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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 plays an important regulatory role in immune cells such as dendritic cells, macrophages and T cells. The recombinant mouse IL-33 was prepared by using Escherichia coli, and its potential and characteristics as a mucosal immune adjuvant were preliminary investigated. Methods: The expression of Trx / IL-33 fusion protein in E. coli DH5α was induced by IPTG. The Trx / IL-3 fusion protein was purified by QSepharose ion exchange and Ni ~ (++) metal chelate affinity chromatography. 33. Further cleavage by enterokinase results in the mature form of IL-33. The mice were immunized intranasally with recombinant HBcAg mixed with purified IL-33 and the HBcAg-specific IgA and IgG1, IgG2a responses were investigated. Results: The purified recombinant IL-33 had the same cell-like biological activity as that of the standard product, indicating that macrophage RAW264.7 expressed TNF-α in vitro. As an adjuvant, HBcAg-specific IgA response in mucosal tissues immunized with nasal mucosa immunized by nasal mucosa and the response level of specific IgG1 in serum and bronchoalveolar lavage fluid were significantly increased as adjuvant, while IgG2a was suppressed. Conclusion: The active IL-33 can be prepared by using Escherichia coli, which has the potential application of mucosal immune adjuvant.