Development of Immune Induction System by Using The Novel Virus Vectors of Cytokines

本研究的目的是利用重组仙台病毒反向遗传系统这一新型重组蛋白生产系统制备细胞因子，并对其进行体内外生物学活性检测。然后，通过检测活性对细胞因子网络进行基础研究，并在此基础上通过蛋白质或病毒载体的药物化，用于开发有效的细胞因子佐剂免疫导向方法。1）能够制备分别含有犬IFNγ、GM-CSF、IL-8和IL-6的cDNA的重组仙台病毒，能够成功地进行浓缩和纯化，并能通过体外实验检测出具有生物活性的产物。该犬实际上在本财政年度接受了药物治疗，并检索了体内有效性。2）从接种编码SENDAI病毒的犬GM-CSF的含胚鸡蛋中精制的GM-CSF指导犬源骨髓细胞或GM-CSF依赖性细胞储备液（TF-1细胞）的增殖，以达到浓度依赖性。当对6只动物的健康狗进行给药时，可以接受外周白细胞、嗜中性粒细胞和单核细胞的显著增加。3）当对利什曼原虫感染小鼠的皮肤病变部分进行给药时，能够控制皮肤病变的加重（扩大），所述狗IL-8是从感染14天开始经5天进行类似发现并精制的。从上述结果可以看出，通过重组仙台病毒的狗细胞因子产生系统是一种强有力的工具[在体内也具有足够的生物活性。

The purpose of this research is making cytokine using the recombinant SENDAI virus reverse genetics system which is a new recombinant protein production system, and examining the biological activity in in vitro and in vivo. Then, it is doing fundamental research of a cytokine network by checking activity, and using for development of the effective cytokine adjuvant immunity guiding method by medication of protein or a virus vector on the base.1) The recombination SENDAI virus which has cDNA of dog IFNγ, GM-CSF, IL-8, and IL6, respectively was able to be made, it was able to succeed in condensing and purification, and the products which show biological activity was able to be checked by in vitro. The dog was actually medicated with this fiscal year, and it searched about the validity in in vivo.2) GM-CSF refined from embryonated chicken eggs which inoculated the dog GM-CSF encoding SENDAI virus guided multiplication of a dog origin marrow cell or a GM-CSF dependability cell stock (TF-1 cell) to concentration dependability. When the healthy dog of six animals was medicated, the remarkable increase in peripheral leukocytes, neutrophiles and monocytes was accepted.3) When the skin pathological change part of a Leishmania protozoan infection mouse was medicated with dog IL-8 which were made to discover similarly and were refined over five days from the 14 days of infection, aggravation (expansion) of a skin pathological change was able to be controlled. Dog cytokine producing system by the recombination SENDAI virus have shown to be a powerful tool [having sufficient living thing activity also in in vivo from the above results.