原头蚴外泌体对树突状细胞活化和细胞因子分泌的影响  

Study on the influence of Protoscoleces exosome on the activation and cytokine secretion of dendritic cells

作　　者：李宗吉[1,3] 梁锦屏[1,2] LI Zong-ji;LIANG Jin-pin(Basic Medicine School of Ningxia Medical University,Yinchuan 750004,China;Key Laboratory of Clinical Pathogen Biology,General Hospital of Ningxia Medical University,Yinchuan 750004,China;Clinical Medicine School of Ningxia Medical University,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学基础医学院,宁夏银川750004 [2]宁夏医科大学总医院临床病原生物重点实验室,宁夏银川750004 [3]宁夏医科大学临床医学院,宁夏银川750004

出　　处：《中国预防兽医学报》2020年第1期70-74,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：宁夏自然科学基金面上项目(NZ17064)。

摘　　要：为分析囊型包虫原头蚴(PSC)分泌的外泌体(Exosome)对树突状细胞(DCs)活化和分泌细胞因子的影响,本研究采用高速离心法结合外泌体提取试剂盒分离原头蚴分泌的外泌体(PE),收集的PE沉淀中加入去离子水低渗裂解离心,提取超滤后的蛋白浓缩液即为原头蚴外泌体超滤裂解物(PEL)。利用免疫磁珠从健康ICR小鼠脾脏中分选DCs,分别加入PE、LPS、PEL、PBS进行刺激培养,24 h后收集培养细胞及培养上清,利用流式细胞术检测DCs表面活化分子的表达水平及细胞因子分泌情况。结果显示:4组DCs表面表达CD40、CD80、CD86、MHCⅡ的比例相比,PEL组结果明显高于PE组,这两组较PBS组均显著升高(p<0.05),但3组均低于LPS组。PE、PEL组DCs培养上清中IL-6、IL-10、IL-12、TNF-α均显著高于PBS组,且PEL组IL-12、TNF-α高于PE组,IL-6、IL-10水平低于PE组。以上结果初步证实PE能刺激DCs活化和释放细胞因子,低渗裂解超滤提纯外泌体蛋白并去除miRNA成分的PEL能够更有效激活DCs活化和促进细胞因子释放,表明外泌体蛋白可诱导抗感染免疫应答,exosomal miRNA成分可能参与虫体免疫逃逸。本研究为后续进一步探索包虫感染及抗感染免疫应答机理提供实验依据。The aim of this study is to analyze the effects of exosome secreted by Protoscoleces(PSC)on the activation and secretion of inflammatory factors in dendritic cells(DCs),and to explore the role of exosome secreted by Protoscoleces in interaction between host and parasite and immune escape Protoscoleces exosome(PE)was separated by high-speed centrifugation.The precipitation of PE was lysed by deionized water and centrifuged,the concentrated liquid extract is PE lysates(PEL).Dendritic cells(DC)were isolated from the spleens of healthy ICR mice by immunomagnetic beads.After 24 hours of stimulation and culture with PE,LPS,PEL and PBS,the cells and culture supernatants were collected.Flow cytometry was used to detect the expression of activated molecules on DC surface and the secretion of cytokines.The result showed that the expression of CD40,CD80,CD86 and MHCⅡon the surface of DCs from PEL group were significantly higher than those of PE group,moreover both of the two groups were significantly higher than those of PBS group(p<0.05),however all PE,PEL and PBS groups s were lower than those of the LPS group.The levels of IL-6,IL-10,IL-12 and TNF-αin DCs culture supernatants of PE and PEL groups were significantly higher than those of PBS group,and the levels of IL-12 and TNF-a in PEL group were higher than those in PE group,but the levels of IL-6 and IL-10 in PEL group were lower than those in PE group.These data showed that PE can stimulate the activation of DCs and release of cytokines.PEL with purified exosome protein and removal of the components of microRNA can induce more effective activation of DCs and promote the release of cytokines,suggesting that exosome protein can induce anti-infective immune response,and exosome microRNA may participate in the immune escape of worms.This study may provide theoretical and experimental data for further investigation on the immunity response of infection and anti-infection of parasites.

关 键 词：外泌体 树突状细胞 细胞因子 免疫逃逸 

分 类 号：S855.9[农业科学—临床兽医学]