文章摘要
柳昭君,赵杰,卜云璇,肖兰莹,时凯丽,宋学宏.IL-8在LPS诱导的草鱼炎症过程中的表达特征[J].水产学报,2017,41(7):1028~1035
IL-8在LPS诱导的草鱼炎症过程中的表达特征
Expression patterns of IL-8 protein during bacterial LPS-induced inflammatory response in grass carp (Ctenopharyngodon idella)
投稿时间:2016-02-03  修订日期:2016-09-04
DOI:10.11964/jfc.20160210278
中文关键词: 草鱼  白细胞介素-8  多克隆抗体  细菌脂多糖  组织表达  炎症应答
英文关键词: Ctenopharyngodon idella  interleukin-8 (IL-8)  polyclonal antibody  bacterial lipopolysaccharide  tissue expression  inflammatory response
基金项目:江苏省自然科学基金(BK2011285);苏州市应用基础(SYN201505)
作者单位E-mail
柳昭君 苏州大学基础医学与生物科学学院, 江苏 苏州 215123  
赵杰 苏州大学基础医学与生物科学学院, 江苏 苏州 215123  
卜云璇 苏州大学基础医学与生物科学学院, 江苏 苏州 215123  
肖兰莹 苏州大学基础医学与生物科学学院, 江苏 苏州 215123  
时凯丽 苏州大学基础医学与生物科学学院, 江苏 苏州 215123  
宋学宏 苏州大学基础医学与生物科学学院, 江苏 苏州 215123 songxh@suda.edu.cn 
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中文摘要:
      采用原核表达获得草鱼白细胞介素-8(IL-8)重组蛋白,并以腹腔与皮下组织交替注射的方法免疫小鼠,制备抗草鱼IL-8的多克隆抗体。采用免疫印迹(Western blot)和酶联免疫吸附(ELISA)技术检测抗体的特异性与效价,运用流式细胞术分析健康草鱼及细菌脂多糖(LPS)刺激后草鱼各免疫相关组织中IL-8的表达特征。结果显示,草鱼IL-8多克隆抗体效价可达1∶40 000。健康草鱼的胸腺、头肾、肝脏、肠道、鳃、脾脏、体肾等组织均表达IL-8蛋白,其中,头肾和鳃中表达量较高。LPS刺激后,各组织IL-8蛋白表达量均显著升高,其中,肠道、肝脏、肾脏在LPS刺激4 h后IL-8蛋白上调达最高峰,头肾、脾脏、鳃于LPS刺激后12 h时表达量最高,胸腺则在24 h时达最高峰。研究表明,IL-8参与了草鱼炎症应答;IL-8表达量的升高,可作为早期炎症监测指标,应用于养殖鱼类炎症性疾病的预警指标。
英文摘要:
      In order to investigate the expression characteristics of IL-8 during the lipopolysaccharide (LPS)-induced inflammatory process in grass carp (Ctenopharyngodon idella), recombinant grass carp IL-8 (rgcIL-8) was generated in prokaryotic expression system, and an anti-rgcIL-8 polyclonal antibody was prepare by immunizing mice with the purified rgcIL-8 alternately via intraperitoneal and subcutaneous injections. The polyclonal antibody was validated for specificity and titer by Western blotting and enzyme-linked immunosorbent assay (ELISA), and was used to detect IL-8 in immune-related tissues from healthy and/or LPS-stimulated grass carp by flow cytometry analysis. Results showed that the antibody titer peaked at 40 000. Flow cytometry analysis confirmed that IL-8 protein was expressed constitutively in all tested tissues of healthy fish, with higher levels in head kidney and gill. Moreover, the expression of IL-8 protein was found to be significantly up-regulated by LPS stimulation in all tested fish tissues, the protein levels peaked in intestine, liver and trunk kidney at 4 h, in head kidney and spleen at 12 h, and in thymus at 24 h following LPS stimulation, respectively. Our results provide further evidence that IL-8 is a key player in host inflammatory responses. Therefore, it is proposed that an elevated expression level of IL-8 protein could be used as a marker for monitoring the early inflammation, and be considered as a risk indicator for inflammatory diseases in aquaculture.
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