重组小鼠白细胞介素-33的原核表达制备及其粘膜免疫佐剂活性  

The Prokaryotic Expression of Recombinant Murine Interlukin(IL)-33 and Its Mucosal Immunization Affect as Adjuvant

作　　者：冯雪军[1,2,3] 龙琼 唐增华[1,2,3] 黄惟巍 刘存宝 杨旭[1,2,3] 孙文佳 白红妹[1,2,3] 马雁冰 

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所,昆明650118 [2]云南省重大传染病疫苗研发重点实验室,昆明650118 [3]云南省重大传染病疫苗工程技术研究中心,昆明650118

出　　处：《中国生物工程杂志》2017年第4期26-32,共7页China Biotechnology

基　　金：云南省对外合作科技项目(2013IA005);云南省应用基础研究重点项目(2011FA023;2016FA023)资助项目

摘　　要：目的:白细胞介素(IL)-33对树突状细胞、巨噬细胞及T细胞等免疫细胞具有重要调控作用。利用大肠杆菌制备重组小鼠的IL-33,并初步考察其作为粘膜免疫佐剂应用的潜能与特点。方法:以IPTG诱导硫氧还蛋白(Trx)/IL-33融合蛋白在大肠杆菌DH5α中的表达,并通过QSepharose离子交换和Ni^(++)金属螯合亲和层析纯化Trx/IL-33,进一步经肠激酶切割获得成熟形式的IL-33。重组HBcAg混合纯化的IL-33后经滴鼻免疫小鼠,考察HBcAg特异的IgA及IgG_1、IgG_(2a)的应答。结果:纯化的重组IL-33具有与标准品相当的促巨噬细胞RAW264.7表达TNF-α的体外细胞生物学活性。作为佐剂可显著增强滴鼻粘膜免疫激发的不同粘膜组织中HBcAg特异的IgA应答,以及血清与支气管肺泡灌洗液中特异IgG_1的应答水平,而抑制IgG_(2a)应答。结论:利用大肠杆菌可制备活性IL-33,其具有粘膜免疫佐剂的应用潜能。Objective： Interleukin （IL）-33 has important immunoregulatory effects on immune eel]s, such as dentritie cells, macropphage and T cells. The purpose is to prepare recombinant murine IL-33 in E. coli cells, and investigate the potentials and characteristics of IL-33 to be employed as an adjuvant in mucosal immunization. Methods： The expression of fusion protein thioredoxin/IL-33 was induced by IPTG in DH5a cells, and the purification was performed through a chromatography procedure consists of Q-Sepharose ionic exchange and Ni ~ ~ chelating affinity. And then, mature IL-33 was released from the fusion protein by enterokinase treatment. The recombinant HBcAg was combined with IL-33 or not, and used to immunize mice intranasally. HBcAg-specific IgA, IgG1 and IgG2~ were detected with ELISAs. Results： Purified IL-33 displayed the similar bioactivity of stimulating RAW264.7 to express TNF-ct as commercially derived recombinant IL-33. Purified IL- 33 which served as an adjuvant significantly enhanced the HBeAg-specific IgA response in multiple mucosal tissues, as well as IgG1 levels in serum and bronchial alveolar lavage fluid, whereas reduced IgG2a levels. Conclusion ： Active IL-33 is successfully prepared from E. coli cells, and has a potential to be used as a molecular adjuvant in mucosal immunization.

关 键 词：重组 白细胞介素-33 表达 粘膜免疫 佐剂 

分 类 号：Q819[生物学—生物工程]