中国公共卫生  2010, Vol. 26 Issue (9): 1117-1119   PDF    
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张军, 王艳, 赵凤红, 钟媛, 于霄云, 李革新, 吕秀强, 孙贵范, 金亚平. 蛋氨酸对砷暴露小鼠脑砷形态及NO代谢影响[J]. 中国公共卫生, 2010, 26(9): 1117-1119.
ZHANG Jun, WANG Yan, ZHAO Feng-hong, et al. Effects of methionine on distribution of arsenicals and NO metabolism in brain of mice exposed to arsenite[J]. Chinese Journal of Public Health, 2010, 26(9): 1117-1119.
蛋氨酸对砷暴露小鼠脑砷形态及NO代谢影响
张军1, 王艳1, 赵凤红2, 钟媛2, 于霄云2, 李革新2, 吕秀强2, 孙贵范2, 金亚平2    
1. 辽宁中医药大学职业技术学院卫生管理系, 沈阳 110101;
2. 中国医科大学公共卫生学院劳动卫生教研室
收稿日期: 2010-01-10.
基金项目: 国家自然科学基金(30571590;30972441)
作者简介: 张军(1962- ),男,福建闽候人,副教授,本科,主要从事预防医学科研工作.
摘要: 目的 探讨外源性蛋氨酸(methionine,Met)对饮水砷暴露小鼠脑中砷形态分布及NO代谢影响.方法 将小鼠随机分为对照组、染砷组及低、中、高剂量Met干预组.饮水染砷4周,最后1周在染砷同时腹腔注射不同剂量Met,末次注射后24 h处死小鼠,取血和脑组织检测砷、NO等指标.结果 与染砷组比较,各Met干预组小鼠血中无机砷(iAs)、一甲基胂(MMA)和总砷(TAs)含量及脑中iAs、二甲基胂(DMA)和TAs含量明显降低;与对照组比较,染砷组小鼠脑中一氧化氮合酶(NOS)活性〔(1.526±0.185)U/(mg·pro)〕及一氧化氮(NO)含量〔(0.472±0.129)μmol/(g·pro)〕明显降低;高剂量Met干预组小鼠脑中NO含量〔(0.666±0.135)μmol/(g.pro)〕明显升高.结论 给予外源性Met可减少血液和脑组织中的砷负荷,进而改善砷对脑内NO代谢影响.
关键词: 砷中毒     蛋氨酸     砷形态     一氧化氮合酶     一氧化氮    
Effects of methionine on distribution of arsenicals and NO metabolism in brain of mice exposed to arsenite
ZHANG Jun, WANG Yan, ZHAO Feng-hong, et al    
Department of Public Health Management, School of Professional Technology, Liaoning University of Traditional Chinese Medicine Shenyang 110101, China
Abstract: Objective To explore the effects of exogenous methionine(Met) on distribution of a rsenicals and nitric oxide metabolism in the brain of mice exposed to arsenite in drinking water.Methods Forty mice were divided randomly into 5g roups with 8 mice in each group.The mice in the exper miental groups were exposed to drinking water contam inated with 50 mg/L arsenic for four consecutive weeks.At the fourth week,the mice treated intraper itoneally with Met at doses of 100,200,and 400 mg/kg·bwand saline solution separately.Twenty four hours afterce ssation of Metadm in istration,the mice were anaesthetized and dissected rapidly.Sam ples of blood and brain were collected mimediately for analysis of arsenic species.Levels of inorganic arsenic(Ais),monomethy larsenic acid(MMA) and dmiethy larsenic acid(DMA) were determ ined with HG AAS method.Activities of nitric oxide synthase(NOS) and the concentrations of nitricoxide(NO) were determined with kits purchased from Nanjing Jiancheng Co.Results Compared to those treated with saline solution,adm inistration of Met significantly decreased the levels of Ais,MMA and total arsenic levels(TAs) in the blood and the levels of Ais,DM A and TAs in the brain.A ctivities of NOS and the levels of NO in As exposure group were significantly lower than those of in the control.However administration of Metcouldame liorate these toxic effects and NO levels in the group treated with 400mg/kg·bw.Met were significantly higher than those in saline solution treated group.Conclusion Exogenous Met could promote methylation of a rsenic,there fore reduce arsenic levels in both blood and brain.Moreover,administration of exogenous methionine couldame liorate the effects of arsenic on NO metabo lism in the brain via decreasing arsenic burden.
Key words: arsenite     methionine     arsenic species     nitric oxide synthase     nitricoxide    

