative PCR(RT-PCR), and concentration of manganese in whole blood by graphite furnace atomic absorption spectrometry(AAS). The remaining rats in each group stopped intraperitoneal injection of manganese and received normal feeding for 5 months. They were anesthetized with ether every month to collect whole blood samples of infrarenal vein and detect changes of PARK2 expression.[Results] One month after intraperitoneal injection of MnCl_2, the blood levels of manganese of the high dose group [(5.31±0.23) mg/L] and low dose group [(3.13±0.16) mg/L] were increased compared with the control group [(2.10±0.15) mg/L](P < 0.05); the PARK2 gene expression levels of the high dose group(6.83±1.97) and the low dose group(13.15±3.84) were lower than that of the control group(26.05±3.66)(P < 0.05); the distance traveled in RPT of the high dose group [(743.56±99.66) cm] was lower than that of the control group [(1 547.09±192.53) cm](P < 0.05); the level of manganese in whole blood was negatively correlated with the RPT distance traveled(r =-0.74) and PARK2 gene expression(r =-0.57)(Ps < 0.05); PARK2 gene expression was positively correlated with the distance traveled in RPT(r = 0.61, P=0.02). After 5 months of cessation of manganese exposure, the blood level of manganese in the high dose group [(2.54±0.20) mg/L] was decreased(P < 0.05), but still higher than that in the control group [(1.80±0.09) mg/L](P < 0.05); the distance traveled of the high dose group was(1 105.73±132.23) cm, which was higher than that before cessation of manganese exposure(P < 0.05).[Conclusion] PARK2 gene expression in whole blood and motor function of rats would recover after cessation of subacute manganese poisoning, and PARK2 gene may be used to determine the damage to motor function caused by manganese.
引文
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[8]CHANG C Y,WU G L,GAO P Y,et al.Upregulated Parkin expression protects mitochondrial homeostasis in DJ-1konckdown cells and cells overexpressing the DJ-1 L166P mutation[J].Mol Cell Biochem,2014,387(1/2):187-195.
[9]邓宇,徐兆发.锰的神经兴奋毒性及其机制[J].中国职业医学,2008,35(2):148-150.
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[11]罗英,许洁,范希敏,等.PARK2基因在锰致大鼠运动功能降低中的作用[J].毒理学杂志,2016,30(1):27-30.
[12]ZHENG W,KIM H,ZHAO Q Q.Comparative toxicokinetics of manganese chloride and methylcyclopentadienyl manganese tricarbonyl(MMT)in sprague-dawley rats[J].Toxicol Sci,2000,54(2):295-301.
[13]TAKEDA A,SAWASHITA J,OKADA S.Biological half-lives of zinc and manganese in rat brain[J].Brain Res,1995,695(1):53-58.
[14]ROMANí-AUMEDES J,CANAL M,MARTíN-FLORES N,et al.Parkin loss of function contributes to RTP801 elevation and neurodegeneration in Parkinson’s disease[J].Cell Death Dis,2014,5(8):e1364.
[15]TAN Y,YU F R,PEREIRA A,et al.Suppression of Nrdp1toxicity by Parkin in Drosophila models[J].Biochem Biophys Res Commun,2011,416(1/2):18-23.
[16]王曼,刘晓莉.多巴胺在运动改善帕金森病学习记忆障碍中的调节作用[J].中国老年学杂志,2016,36(10):2542-2545.
[17]陶冬,邓妍,张蓓,等.锰对大鼠纹状体神经细胞PARK2表达的影响[J].遵义医学院学报,2016,39(6):588-592.
[2]SHI S S,ZHAO J Y,YANG L,et al.KHSRP participates in manganese-induced neurotoxicity in rat striatum and PC12cells[J].J Mol Neurosci,2015,55(2):454-465.
[3]SHIMURA H,HATTORI N,KUBO S I,et al.Familial Parkinson disease gene product,parkin,is a ubiquitin-protein ligase[J].Nat Genet,2000,25(3):302-305.
[4]FAN Q Y,ZHOU Y,YU C Y,et al.Cross-sectional study of expression of divalent metal transporter-1,transferrin,and hepcidin in blood of smelters who are occupationally exposed to manganese[J].Peer J,2016,4:e2413.
[5]FAN X M,LUO Y,FAN Q Y,et al.Reduced expression of PARK2 in manganese-exposed smelting workers[J].Neurotoxicology,2017,62:258-264.
[6]邓妍,隋典朋,许洁,等.锰接触所引起全血中PARK2基因表达变化[J].遵义医学院学报,2011,34(6):567-569.
[7]CHASE B A,MARKOPOULOU K.PARK2,parkin[M]//METMAN L V,KOMPOLITI K.Encyclopedia of Movement Disorders.Amsterdam:Elsevier Academic Press,2010:383-386.
[8]CHANG C Y,WU G L,GAO P Y,et al.Upregulated Parkin expression protects mitochondrial homeostasis in DJ-1konckdown cells and cells overexpressing the DJ-1 L166P mutation[J].Mol Cell Biochem,2014,387(1/2):187-195.
[9]邓宇,徐兆发.锰的神经兴奋毒性及其机制[J].中国职业医学,2008,35(2):148-150.
[10]JIANG Y M,MO X A,DU F Q,et al.Effective treatment of manganese-induced occupational Parkinsonism with p-Aminosalicylic acid:a case of 17-year follow-up study[J].J Occup Environ Med,2006,48(6):644-649.
[11]罗英,许洁,范希敏,等.PARK2基因在锰致大鼠运动功能降低中的作用[J].毒理学杂志,2016,30(1):27-30.
[12]ZHENG W,KIM H,ZHAO Q Q.Comparative toxicokinetics of manganese chloride and methylcyclopentadienyl manganese tricarbonyl(MMT)in sprague-dawley rats[J].Toxicol Sci,2000,54(2):295-301.
[13]TAKEDA A,SAWASHITA J,OKADA S.Biological half-lives of zinc and manganese in rat brain[J].Brain Res,1995,695(1):53-58.
[14]ROMANí-AUMEDES J,CANAL M,MARTíN-FLORES N,et al.Parkin loss of function contributes to RTP801 elevation and neurodegeneration in Parkinson’s disease[J].Cell Death Dis,2014,5(8):e1364.
[15]TAN Y,YU F R,PEREIRA A,et al.Suppression of Nrdp1toxicity by Parkin in Drosophila models[J].Biochem Biophys Res Commun,2011,416(1/2):18-23.
[16]王曼,刘晓莉.多巴胺在运动改善帕金森病学习记忆障碍中的调节作用[J].中国老年学杂志,2016,36(10):2542-2545.
[17]陶冬,邓妍,张蓓,等.锰对大鼠纹状体神经细胞PARK2表达的影响[J].遵义医学院学报,2016,39(6):588-592.