白藜芦醇对氧糖剥夺/再复氧损伤后小胶质细胞系N9活化的影响

doi:10.16098/j.issn.0529-1356.2019.02.001

摘要/Abstract

摘要：

目的探讨白藜芦醇对氧糖剥夺/再复氧损伤（OGD/R）后小胶质细胞系N9活化的影响。方法体外培养的N9小胶质细胞行氧糖剥夺150 min，复氧培养24 h。实验分为正常组（Nor）、对照组(Ctrl)和白藜芦醇预处理组(Res)。细胞计数盒-8（CCK-8）法测细胞活力，流式细胞仪检测细胞凋亡，免疫荧光法检测Iba1蛋白表达，Western blotting检测钙离子接头蛋白1（Iba1）、CD11b、Caspase-3、Bax、白细胞介素-10(IL-10)、肿瘤坏死因子α（TNF-α）和IL-1β蛋白表达，ELISA法检测培养细胞上清液中IL-10、TNF-α和IL-1β蛋白含量。结果 CCK-8 法检测结果显示，OGD/R损伤后对照组、1、5、20、40和80 μmol/L白藜芦醇组细胞活力均较正常组降低（P＜0.05，n=3），但5、20和 40 μmol/L白藜芦醇组细胞活力较对照组显著增强（P＜0.05，n=3），以20 μmol/L组最强。流式细胞术、免疫荧光、Western blotting和ELISA法显示，对照组和白藜芦醇组CD11b、Iba1、Caspase-3、Bax、IL-10、TNF-α、IL-1β蛋白表达或含量或凋亡细胞百分比均显著高于正常组（P＜0.05，n=3），但白藜芦醇组除IL-10蛋白表达或含量较对照组增高外，其余均低于对照组（P＜0.05，n=3）。结论白藜芦醇预处理可抑制OGD/R后小胶质细胞的活化及凋亡，减轻炎症反应。

关键词: 白藜芦醇, 小胶质细胞, 活化, 炎症反应, 免疫印迹法

Abstract:

Objective To investigate the effect of resveratrol on the activation of microglia cell line N9 after oxygen-glucose deprivation/reoxygenation (OGD/R) injury in vitro.Methods N9 microglias were cultured under oxygen-glucose deprivation for 150 minutes and reoxygenation for 24 hours. The experiments were divided into normal (Nor), control (Ctrl) and resveratrol pretreatment group (Res). Cell viability was determined by cell counting kit-8(CCK-8) assay. The percentage of apoptosis was measured by flow cytometry. Expression of ionized calcium binding adaptor molecule-1(Iba1), CD11b, Caspase-3, Bax，interleukin-10 (IL-10), tumor necrosis factor α(TNF-α) and IL-1β proteins were detected by immunofluorescence. The levels of IL-10, TNF-α and IL-1βprotein in the supernatant were measured by ELISA. Results CCK-8 assay showed that the viability of microglia in the control and 1,5,20,40,80 μmol/L resveratrol groups significantly decreased compared with the normal group after OGD/R injury (P＜0.05，n=3), and significantly enhanced in the 5, 20, and 40 μmol/L resveratrol group than those in the control group, the peak was 20 μmol/L resveratrol group (P＜0.05,n=3). Immunofluorescence, Western blotting and ELISA or flow cytometry method showed that expressions or levels of CD11b, Iba1, Caspase3, Bax, IL-10, TNF-α, IL-1β proteins and the percent of apoptosis significantly increased in the control and the resveratrol group after OGD/R injury (P<0.05, n=3). However, except the expression or level of IL-10 protein was higher, the rests were lower in the resveratrol group than those of the control group (P<0.05, n=3). Conclusion Resveratrol pretreatment can inhibit the activation and apoptosis of microglia, reduce inflammation after OGD/R.

Key words: Resveratrol, Microglia, Activation, Inflammation, Western blotting

廖鸿雁刘杰刘菁郭霜杨琴. 白藜芦醇对氧糖剥夺/再复氧损伤后小胶质细胞系N9活化的影响[J]. 解剖学报, 2019, 50(2): 137-144.

LIAO Hong-yan LIU Jie LIU Jing GUO Shuang YANG Qin. Effect of resveratrol on activation of microglia cell line N9 after oxygen-glucose deprivation/reoxygenation injury in vitro[J]. Acta Anatomica Sinica, 2019, 50(2): 137-144.

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