饮水型砷中毒是中国主要地方病之一〔1〕。无机砷(inorganicarsenic,iAs)进入人体后,被甲基化代谢形成一甲基胂(monomethylarsonicacid,MMA)和二甲基胂(dimethylarsenicacid,DMA)〔2〕。蛋氨酸(methionine,Met)在砷甲基化代谢中起重要作用〔3〕。一氧化氮(nitricoxide,NO)是中枢神经系统中的重要信使分子〔4〕。砷对脑中NO水平及相关代谢酶活性的影响可能是砷对中枢神经系统毒作用的重要途径之一。本研究通过给予砷暴露小鼠不同剂量Me,t探讨其对小鼠脑中砷形态及NO代谢影响,为砷中毒防治提供理论依据。

1 材料与方法 1.1 试剂与仪器

亚砷酸钠(NaAsO2)(上海分析试剂厂),分析纯,Met(美国Sigma公司),优级纯;一氧化氮合酶(NOS)与NO试剂盒(南京建成生物工程研究所);分光光度计(上海凤凰光学仪器公司)。

1.2 实验动物

健康雌性清洁级昆明种小鼠(中国医科大学实验动物中心)40只,体重(25.0±2.0)g。动物室温度(20±2)℃,相对湿度为50%~60%,适应性喂养1周。

1.3 分组与处理

将小鼠随机分为5组,分别为对照、染砷组及低、中、高剂量Met干预组,每组8只小鼠。对照组小鼠饮蒸馏水,其余各组小鼠自由饮用含砷水(50mg/L),连续4周。在第4周,低、中、高Met干预组分别腹腔注射100,200,400mg/kgMet溶液(生理盐水配制),对照与染砷组腹腔注射生理盐水,共7d。末次注射24h后,处死小鼠,取血和脑组织,保存于-40℃冰箱待测。

1.4 砷含量测定

取50mg脑组织,加入1.0mL三蒸水,制成组织匀浆,并与2.0mL1.5mol/LH2SO4混合;取0.1mL全血与2.0mL1.0mol/LH2SO4混合。超声消化10min。采用氢化物发生-超低温捕集-原子吸收分光光度法(HG-AAS)测定消化液中iAs、MMA和DMA含量。并计算血和脑组织中总砷含量(TAs=iAs+MMA+DMA)。

1.5 NOS活性及NO含量测定

按试剂盒说明书进行。

1.6 统计分析

采用SPSS 13.0进行统计分析。多组间比较采用单因素方差分析(ANOVA),组间两两比较采用q检验(SNK),以P<0.05为差异有统计学意义。

2 结 果 2.1 各组小鼠血中不同形态砷含量比较 (表 1)
表 1 各组小鼠血中不同形态砷含量比较 ( x± s, ng /mL, n= 8)

单纯染砷组及各Met干预组小鼠血中iAs、MMA、DMA和TAs含量均明显高于对照组(P<0.05)。各Met干预组小鼠血中iAs、MMA和TAs含量与单纯染砷组比较均明显降低(P<0.05)。对照组小鼠血中MMA和DMA含量未检出。

2.2 各组小鼠脑中不同形态砷含量比较 (表 2)
表 2 各组小鼠脑中不同形态砷含量的比较 ( x± s, ng /mL, n= 8)

单纯染砷组小鼠脑中iAs、DMA和TAs含量均明显高于对照组(P<0.05)。各剂量Met干预组小鼠脑中iAs、DMA和TAs含量与单纯染砷组比较均明显降低(P<0.05)。各组小鼠脑中MMA含量及对照组小鼠脑中DMA均未检出。

2.3 各组小鼠脑组织NOS活性和NO含量比较 (表 3)
表 3 各组小鼠脑组织NOS活性与NO含量的比较 ( x± s , n= 8)

单纯染砷组小鼠脑组织NOS活性和NO含量与对照组比较明显降低;各Met干预组小鼠脑组织NOS活性和NO含量与单纯染砷组比较均有升高,并与对照组比较差异无统计学意义,高剂量Met干预组小鼠脑中NO含量与单纯染砷组比较差异有统计学意义(P<0.05)。

3 讨 论

iAs与其甲基代谢物由肝脏入血后,随血液循环进入机体其他组织器官。研究表明,某些肝外组织也具有砷甲基化代谢能力〔5〕。本研究结果表明,Met干预可明显降低小鼠血中iAs、MMA及TAs的含量,提示Met通过促进iAs在体内的甲基化代谢过程,有效降低了血液中的砷负荷。NO在中枢神经系统中起重要生理调节作用,NO含量变化会严重影响脑组织功能。本研究发现,单纯染砷组小鼠脑中NOS活性和NO含量明显低于对照组,与SergioZ等〔6〕研究结果一致,进一步证明砷暴露可明显干扰脑组织NO代谢。给予Met可有效降低进入脑中的iAs、DMA及TAs含量,从而减少了脑组织的砷暴露水平,可能与Met有效降低血中各形态砷的含量有关。Met干预可提升小鼠脑组织NOS活性和NO含量,明显改善砷对脑组织NO代谢的影响,可能与外源性Met降低了脑中的砷负荷有关。

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