氧葡萄糖剥夺-再恢复小鼠小胶质细胞的激活及Toll样受体9表达的变化  

Microglial activation and changes of Toll-like receptor 9 expression after oxygen-glucose deprivation and reoxygenation in rat

作　　者：杨碧莹[1] 马珊珊[2] 潘经锐[1] 纪原[1] 彭晴霞 王艺东[1] 

机构地区：[1]中山大学孙逸仙纪念医院神经科,广州510120 [2]中山大学中山医学院药理教研室

出　　处：《中国脑血管病杂志》2013年第2期84-88,共5页Chinese Journal of Cerebrovascular Diseases

基　　金：国家自然科学基金面上项目(81171103);广东省自然科学基金资助项目(S2011010006043)

摘　　要：目的观察氧葡萄糖剥夺-再恢复(OGDR)对小鼠BV-2小胶质细胞的激活以及Toll样受体9(TLR9)表达的影响。方法采用OGDR法建立BV-2细胞缺氧、缺糖模型,以常氧培养的BV-2细胞作为对照。使用倒置相差显微镜观察BV-2细胞在OGDR后0、6、12、24、48、72 h形态的变化,CCK8法检测OGDR后不同时间点细胞存活率的变化,反转录PCR和Western Blot检测细胞内TLR9 mRNA和蛋白的表达。结果①OGDR后BV-2细胞由原来静止的分枝状变成激活的阿米巴状。②OGDR后,细胞存活率明显下降,0 h为对照组的(65.7±9.2)%;12 h下降至最低,为对照组的(44.8±2.3)%,后逐渐升高,48、72 h分别为对照组的(60.8±10.2)%、(72.3±10.0)%。③OGDR后,随着时间的延长,TLR9 mRNA表达逐渐升高,24 h达高峰,后逐渐降低,但72 h仍高于0 h时间点的表达。TLR9蛋白表达亦呈升高趋势,在72 h达高峰。对照组各时间点TLR9 mRNA、TLR9蛋白表达差异均无统计学意义。④相同时间点的比较,OGDR组OGDR后6～72 h,TLR9 mRNA表达均明显高于对照组(P<0.05,或P<0.01,);12～72 h,TLR9蛋白表达显著高于对照组(P<0.05,或P<0.01)。结论 OGDR后BV-2细胞被激活,其细胞内TLR9表达随着时间的延长逐渐增高,可能在脑缺血-再灌注后的炎性反应中,发挥重要作用。Objective To observe the effect of oxygen-glucose deprivation and reoxygenation on microglial （BV-2 microglia） activation and the expression of Toll-like receptor 9 （TLR9） in a rat model. Methods An oxygen and glucose deprivation rat model of BV-2 microglia was induced by the oxygen-glucose deprivation and reoxygenation method. The morphological changes of the BV-2 microglia were observed with an inverted phase contrast microscope at 0, 6, 12, 24, 48, and 72 h after oxygen-glucose deprivation and reoxygenation. The CCK 8 assay was used to detect the changes in cell viability at different time points after oxygen-glucose deprivation and reoxygenation. Reverse transcription-polymerase chain reaction （RT-PCR） and Western blot were used to detect intracellular TLR9 mRNA and protein expression. Results ①The BV-2 microglia changed from a state of original resting branches to a state of activated amoeboid after oxygen-glucose deprivation and reoxygenation. ②Cell smvival rate decreased significantly after oxygen-glucose deprivation and reoxygenation, and 0 h was 65.7±9.2% of the control group ; it reached bottom at 12 h and was 44.8±2.3% of the control group, and then it increased gradually at 48 h （60.8±10.2% ） and 72 h （72.3±10.0% ）. ③With the extension of time, the TLR9 mRNA expression increased gradually after oxygenglucose deprivation and reoxygenation. It reached the peak at 24 h, and then declined gradually, but at 72 h it was still higher than that at 0 h. TLR9 protein expression also had a tendency of increasing, but it reached the peak at 72 h. There were no significant differences in the expression of TLR9 mRNA and TLR9 protein among all time points in the control group. @The comparison of the same time points：at 6 to 72 h in the oxygen-glucose deprivation and reoxygenation group, the TLR9 mRNA expression was significantly higher than that in the control group （P〈0.05 or P〈0.01） ; at 12 to 72 h, the TLR9 protein expression was significantly higher than that in the contro

关 键 词：TOLL样受体9 小神经胶质细胞 氧葡萄糖剥夺 小鼠 

分 类 号：R363[医药卫生—病理学]