摘要
目的在离体细胞水平鉴定脑缺血/再灌注损伤中产生白细胞介素-17A(IL-17A)的脑固有神经细胞类型。方法利用原代培养小鼠脑皮层神经元、小胶质细胞和星形胶质细胞1-4 h氧-糖剥夺/24 h复糖复氧(OGD/R)模拟细胞离体缺血/再灌注损伤模型,用免疫荧光双标、实时定量聚合酶链式反应(RT-qPCR)和蛋白免疫印迹(Western blot)等技术,确定产生IL-17A的神经细胞类型。结果三种神经细胞中,除NeuN阳性神经元外,小胶质细胞和星形胶质细胞在OGD/R处理后,均可表达IL-17A蛋白,且分别与其特定标志物Iba-1和GFAP存在共定位现象。星形胶质细胞经过1-6 hOGD/24 h R处理后,IL-17A mRNA表达水平随OGD时间延长显著增高,且在4 hOGD/R时达高峰[(2.74±2.48),P〈0.001,每组n=5]。此外,OGD/R可促进星形胶质细胞表达IL-17A蛋白[(3.17±0.91),P〈0.05,每组n=5]。结论脑缺血/再灌注损伤中星形胶质细胞可能是产生IL-17A的主要来源。
Objective To identify the neural cell-type as a source of IL-17 A during cerebral ischemia/reperfusion injuries in vitro. Methods Primary cultured mouse cortical neurons,microglia and astrocytes were subjected to 1 - 4 h oxygen-glucose deprivation/24 h reoxygenation( OGD/R) simulating cell ischemia/reperfusion injury model in vitro,and then the double immunofluorescence staining with IL-17 A and their specific markers of NeuN,Iba-1 and GFAP,RT-qPCR and Western blot were performed to determine the neural cell-type of IL-17 A production. Results The primary cultured microglia and astrocytes( but not NeuN + neurons) expressed IL-17 A and respectively co-localized with their specific markers Iba-1 and GFAP after OGD/R treatment. The mRNA expression level of IL-17 A increased withOGD duration and reached the peak at 4 hOGD [( 2. 74 ± 2. 48),P 0. 001,n = 5 per group]in astrocytes after1 - 6 hOGD/24 h R treatment. In addition,4 hOGD/R treatment could promote IL-17 A protein expression in primary cultured astrocytes [( 3. 17 ± 0. 91),P〈 0. 05,n = 5 per group]. Conclusions Astrocytes is a potential source of IL-17 A production during cerebral ischemia/reperfusion injuries.
出处
《基础医学与临床》
CSCD
2016年第12期1618-1623,共6页
Basic and Clinical Medicine
基金
国家自然科学基金(31471142
81301015)
北京市自然科学基金重点项目(7141001)