重组IFN-α-IL-18体外对鸡外周血淋巴细胞转化及核因子κB的影响  

Effects of recombinant IFN-α-IL-18 on chicken lymphocyte transformation and NF-κB activity in vitro

作　　者：杨惠[1] 李银聚[1] 刘一尘[1] 程相朝[1] 杨丹芳[1] 韩海锋[1] 金修哲[1] 

机构地区：[1]河南省动物疫病与公共安全院士工作站/河南科技大学动物疫病与公共卫生重点实验室,洛阳471003

出　　处：《中国免疫学杂志》2015年第8期1040-1044,1048,共6页Chinese Journal of Immunology

基　　金：河南省科技攻关项目(132102110117)

摘　　要：目的:探讨鸡IFN-α-IL-18融合蛋白对鸡外周血淋巴细胞转化及核因子κB(NF-κB)的影响。方法:构建重组酵母表达质粒p PICZ-IFN-α、p PICZ-IL-18和p PICZ-IFN-α-IL-18,电转化酵母菌X-33后以1%甲醇诱导,SDS-PAGE和Western blot鉴定表达IFN-α、IL-18及IFN-α-IL-18蛋白,淋巴细胞转化试验初步检测表达产物生物学活性,双抗体夹心法测定不同时段淋巴细胞中总NF-κB和细胞核中NF-κB的含量。结果:酵母培养上清液中存在分子量分别为22、23和43 k D的目的蛋白;表达的IFN-α-IL-18和IL-18均能明显促进鸡外周血淋巴细胞转化(P<0.05),IFN-α对淋巴细胞转化有促进作用,但总体差异不显著(P>0.05)。与对照组相比,经IFN-α-IL-18和IL-18作用的鸡淋巴细胞中总NF-κB含量明显增加(P<0.05),细胞核中NF-κB含量差异极显著(P<0.01),IFN-α作用的淋巴细胞中总NF-κB含量与对照组相比差异不显著(P>0.05),细胞核中NF-κB含量显著增加(P<0.05)。结论:IFN-α-IL-18及IL-18能够明显促进淋巴细胞转化,IFN-α促进淋巴细胞转化活性较弱,这种促淋转机制与NF-κB的表达、活化及核质转运有关,为进一步研究IFN-α-IL-18融合蛋白的功能及其在疫病防控中的作用奠定了基础。Objective： To discuss the effects of recombinant IFN-α-IL-18 fusion protein on chicken lymphocyte transformation and the chicken nuclear factor kappa B（ NF-κB） activity. Methods： The recombinant plasmids p PICZ-IFN-α,p PICZ-IL-18 and p PICZ-IFN-α-IL-18 were constructed,and transformed into P. Pastoris X-33 strain by electroporation. The recombinant proteins were expressed under the induction of 1% methanol,and detected by SDS-PAGE and Western blot. The biological activities of the expressed products were detected by the lymphocyte transformation test,the NF-κB concentration in chicken lymphocyte and lymphocytic nucleus were detected at different times with ELISA. Results： SDS-PAGE and Western blot showed that the expressed products existed in supernatant,and the molecular weights were about 22 k D,23 k D and 43 k D,respectively. The expressed products IL-18 and IFN-α-IL-18 could stimulate chicken lymphocyte transformation（ P〈0. 05）,the biological activity of IFN-α stimulating lymphocyte transform was feeble（ P〉0. 05）. Compared with the control group,the total NF-κB concentration in chicken lymphocyte induced by IL-18 and IFN-α-IL-18 were increased（ P〈0. 05）,and the NF-κB in lymphocytic nucleus was increased significantly（ P〈0. 01）. Otherwise,the total NF-κB in lymphocyte induced by IFN-α increase was limited（ P〈0. 05）,but the NF-κB in lymphocytic nucleus showed the remarkable increase（ P〈0. 05）. Conclusion： IFN-α-IL-18 and IL-18 can promote lymphocyte transformation significantly,the activity of IFN-α induced lymphocyte transformation is imperfect. The biological activity of stimulating lymphocyte transformation is associated with the NF-κB expression,activation and nucleo-cytoplasmic transport. The study built foundation for the function of IFN-α-IL-18 fusion protein and the exploration of the role of controlling epidemic diseases.

关 键 词：IFN-α-IL-18 酵母表达 淋巴细胞转化试验 核因子ΚB 

分 类 号：S852.4[农业科学—基础兽医学]