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Development of a novel method for evaluating immunostimulants in fish based on gene expression.

开发一种基于基因表达评估鱼类免疫刺激剂的新方法。

基本信息

批准号：
10556046
负责人：
YANO Tomoki
金额：
$ 8.7万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

To analyze mode of action of immunostimulants such as b-1,3-glucans and sodium alginate, which have been reported to enhance biodefence of carp (Cyprinus carpio), genes of which expression was stimulated by the immunostimulants were identified by means of the suppression subtractive hybridization. As a result, about 50 genes including acute phase protein known in mammals, such as pentraxin and serum amyloid A, as well as several chemokines, chemokine-receptors, a novel C-type lectin, a novel complement factor B isotypes, glia-maturation factor, IL-1 beta, and pre-B cell enhancing factor were identified. Their complete primary structures were also determined from respective full-length cDNA clones, which were isolated from a carp leukocyte cDNA library. Interestingly, the b-1,3-glucan stimulated expression of a novel complement factor B isotype in carp kidney and spleen, suggesting a new role of factor B in bony fish. On the other hand, sodium alginate induced expression of chemokines and chemokine receptors, in agreement with the results that peritoneal injection of sodium alginate stimulated the migration of leukocytes from head kidney to peritoneal cavity. The present data suggests that each immunostimulant gave distinct pattern of gene expression in leukocytes and therefore, identification of the expressed genes gives an important insight into the mechanism of immunostimulants-mediated biodefense in bony fish. The approach in the present study would be useful to evaluate and to screen immunostimulants in bony fish.

为分析免疫增强剂B-1，3-葡聚糖和海藻酸钠对鲤鱼生物防御的作用机制，采用抑制性消减杂交技术对免疫增强剂刺激鲤鱼表达的基因进行了鉴定。结果，鉴定了约50个基因，包括哺乳动物中已知的急性期蛋白，如五聚蛋白和血清淀粉样蛋白A，以及几种趋化因子、趋化因子受体、新型C型凝集素、新型补体因子B同种型、神经胶质成熟因子、IL-1 β和前B细胞增强因子。它们的完整的一级结构也确定了各自的全长cDNA克隆，这是从鲤鱼白细胞cDNA文库中分离。有趣的是，B-1，3-葡聚糖刺激了鲤鱼肾脏和脾脏中一种新的补体因子B同种型的表达，表明B因子在硬骨鱼中的新作用。另一方面，海藻酸钠诱导趋化因子和趋化因子受体的表达，与腹腔注射海藻酸钠刺激白细胞从头肾向腹腔迁移的结果一致。目前的数据表明，每种免疫刺激剂在白细胞中的基因表达模式不同，因此，表达的基因的鉴定提供了一个重要的洞察机制的免疫刺激剂介导的生物防御硬骨鱼。本研究的方法将有助于硬骨鱼免疫刺激剂的评价和筛选。