摘要
[目的]探讨对氨基水杨酸钠(PAS-Na)对体外染锰大鼠丘脑原代小胶质细胞氧化损伤、炎症反应的影响。[方法]将提纯、鉴定后的SD大鼠丘脑小胶质细胞分别给予含不同浓度氯化锰(MnCl_2)(0、200、300、400、500μmol/L)、PAS-Na(50、150、450μmol/L)的完全培养基(DMEM+FBS)培养24 h,采用MTT法检测细胞存活率,根据结果选择染锰的毒性剂量并选定PAS-Na的无毒剂量。随后将细胞随机分为对照组、400μmol/L MnCl_2组、400μmol/L MnCl_2+(50、150、450μmol/L)PAS-Na组、450μmol/L PAS-Na组,共6组。采用MTT法检测各组小胶质细胞存活率;运用DCFH-DA探针标记,荧光倒置显微镜下观察小胶质细胞内活性氧(ROS)的产生情况;ELISA法测定炎症因子白介素(IL)-1β、IL-6、肿瘤坏死因子(TNF)-α的蛋白分泌量和荧光定量PCR法测IL-1β,TNF-αmRNA表达水平。[结果]400、500μmol/L MnCl_2染毒组的细胞存活率为(85.83±12.53)%、(83.30±6.33)%,低于对照组(均P<0.05),故选择400μmol/L作为锰的染毒剂量;与对照组比较,50、150、450μmol/L PAS-Na组的细胞存活率无明显变化(均P>0.05)。400μmol/L MnCl_2+(50、150、450μmol/L)PAS-Na组的细胞存活率分别为(96.00±18.11)%、(106.13±18.32)%、(110.21±15.13)%,均比400μmol/L MnCl_2组高(均P<0.05)。与对照组比较,400μmol/L MnCl_2组细胞ROS水平、IL-1β和TNF-α的蛋白及mR NA表达水平均升高(均P<0.05)。与400μmol/L MnCl_2组比较,400μmol/L MnCl_2+(50、150、450μmol/L)PAS-Na组的ROS水平、TNF-α分泌量和IL-1βmRNA表达量均降低(均P<0.05),400μmol/L MnCl_2+(150、450μmol/L)PAS-Na组的TNF-αmRNA表达量降低(均P<0.05)。[结论]PAS-Na可减轻锰致大鼠丘脑原代小胶质细胞氧化损伤、炎症反应的程度。
[Objective] To investigate the effects of sodium p-aminosalicylic acid(PAS-Na) on oxidative damage and inflammatory response of rat thalamic primary microglia induced by manganese. [Methods] Purified and identified SD rat’s thalamic microglia were cultured with different concentrations of MnCl2(0, 200, 300,400, and 500 μmol/L, respectively) and PAS-Na(50, 150, and 450 μmol/L, respectively) in complete mediums(DMEM+FBS) for 24 h. Cell viability was determined by MTT to identify a suitable dose of manganese exposure and non-toxic dose of PAS-Na exposure. Then cells were randomly divided into control group, 400 μmol/L MnCl2 group, 400 μmol/L MnCl2+PAS-Na(50, 150, and 450 μmol/L, respectively) groups, and 450 μmol/L PAS-Na group. The survival rate of microglia in each group was detected by MTT assay. The level of intercellular reactive oxygen species(ROS) was detected under fluorescence inverse microscope with DCFH-DA probe mark. The protein expressions of interleukin(IL)-1β, IL-6, and tumor necrosis factor(TNF)-α were detected by ELISA, and the mRNA expression levels of IL-1β and TNF-α were determined by RT-PCR.[Results] The survival rates of cells treated with 400 and 500 μmol/L MnCl2 were(85.83±12.53)% and(83.30±6.33)% , respectively, lower than that of the control group(Ps 〈 0.05). Therefore, the exposure dose of 400 μmol/L was chosen. Compared with the control group, the survival rates of the 50, 150, and 450 μmol/L PAS-Na groups did not change(Ps 〉 0.05). The survival rates of the 400 μmol/L MnCl2+PAS-Na(50, 150, 450 μmol/L) groups were(96.00±18.11)% ,(106.13±18.32)% , and(110.21±15.13)% , respectively, higher than that of the 400 μmol/L MnCl2 group(Ps 〈 0.05). Compared with the control group, the levels of ROS, the protein and mRNA expression levels of IL-1β and TNF-α in the 400 μmol/L MnCl2 group increased(Ps 〈 0.05). Compared with the 400 μmol/L MnCl2 group, the levels of ROS, protein expression levels of TNF-α, and the mR NA expression levels of IL-1β in the 400 μmol/L MnCl2+PAS-Na groups(50, 150, 450 μmol/L) decreased(Ps 〈 0.05), and the mRNA expression levels of TNF-α in the 400 μmol/L MnCl2+PAS-Na groups(150, 450 μmol/L) also decreased(Ps 〈 0.05). [Conclusion] PAS-Na may alleviate the oxidative damage and inflammatory response of thalamic primary microglia in rats induced by manganese.
作者
彭东杰
罗旖旎
秦文霞
梁典胤
谢秉言
许放
李少军
姜岳明
PENG Dong-jie;LUO Yi-ni;QIN Wen-xia;LIANG Dian-yin;XIE Bing-yan;XU Fang;LI Shao-jun;JIANG Yue-ming(Department of Health Toxicology, School of Public Health;Guangxi Colleges and Universities Key Laboratory of Prevention and Control of Highly Prevalent Diseases, Nanning, Guangxi 530021, China;Department of Preventive Health, Wuxiang Hospital, Second People's Hospital of Nanning, Nanning, Guangxi 530031, China)
出处
《环境与职业医学》
CAS
CSCD
北大核心
2018年第4期309-315,共7页
Journal of Environmental and Occupational Medicine
基金
国家自然科学基金(编号:81460505
81773476)
关键词
对氨基水杨酸钠
锰
小胶质细胞
氧化应激
炎症
sodium p-aminosalicylic acid
manganese
microglia
oxidative stress
inflammation