晚期氧化蛋白产物诱导小胶质细胞M1型激活的研究

Advanced oxidation protein products induce M1 activation of BV2 cells： an experimental study

目的观察晚期氧化蛋白产物（AOPPs）能否诱导小胶质细胞M1型激活。方法小鼠血清白蛋白（MSA）与次氯酸钠溶液反应制成AOPPs。噻唑蓝（MTr）法检测经不同浓度AOPPs处理24h后BV2细胞的活力，以选择AOPPs的工作浓度。根据浓度效应对BV2细胞进行分组处理，二氯荧光素双醋酸盐（DCFH-DA）法和激光共聚焦显微镜检测细胞内活性氧（ROS）水平，Westernblot技术检测细胞内还原型辅酶1I（NADPH）氧化酶4（NOX4）、中性粒细胞黏附分子（CDllb）和肿瘤坏死因子．仅（TNF-α）的表达。结果400μg／ml和800μg／mlAOPPs作用后BV2细胞活力较其他浓度明显降低[（50．052±2．163）％、（36．505±1．589）％，P=0．000]。与对照组比较，随AOPPs作用时间的延长和浓度的增加，BV2细胞胞内ROS产生增多，NOX4、CD11b和TNF-α蛋白表达量均增加（ROS：P=0．000；NOX4：PLPs=0．001，P50μg／ml：0．000，P100μg／ml=0．004，P200μg／ml=0．000；CDllb：P=0．000：TNF-α：P=0．000）。结论AOPPs可以通过激活BV2细胞生成ROS产生M1型激活。

Objective To investigate whether advance oxidation protein products （AOPPs） would induce M1 activation of BV2 cells. Methods AOPPs were prepared by incubation of mouse serum albu- min （MSA） with sodium hypochlorite solution. Methyl thiazol tetrazolium （MTT）assay was used to deter- mine the effects of different concentration of AOPPs on BV2 cells activity by incubating the cells for 24 h for selecting optimum working concentration of AOPPs. BV2 cells were treated based on concentration effect. Intracellular reactive oxygen species （ROS） production was determined with dichlorofluorescein diaeetate （DCFH -DA） and Confocal laser scanning microscope system. And the protein expression of nicotinamide adenine dinucleotide （ NADPH ） oxidase （ NOX ） 4 （ NOX4 ） , cluster of differentiation molecule 11 b （CD1 l b） and tumor necrosis factor （TNF -α） were examined by Western blotting. Results Viability of BV2 cells treated with 400 μg/ml and 800μg/ml AOPPs was significantly lowered [ （50. 052 ± 2. 163）%, （36.505 ＋ 1.589）%, P= 0.000]. AOPPs, in a concentration- dependent manner, in- creased the expression of NOX4, CDllb and TNF - α （NOX4： PEts = 0.001, P50μg/ml = 0.000, P100μ = 0. 004, P200μg/ml=0. 000 ; CD11 b ： P = 0. 000 ; TNF - ct ： P = 0. 000 ） ; prolonged exposure to AOPPs and increase of AOPPs concentration both led to an increase of intracellular ROS production （P=0. 000）. Conclusion AOPPs may induce M1 activation of BV2 cells through producing ROS.