肥大细胞对重组口蹄疫病毒VP1-VP4应答的蛋白质表达谱检测  

Protein expression profile of mast cells in response to recombinant VP1-VP4 of foot-and-mouth disease virus

作　　者：王燕[1] 张莎莎[1] 王蕾[1] 刘雪芳[1] 张恩[1] 李娜[1] 高云欢[1] 张义明[1] 李丽敏[1] 王家鑫[1] 

机构地区：[1]河北农业大学动物医学院免疫学实验室,河北保定071000

出　　处：《生物工程学报》2016年第9期1194-1203,共10页Chinese Journal of Biotechnology

基　　金：国家自然科学基金(No.31402174);河北省高等学校专业综合改革项目;河北省教育厅重点项目(No.ZD2015040)资助~~

摘　　要：为揭示肥大细胞抗口蹄疫病毒VP1-VP4蛋白的天然免疫作用,以重组口蹄疫病毒VP1-VP4蛋白刺激小鼠腹腔肥大细胞(Peritoneal mast cells,PMCs),用高通量ELISA芯片检测PMCs的蛋白质表达谱。结果显示,VP1-VP4蛋白刺激的PMCs(VP1-VP4组)表达CCL19、L-selectin、CCL17和TNF-α的水平极显著低于对照组(PMCs)(P<0.001),而VP1-VP4蛋白刺激经甘露糖受体(Mannose receptor,MR)抑制剂预处理的PMCs(MR组)表达CCL19、IL-15、IL-9、G-CSF和Galectin-1的水平则极显著高于对照组(P<0.01),IL-10表达水平也有显著升高(P<0.05)。MR组与VP1-VP4组相比,PMCs表达IL-10、IL-17、CCL20、IL-15、IL-9、L-selectin、CCL17、TNF-α和CCL19的水平极显著升高(P<0.01),CCL21和G-CSF的表达也显著高于VP1-VP4组(P<0.05)。生物信息学差异表达分析结果显示,与对照组相比,VP1-VP4组PMCs表达的L-selectin和CCL17为下调性差异表达蛋白(Log2(ratio)≤–1)。MR组与VP1-VP4相比,PMCs表达的CCL20、CCL19、L-selectin和IL-15为上调性差异表达蛋白(Log_2(ratio)≥1)。这表明,PMCs可自发分泌CCL19、L-selectin、CCL17和TNF-α,而VP1-VP4则对PMCs的天然免疫功能具有抑制作用。由于阻断MR后PMCs的蛋白质表达水平显著升高,所以VP1-VP4对小鼠PMCs的免疫抑制作用可能是由MR介导的。To reveal the innate immunity of mast cells against recombinant VP1-VP4 protein of foot-and-mouth disease virus（FMDV）, mouse peritoneal mast cells（PMCs） were pulsed with recombinant VP1-VP4 protein. The supernatants harvested from PMCs cultures were applied to the high throughput ELISA array. Our results show that the expression levels of CCL19, L-selectin, CCL17, and TNF alpha released from PMCs pulsed with recombinant VP1-VP4 were significantly down-regulated compared with PMCs alone（P0.001）. Surprisingly, in comparison with PMCs alone, the expression levels of CCL19, IL-15, IL-9, G-CSF, and Galectin-1 in PMCs with the mannose receptor（MR） inhibitor were significantly up-regulated（P0.01）, and the expression level of IL-10 was also remarkably up-regulated（P0.05）. Importantly, the protein expression levels in PMCs treated with MR inhibitor were higher than PMCs pulsed with VP1-VP4, including IL-10, IL-17, CCL20, IL-15, IL-9, L-selectin, CCL17, TNF alpha, and CCL19（P0.01） as well as CCL21, and G-CSF（P0.05）. Differential expression analysis in bioinformatics shows that both L-selectin and CCL17 were recognized as differentially expressed protein molecules（Log2（ratio）≤–1） when compared with PMCs alone. Furthermore, the up-regulation of the expression levels of CCL20, CCL19, L-selectin, and IL-15 in PMCs treated with MR inhibitor was defined as differential expression（Log_2（ratio）≥1）. These data indicate that PMCs are capable of secreting CCL19, L-selectin, CCL17, and TNF alpha spontaneously and the recombinant VP1-VP4 has an inhibitive potential to PMCs during their performance of innate immune response. Given the protein expression levels from PMCs pre-treated with MR inhibitor were significantly increased, it can be deduced that immunosuppression of FMDV is presumably initiated by the VP1 recognition of MR on mast cells.

关 键 词：肥大细胞 甘露糖受体 口蹄疫病毒 天然免疫 差异表达 

分 类 号：S852.65[农业科学—基础兽医学]