右美托咪定预处理对LPS激活星形胶质细胞HMGB1基因表达的影响

陈筱诗 肖莹莹 周娇 曾文强 杨进国

[摘要] 目的 觀察右美托咪定（DEX）预处理对细菌脂多糖（LPS）激活的星形胶质细胞（AS）高迁移率族蛋白1（HMGB1）在基因水平表达的影响，探究其与烟碱样乙酰胆碱受体α7亚型（α7nAChR）的关系。 方法 选取新生1～2 d的SD大鼠，分离、培养大脑皮质AS。将细胞接种于细胞培养板上，按照随机数字表法分为空白对照组、LPS组、DEX预处理组、α银环蛇毒素（α-BGT，为α7nAChR拮抗剂）组和α-BGT预处理组。免疫细胞化学技术检测AS特异性标志物胶质纤维酸性蛋白（GFAP）的表达，甲基噻唑基四唑（MTT）法检测细胞活力，实时荧光定量PCR检测各组HMGB1的mRNA表达水平。 结果 第三代AS纯度达95%以上。活化的AS HMGB1 mRNA表达水平上调，与空白对照组比较差异有高度统计学意义（P < 0.01）。与LPS组比较，DEX预处理组HMGB1 mRNA表达明显降低（P < 0.01）。与DEX预处理组比较，α-BGT预处理有显著逆转DEX预处理的作用（P < 0.01），但是α-BGT单独作用于AS则对HMGB1基因表达并无影响（P > 0.05）。 结论 DEX预处理下调HMGB1表达，阻断α7nAChR后DEX预处理作用消失，提示DEX预处理的抗炎作用与其激活α7nAChR相关。

[关键词] 星形胶质细胞；脂多糖；右美托咪定；高迁移率族蛋白1；神经炎

[中图分类号] R741 [文献标识码] A [文章编号] 1673-7210（2018）04（c）-0008-05

Effect of Dexmedetomidine pretreatment on HMGB1 mRNA expression in LPS-stimulated astrocytes

CHEN Xiaoshi1 XIAO Yingying1 ZHOU Jiao2 ZENG Wenqiang1 YANG Jinguo1

1.Department of Anesthesiology， Affiliated Dongfeng Hospital， Hubei University of Medicine， Hubei Province， Shiyan 442008， China； 2.Department of Urology， Affiliated Dongfeng Hospital， Hubei University of Medicine， Hubei Province， Shiyan 442008， China

[Abstract] Objective To investigate the effect of Dexmedetomidine pretreatment on the high mobility group box 1 （HMGB1） mRNA expression in lipopolysaccharide （LPS）-stimulated astrocytes （AS）， and to explore its relationship with α7 subtype of nicotinic acetylcholine receptor （α7nAChR）. Methods The new-born 1-2 day SD rats were selected， and the AS of cerebral cortex was isolated and cultivated. The cells were inoculated on cell culture plates， and they were divided into blank control group， LPS group， DEX-pretreatment group， α-bungatotoxin （α-BGT， which was α7nAChR antagonist） group and α-BGT-pretreatment group. The immunocytochemical technique was used to determine the expression of AS specific marker glial fibrillary acidic protein （GFAP）. Methyl thiazolyl tetrazolium （MTT） was to used detect cytoactive. Real-time quantitative PCR was used to detect the mRNA expression levels of HMGB1. Results The purity of the 3rd generation of AS reached above 95%. The expression of AS HMGB1 mRNA was up-regulated， which had statistically significant differences compared with those of blank control group （P < 0.01）. Compared with LPS group， the expression of HMGB1 mRNA in DEX-pretreatment group was decreased significantly （P < 0.01）. Compared with DEX-pretreatment group， α-BGT-pretreatment had significant effects of reversing DEX-pretreatment （P < 0.01）， while single application of α-BGT for AS had no significant effects on the expression of HMGB1 gene （P > 0.05）. Conclusion Dexmedetomidine pretreatment can reduce the expression of HMGB1， the DEX-pretreatment effect disappears after cutting off α7nAChR， which indicates that the anti-inflammatory action of DEX-pretreatment may be related to activating α7nAChR.

[Key words] Astrocyte； Lipopolysaccharide； Dexmed？鄄etomidine； High mobility group box 1； Neuritis

在神经退行性疾病和脑卒中等疾病中，神经炎是其中一个重要的病理过程，涉及脑中小胶质细胞和星形胶质细胞（astrocyte，AS）的激活[1]。在神经系统与先天免疫系统相互作用对抗全身炎性反应的过程中，胆碱能抗炎通路起到重要的桥梁作用[2]。越来越多的证据表明，烟碱样乙酰胆碱受体α7亚型（α7nAChR）在介导胆碱能抗炎通路中扮演关键角色[3]。已知α7nAChR激活后抑制高迁移率族蛋白1（high mobility group box 1，HMGB1）的释放[4]。HMGB1是重要的晚期炎症介质，是阿尔兹海默病、帕金森病、脊髓损伤和脑出血等疾病的一个危险因子。有研究[5]指出，右美托咪定（DEX）通过迷走神经和α7nAChR依赖性机制抑制全身反应。……