敲低小胶质细胞中过氧还原素6(PRDX6)降低氧糖剥夺再复氧神经元的存活  

Knockdown of PRDX6 in microglia reduces neuron viability after OGD/R injury

作　　者：谭丽[1] 赵涌[1] 姜蓓蓓[1] 杨波[1] 张徽[1] 

机构地区：[1]重庆医科大学基础医学院病理学教研室,分子医学与肿瘤研究中心,重庆400016

出　　处：《细胞与分子免疫学杂志》2016年第8期1014-1020,共7页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81271460,81301125)

摘　　要：目的观察敲低小胶质细胞中过氧还原素6(PRDX6)对氧糖剥夺再复氧(OGD/R)神经元存活的影响。方法小胶质细胞采用PRDX6-siRNA慢病毒感染,同时培养基中加入非Ca2+依赖性磷脂酶A2(i PLA2)活性抑制剂MJ33,24 h后与原代神经元共培养,并进行OGD/R处理。实验分为正常组、OGD/R组、阴性对照siRNA处理的OGD/R组、PRDX6-siRNA处理的OGD/R组、PRDX6-siRNA联合MJ33处理的OGD/R组。Western blot法检测小胶质细胞PRDX6蛋白的表达,实时定量PCR检测小胶质细胞PRDX6 mRNA的表达。ELISA检测i PLA2活性;MTS、乳酸脱氢酶(LDH)法检测神经元的存活率;采用比色法检测超氧化物歧化酶(SOD)和丙二醛(MDA)含量以反映神经元的氧化应激水平。结果与OGD/R组比较,PRDX6-siRNA处理的神经元存活率降低,MDA增加、SOD降低、氧化应激损伤加重;与PRDX6-siRNA组比较,同时加入i PLA2活性抑制剂MJ33组神经元存活率增加,MDA降低、SOD增加、氧化应激水平下降。结论小胶质细胞PRDX6对神经元氧糖剥夺损伤有保护作用,而i PLA2活性对PRDX6的作用有影响。Objective To observe the effects of peroxiredoxin 6 （PRDX6） knockdown in the microglia on neuron viability after oxygen-glucose deprivation and reoxygenation （OGD/R）. Methods Microglia was treated with lentivirus PRDX6-siRNA and Ca2＋ -independent phospholipase A2 （ iPI_A2 ） inhibitor, l-hexadecyl-3-（ trifluoroethgl ）-sn-glycerol-2 phosphomethanol （MJ33）. Twenty-four hours later, it was co-cultured with primary neuron to establish the microglia-neuron co-culture OGD/R model. According to the different treatment of microgtia, the ceils were divided into normal group, OGD/R group, negative control-siRNA treated OGD/R group, PRDX6-siRNA treated OGD/R group and PRDX6-siRNA combined with M J33 treated OGD/R group. Western blot analysis and real-time quantitative PCR were respectively performed to detect PRDX6 protein and mRNA levels after knockdown of PRDX6 in microglia. The iPI_A2 activity was measured by ELISA. MTS and lactate dehydrogenase （LDH） assay were used to measure neuron viability and cell damage. The oxidative stress level of neuron was determined by measuring superoxide dismutase （SOD） and malonaldehyde （MDA） content. Results In PRDX6-siRNA group, neuron viability was inhibited and oxidative stress damage was aggravated compared with OGD/R group. In PRDX6- siRNA combined with MJ33 group, cell viability was promoted and oxidative stress damage was alleviated compared with PRDX6-siRNA group. Conclusion PRDX6 in microglia protects neuron against OGD/R-induced injury, and iPLA2 activity has an effect on PRDX6.

关 键 词：氧糖剥夺 小胶质细胞 神经元 共培养 过氧还原素6(PRDX6) 小干涉RNA(siRNA) MJ33 

分 类 号：R392.12[医药卫生—免疫学]