日本血吸虫SjIR3基因核酸疫苗的研究

发布时间：2018-02-06 06:26

本文关键词： 日本血吸虫 SjIR3 mGM-CSF 核酸疫苗免疫保护力　出处：《湖南农业大学》2007年硕士论文　论文类型：学位论文

【摘要】： 疫苗是防治传染病的有效手段，作为一种相对便宜且长效的防治措施，研制出血吸虫病的疫苗一直是科学家们追求的目标。血吸虫病疫苗的研究经历了从死疫苗、致弱活疫苗、亚单位疫苗到基因工程疫苗的探索过程。核酸疫苗是血吸虫疫苗研究的重要内容。现阶段研制的血吸虫核酸疫苗候选分子所诱导的保护力比较弱，保护不完全，因此有必要提高疫苗分子的免疫诱导力和寻找新的疫苗候选分子。 1．本研究是通过由日本血吸虫cDNA文库筛选出的新基因(SjIR3)与真核表达载体pCDNA3.0构建出重组质粒SjIR3／pC，并以生理盐水(100μl)、空白质粒pCDNA3.0(50μg)为对照免疫KM小鼠(每组18只)，免疫方案是0-2-4-6，每次免疫前及攻击感染前于尾静脉采血进行ELISA实验检测抗体滴度。末次免疫后2w，各组剖杀任意6只小鼠取脾细胞，分别用ConA或rSjIR3重组蛋白诱导，，MTT法检测淋巴细胞增殖。各组余下小鼠经腹部感染日本血吸虫尾蚴40±1条，45d后宰杀，观察实验组小鼠的减虫率和减卵率。其结果显示，SjIR3／pC可诱导小鼠产生较高抗体水平(0.78±0.05)，ConA或rSjIR3重组蛋白诱导淋巴细胞增殖结果分别为0.57±0.02、0.68±0.01，减虫率和减卵率分别是29.4％、36.6％。 2．首次构建了鼠粒细胞-巨噬细胞集落刺激因子(mGM-CSF)与血吸虫基因的共表达载体(SjIR3-GMCSF／pC)，并设计了生理盐水(100μl)、空白质粒pCDNA3.0、SjIR3／pC、mGM—CSF／pC和SjIR3-GMCSF／pC(50μg)五组小鼠实验，其免疫方案、攻击感染、检测方法同前。结果显示，SjIR3／pC组与SjIR3-GMCSF／pC诱导小鼠产生的抗体水平分别为0.78±0.04、0.91±0.03；ConA或rSjIR3重组蛋白诱导淋巴细胞的转化率明显增加，其A_(570)分别为0.56±0.07和0.68±0.05、0.56±0.07和0.77±0.04；减虫率和减卵率分别是39.81％、34.75％和33.96％、28.43％。结论：SjIR3 DNA疫苗能够诱导受攻击感染的小鼠产生体液免疫和细胞免疫而具有部分抗攻击感染的免疫保护作用。mGM-CSF可增强SjIR3DNA疫苗诱导小鼠产生的体液免疫和细胞免疫应答和抗攻击感染的免疫保护作用。SjIR3 DNA疫苗免疫诱导家畜等产生的免疫应答和抗攻击感染的保护效果值得进一步研究，SjIR3 DNA疫苗可能成为血吸虫“鸡尾酒疫苗”的组成成分。
[Abstract]:The vaccine is effective in the prevention and treatment of infectious diseases, as a relatively inexpensive and effective prevention measures, development of schistosomiasis vaccine scientists are pursuing. Research on schistosomiasis vaccine has experienced from dead vaccine, attenuated vaccine, subunit vaccine to explore the process of genetic engineering vaccine of nucleic acid vaccine is important. The content of vaccine against Schistosoma japonicum. Protection induced by current developed schistosome nucleic acid vaccine candidate molecules is relatively weak, the protection is not complete, so it is necessary to improve the vaccine induced molecular force and looking for new molecules for vaccine.
1. in this study by screening new genes of Schistosoma japonicum cDNA library from Japan (SjIR3) and eukaryotic expression vector pCDNA3.0 to construct recombinant plasmid SjIR3 / pC, and normal saline (100 L), blank plasmid pCDNA3.0 (50 g) were immunized KM mice (n = 18), immune scheme 0-2-4-6, before each vaccination and attack before infection in the tail vein blood antibody titer ELISA assay. 2W after the last immunization, 6 mice of each group were sacrificed randomly from spleen cells treated with ConA or rSjIR3 recombinant protein induced lymphocyte proliferation was detected by MTT. All remaining mice by abdominal infection of Schistosoma japonicum cercariae 40 + 1, 45d after slaughter, the experimental group of mice the worm and egg reduction rates. The results showed that SjIR3 / pC can produce higher levels of antibodies in mice induced by (0.78 + 0.05), ConA or rSjIR3 recombinant protein induced lymphocyte proliferation results were 0.57 + 0.02,0.6 8 + 0.01, the rate of worm reduction and the rate of egg reduction were 29.4%, 36.6%.
2. for the first time established murine granulocyte macrophage colony stimulating factor (mGM-CSF) gene co expression with schistosome vector (SjIR3-GMCSF / pC), and the design of normal saline (100 L), blank plasmid pCDNA3.0, SjIR3 / pC, mGM, CSF / pC and SjIR3-GMCSF / pC (50 g) five groups the mice, the immune attack scheme, infection, with the detection method. The results showed that the antibody level of SjIR3 / pC and SjIR3-GMCSF / pC mice induced by respectively 0.78 + 0.04,0.91 + 0.03; lymphocytes induced by ConA or rSjIR3 recombinant protein conversion rate increased significantly, the A_ (570) were 0.56 + 0.07 and 0.68 + 0.05,0.56 + 0.07 and 0.77 + 0.04; the worm and egg reduction rates were 39.81%, 34.75% and 33.96%, 28.43%.
Conclusion: SjIR3 DNA vaccine can be worthy of further research on the immune response induced by infection induced humoral and cellular immunity and immune effects of.MGM-CSF protection against challenge infection can enhance SjIR3DNA vaccine induced humoral and cellular immune response and anti infection immune protection against the immune effect of.SjIR3 induced by DNA vaccine in livestock. The attack and anti infection protection effect, composition of SjIR3 DNA vaccine of Schistosoma japonicum may become a "cocktail vaccine".

【学位授予单位】：湖南农业大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R392

【相似文献】