PP2Ac去甲基化在锰暴露诱导大鼠肝脏炎症性损伤中的作用研究  

The role of PP2Ac demethylation in inflammatory liver injury induced by manganese exposure in rats

作　　者：黄丽媛 陆建勇 梁宁静 袁江浪 孔星星 李习艺[1] 唐深[2] 陆彩玲 Huang Liyuan;Lu Jianyong;Liang Ningjing;Yuan Jianglang;Kong Xingxing;Li Xiyi;Tang Shen;Lu Cailing(School of Public Health,Guangxi Medical University,Nanning 530021,China;School of Basic Medical Sciences,Guangxi Medical University,Nanning 530021,China)

机构地区：[1]广西医科大学公共卫生学院,南宁530021 [2]广西医科大学基础医学院,南宁530021

出　　处：《广西医科大学学报》2022年第12期1911-1917,共7页Journal of Guangxi Medical University

基　　金：国家自然科学基金项目(No.81760576,No.81860585,No.82160612);广西自然科学基金项目(No.2021GXNSFAA196019)。

摘　　要：目的:研究亚急性及慢性锰暴露对SD大鼠肝脏的损伤效应,探索PP2Ac去甲基化修饰在锰暴露致肝脏炎症性损伤中的作用。方法:40只雄性SD大鼠按体重随机入组,分别设置4周(4W)和16周(16W)两个暴露时间点,每个时间点分两组(即4W、16W对照组和4W、16W锰暴露组),每组10只;锰暴露组腹腔注射MnCl_(2)·4H_(2)O(15 mg/kg),对照组注射同体积的生理盐水,每天1次,持续4W、16W。实验终点时分离大鼠肝脏,ICP-MS法检测血液及肝组织中锰的含量。苏木精—伊红(HE)染色观察肝脏的病理改变。实时荧光定量聚合酶链式反应(RT-qPCR)法检测肝脏炎症相关因子肿瘤坏死因子α(TNF-α)、白介素6(IL-6)、趋化因子CCL-2的表达。Western blotting法检测肝组织总PP2Ac、去甲基化PP2Ac及PP2Ac甲基化调控蛋白LCMT-1及PME-1的表达。结果:与4 W对照组相比,4 W、16 W锰暴露组全血锰含量显著升高(P<0.01),4 W锰暴露组肝组织锰含量显著下降(P<0.05),而16 W锰暴露组肝组织锰含量显著升高(P<0.01);HE染色结果显示,与4 W对照组相比,4 W锰暴露组出现肝血窦扩张,炎症细胞浸润,16 W锰暴露组与16 W对照组相比,肝索排列紊乱,细胞间隙增大,肝窦进一步扩张充血,大量肝细胞膜破裂;与4 W对照组相比,4 W锰暴露组仅肝脏炎症因子IL-6的表达呈上升趋势,16 W锰暴露组CCL-2、IL-6的mRNA表达与16 W对照组相比呈升高趋势,并且TNF-α的mRNA表达显著升高(P<0.05);4 W、16 W锰暴露组的去甲基化PP2Ac表达水平升高(P<0.05),仅16W锰暴露组的甲基化PP2Ac修饰蛋白LCMT-1表达水平降低(P<0.01),不同时点锰暴露组PME-1、总PP2Ac蛋白表达无明显变化。结论:锰诱导大鼠肝脏产生炎症性损伤,可能与上调肝脏PP2Ac去甲基化有关。Objective:To study the damage effects of subacute and chronic manganese exposure on the liver of SD rats and explore the role of catalytic subunit of protein phosphatase 2A(PP2Ac)demethylation modification in the inflammatory liver damage induced by manganese exposure.Methods:Forty male SD rats were randomly enrolled according to body weight,and two exposure time points were set at 4 weeks(4 W)and 16 weeks(16 W),respectively.Each time point was divided into two groups(4 W and 16 W control group,4 W and 16 W manganese exposure group),with 10 rats in each group.The manganese exposure group was intraperitoneally injected with MnCl_(2)·4H_(2)O(15 mg/kg),while the control group was intraperitoneally injected with the same volume of normal saline,once a day for 4 W and 16 W.At the end of the experiment,the rat liver was isolated,and the content of manganese in the blood and liver tissues was detected by ICP-MS.The pathological changes of the liver were observed by HE staining.The expressions of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and chemokine CCL-2 were detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).The expressions of total PP2Ac,demethylated PP2Ac and PP2Ac methylated regulatory proteins LCMT-1 and PME-1 in liver tissues were analyzed by western blotting.Results:Compared with the 4 W control group,the whole blood manganese content in 4 W and 16 W manganese exposure groups significantly increased(P<0.01),the liver manganese content in 4 W manganese exposure group significantly decreased(P<0.05),and the liver manganese content in 16 W manganese exposure group significantly increased(P<0.01).HE staining results showed that compared with the 4 W control group,the 4 W manganese exposure group showed hepatic sinusoidal dilatation and inflammatory cell infiltration,while the 16 W manganese exposure group showed disordered hepatic cord arrangement,enlarged intercellular space,further dilatation and congestion of hepatic sinuses,and a large number of hepatic cell membrane r

关 键 词：锰暴露 肝损伤 炎症 PP2Ac 

分 类 号：R575.1[医药卫生—消化系统]