以pcD-IL-2为基因佐剂的含日本血吸虫SjFABPc基因真核表达重组质粒裸DNA免疫小鼠的研究

A STUDY ON DNA-BASED IMMUNIZATION WITH EUKARYOTIC EXPRESSION RECOMBINANT PLASMID CONTAINING GENE ENCODING SCHISTOSOMA JAPONICUM FATTY ACID-BINDING PROTEIN AND CO-DELIVERY OF GENETIC ADJUVANT PCD-IL-2 IN MICE

目的 在小鼠模型中评价含日本血吸虫脂肪酸结合蛋白分子基因的真核表达质粒pcD -SjFABPc与含IL - 2基因佐剂的质粒构建体 pCD -IL - 2经肌注、皮内途径导入机体后所诱生的免疫反应。 方法 碱裂解法大量制备重组质粒 pcD-SjFABPc、pCD -IL - 2重组质粒及空质粒 pcDNA3,经肌肉及皮内注射免疫BALB/c小鼠 ,每只鼠注射 10 0 μg ,两周后同量加强免疫一次。分别于末次免疫后的第 2 0d、5 0d及 65d用MTT法检测脾脏T淋巴细胞增殖与NK细胞活性 ,并用双夹心ELISA测定血清细胞因子IL - 2、IFN -γ及IL - 10含量 ;ELISA法测定免疫鼠血清IgG抗体。 结果 NK细胞杀伤活性及脾淋巴细胞增殖测定 ,加佐剂组 ( pcD -SjFABPc联合 pCD -IL - 2组 )较不加佐剂组 ( pcD -SjFABPc组 )NK细胞杀伤及脾淋巴细胞增殖活性皆增加 (P <0 .0 5 )。两途径三次检测IL - 2及IFN -γ值均明显高于NS对照组和空质粒组 ,且加佐剂组的明显高于不加佐剂组的 (P <0 .0 5 ) ;不同途径各组IL - 10值与NS及空质粒对照组的比较则无显著性差别 (P >0 .0 5 )。注射质粒后 2 0d和 5 0d ,未测出抗体 ;免疫后 65d检测 ,pcD -SjFABPc和pcD -SjFABPc联合 pcD -IL - 2免疫组的OD值明显高于对照组 (P <0 .0 1) ,而该两组OD490 值之间无显著性差异 (P >0 .0

Aim To,evaluate the immune responses of mice by intramuscular or intracutaneous injection immunized with DNA-bsaed plasmid pcD-SjFABPc and co-delivery of pcD-IL-2.Methods Large scale preparation of plasmids DNA by alkaline lysis,mice were inoculated DNA plasmids by intramuscular and intracutaneous injection at the dosage of 100μg each mouse respectively.A booster vaccine was given with the same dosage after two weeks.Control groups were injected with pcDNA 3 plasmid only or normal saline respectively.20,50 and 65 days after the final booster injection,following tests were carried out 3 times separately.The splenocytes from experimental mice were used to detect the killing activity of NK cells and the proliferation activity of T lymphcytes by means of MTT assay,the sera from the mice were used to detect the cytokines,IFN-γ,IL-2 and IL-10 by means of sandwich ABC-ELISA,as well as the titer of IgG antibody by ELISA.Results The activities of NK and proliferation of spleen T lymph cells by MTT assay in the pcD-SjFABPc plus pcD-IL-2 experiment group were higher than those of in the only pcD-SjFABPc groups( P<0.05 ).The levels of IL-2 and IFN-γ in pcD-SjFABPc co-administration with pcD-IL-2 immunized mice were increased significantly compared with those of pcD-SjFABPc ( P<0.05).There was no significant change in IL-10 among these groups(P>0.05).After 65 days of immunization,the OD490 values of IgG antibodies between pcD-SjFABPc group and pcD-SjFABPc co-administration with pcD-IL-2 one were not significant difference(P>0.05).Conclusion Genetic adjuvant pcD-IL-2 could enhance effectiveness of cellular immuno-response in DNA immunized mice,as well as the same profile of products of cytokines.There was no significant effect on enhancement of production of specific IgG antibodies.