日本血吸虫多价DNA疫苗的构建及免疫保护性研究

发布时间：2018-02-25 23:00

本文关键词： 日本血吸虫 IL-18 核酸疫苗免疫　出处：《吉林农业大学》2008年硕士论文　论文类型：学位论文

【摘要】： 日本血吸虫病是一种危害严重的人畜共患寄生虫病,目前仍以药物(吡喹酮)治疗为主。但是药物治疗不能阻断血吸虫病的传播与流行。因此加强血吸虫疫苗的研究是控制我国血吸虫病的有效措施。本研究应用基因工程技术构建了日本血吸虫核酸疫苗pVAX-GST、pVAX-GST-FABP和pIRESneo-GST-FABP-Sj23,并且克隆了小鼠IL-18,构建了真核表达载体pVAX-IL-18。以小鼠为动物模型评价了其免疫保护效果。日本血吸虫核酸疫苗的构建运用基因工程重组技术将日本血吸虫保护性抗原基因GST、FABP、Sj23克隆入真核表达载体pVAX和pIRESneo中,通过酶切鉴定构建成了日本血吸虫真核表达载体pVAX-GST、pVAX-GST-FABP和pIRESneo-GST-FABP-Sj23。小鼠IL-18真核表达载体的构建应用RT-PCR技术从小鼠脾淋巴细胞中扩增了小鼠的白介素-18(IL-18),并将其克隆至pMD18-T载体,经序列分析后将IL-18克隆至真核表达载体pVAX中,构建了重组质粒pVAX-IL-18。动物免疫保护性试验将构建的日本血吸虫核酸疫苗及pVAX-IL-18以肌肉注射的方式免疫BALB/c小鼠,二次免疫后,用日本血吸虫尾蚴分别攻击感染小鼠,42天后剖杀冲虫并计数。免疫试验结果表明,日本血吸虫核酸疫苗均能诱导小鼠产生特异性的IgG抗体,pIRESneo-GST-FABP-Sj23+IL-18免疫组所诱导的IgG水平最高;且免疫组都产生了较高水平的IFN-γ,其中联合IL-18组所诱导的IFN-γ明显高于单独免疫组(P0.01);在动物保护性实验中pIRESneo-GST-FABP-Sj23、pIRESneo-GST-FABP-Sj23+IL-18分别获得了45.0%、69.8%的减卵率,40.0%、47.5%的减虫率。上述结果表明日本血吸虫多价核酸疫苗在动物免疫实验中获得了较好的免疫保护效果,并且IL-18能增强日本血吸虫核酸疫苗的免疫保护作用。
[Abstract]:Schistosomiasis japonicum is a serious zoonotic parasitic disease. At present, the drug (praziquantel) is still the main treatment, but the drug treatment can not stop the transmission and prevalence of schistosomiasis. Therefore, strengthening the research of schistosomiasis vaccine is an effective measure to control schistosomiasis in China. In this study, the nucleic acid vaccine pVAX-GST-FABP and pIRESneo-GST-FABP-Sj23 of Schistosoma japonicum were constructed by genetic engineering technique, and the mouse IL-18 was cloned, and the eukaryotic expression vector pVAX-IL-18 was constructed. Construction of Schistosoma japonicum Nucleic Acid Vaccine by genetic Engineering Recombinant technique, the protective antigen gene GST-FABPN-Sj23 of Schistosoma japonicum was cloned into eukaryotic expression vectors pVAX and pIRESneo. The eukaryotic expression vectors pVAX-GST-pVAX-GST-FABP and pIRESneo-GST-FABP-Sj23 of Schistosoma japonicum were constructed by restriction endonuclease digestion. Construction of murine IL-18 eukaryotic expression vector using RT-PCR technique, the mouse interleukin-18 IL-18G was amplified from mouse spleen lymphocytes and cloned into pMD18-T vector. After sequence analysis, IL-18 was cloned into eukaryotic expression vector pVAX, and a recombinant plasmid pVAX-IL-18 was constructed. The BALB/c mice were immunized with the constructed Schistosoma japonicum nucleic acid vaccine and pVAX-IL-18 by intramuscular injection. Schistosoma japonicum cercariae were used to kill and count Schistosoma japonicum in mice after 42 days of infection. The results of immunological test showed that the mice immunized with specific IgG antibody pIRESneo-GST-FABP-Sj23 IL-18 had the highest level of IgG. The IFN- 纬 in the combined IL-18 group was significantly higher than that in the single immunization group (P0.01), and in the animal protective experiment, pIRESneo-GST-FABP-Sj23 pIRESneo-GST-FABP-Sj23 IL-18 obtained 45.09.8% oocyte reduction rate and 47.5% worm reduction rate respectively. The polyvalent nucleic acid vaccine of Schistosoma japonicum has a good protective effect in animal immune experiment. And IL-18 can enhance the immune protection of Schistosoma japonicum nucleic acid vaccine.
【学位授予单位】：吉林农业大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392

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