CXCR2拮抗剂抑制NETs缓解实验性自身免疫性脑脊髓炎病情  

CXCR2 antagonist inhibits NETs and relieves experimental autoimmune encephalomyelitis

作　　者：刘香易 彭靖兰 杨长传 陈思源 魏霞蔚 严亨秀[1] LIU Xiangyi;PENG Jinglan;YANG Zhangchang;CHEN Siyuan;WEI Xiawei;YAN Hengxiu(School of Life Science and Technology,College of Pharmacy,Southwest Minzu University,Chengdu 610041,China;State Key Laboratory of Biotherapy/Collaborative Innovation Center,West China School of Pharmacy,Sichuan University,Chengdu 610041,China)

机构地区：[1]西南民族大学生命科学与技术学院/药学院,成都610041 [2]四川大学华西药学院生物治疗/协同创新中心国家重点实验室,成都610041

出　　处：《免疫学杂志》2021年第6期481-488,共8页Immunological Journal

基　　金：四川省科技厅应用基础研究项目(2021YJ0256);中央高校基本科研业务费专项基金项目(2018NZD16)。

摘　　要：目的探讨CXCR2拮抗剂通过减少中性粒细胞迁移并抑制中性粒细胞胞外诱捕网(neutrophil extracellular traps,NETs)形成对实验性自身免疫性脑脊髓炎(experimental autoimmune encephalomyelitis,EAE)的保护作用。方法用髓鞘少突胶质细胞糖蛋白_(35-55)(myelin oligodendrocyte glycoprotein_(35–55),MOG_(35-55))诱导建立多发性硬化症(multiple sclerosis,MS)的小鼠疾病模型EAE。将C57BL/6小鼠随机分为正常对照组(n=7)、EAE模型组(n=7)、CXCR2拮抗剂处理组(n=7),CXCR2拮抗剂于造模后每天腹腔注射1次(4 mg/kg),持续给药23 d,每日进行神经功能评分;在发病高峰期取小鼠脑和脊髓,采用HE染色进行组织病理评分;取免疫后第7天和第14天小鼠血液和脑、脊髓、脑膜、骨髓等组织,采用流式细胞术分析不同部位的中性粒细胞的百分比,脑组织酯酶染色观察中性粒细胞数量的变化,脑和脊髓组织免疫荧光染色观察NETs的变化;使用伊文思蓝染色的方法检测血脑屏障的通透性。结果CXCR2拮抗剂治疗组发病延迟4 d,其临床评分显著低于EAE模型组;同时其脊髓和脑组织的淋巴细胞浸润和"血管袖套"样结构明显降低、炎性评分与EAE组相比有显著性差异。CXCR2拮抗剂抑制中粒细胞(CD45^(high)CD11b^(+)Ly6G^(+))在脑膜的早期浸润;流式细胞术及酯酶染色均显示脑组织和脊髓的中性粒细胞的浸润也被抑制;脑和脊髓CD31^(+)Ly6G^(+)细胞减少;EAE模型小鼠脑和脊髓NETs(NE^(+)MPO^(+))显著高于正常小鼠,而被CXCR2拮抗剂显著抑制,且伊文思蓝在脑实质的含量也显著降低。结论CXCR2拮抗剂能够有效地延缓和减轻小鼠的临床表现和中枢神经系统炎性细胞的浸润;通过阻止中性粒细胞跨脑膜向中枢神经系统迁移以及抑制炎症部位NETs的形成来缓解病情,并减轻了对血脑屏障通透性的破坏,提示CXCR2抑制剂可成为中枢神经系统炎性疾病治疗的另一种潜在方案。This study was designed to explore the protective effect of CXCR2 antagonists on experimental auto-immune encephalomyelitis(EAE) by reducing neutrophil migration and inhibiting neutrophil extracellular traps(NETs). EAE model was induced by myelin oligodendrocyte glycoprotein_(35-55)(MOG_(35-55)). C57 BL/6 mice were random-ly divided into normal control group(n=7), EAE model group(n=7) and CXCR2 antagonist group(n=7), CXCR2 an-tagonist was injected intraperitoneally(4 mg/kg) once a day for 23 days after the model was constructed. Neurologi-cal function scoring was performed daily;histopathological changes were observed by HE staining;the number of neutrophils and extracellular trap(NETs) in central nervous system was observed by flow cytometry, esterase stain-ing and immunofluorescence. Furthermore, the permea-bility of blood-brain barrier was detected by Evans bluestaining. Compared with EAE model group, CXCR2 an-tagonist treatment could effectively relieve the clinicalmanifestations of EAE mice, alleviate the pathological in-jury in brain and spinal cord, significantly reduce neutro-phil infiltration and NETs formation in brain and spinalcord, and down-regulate the content of Evans blue in brain parenchyma significantly. Taken together, CXCR2 antag-onists can effectively alleviate the clinical manifestations of EAE mice, preserve the blood-brain barrier function,and reduce the accumulation of neutrophils and NETs in inflammatory sites, which may represent another potentialtreatment for inflammatory diseases of the central nervous system.

关 键 词：EAE模型 CXCR2拮抗剂 中性粒细胞 中性粒细胞胞外诱捕网 血脑屏障 

分 类 号：R392.5[医药卫生—免疫学]