3种壳聚糖对团头鲂体外头肾吞噬细胞呼吸爆发功能的影响  

Effect of three chitosan on respiratory burst activity of head-kidney phagocytes in Wuchang bream(Megalobrama amblycephala)

作　　者：郗明君[1] 刘立春[1] 张涓[1] 汤孝成[1] 刘小玲[1] 林蠡[1] 

机构地区：[1]华中农业大学水产学院/淡水水产健康养殖湖北省协同创新中心/农业部淡水生物繁育重点实验室,武汉430070

出　　处：《华中农业大学学报》2014年第3期72-77,共6页Journal of Huazhong Agricultural University

基　　金：中国水产科学研究院淡水生态与健康养殖重点开放实验室开放课题(2010FEA03012);国家重点基础研究发展计划("973")项目(2009CB118706)

摘　　要：采用51%Percoll非连续密度梯度离心法分离团头鲂头肾吞噬细胞,在光镜和电镜下观察所分离细胞的显微、超微结构及吞噬作用;以二氢罗丹明(DHR123)为荧光探针,流式细胞仪检测细胞在佛波豆蔻酸乙酯(PMA)刺激下产生的呼吸爆发活动;光镜下观察壳聚糖对吞噬细胞吞噬功能的影响。结果表明:分离得到的细胞具有吞噬细胞(巨噬细胞与中性粒细胞)的形态学及功能特征;3种壳聚糖组(100μg/mL)均能提高吞噬细胞的呼吸爆发功能,在发荧光细胞比例及细胞发荧光强度上均极显著高于对照组。其中水溶性壳聚糖组的发荧光细胞比例显著高于其他试验组,而在细胞发荧光强度上,普通壳聚糖作用相对较高;同时壳寡糖能增强吞噬细胞的吞噬活性。The aim of this study was to investigate the effects of oligochitosan（OCS）,water soluble chitosan（WSC）and acid soluble chitosan（ASC）on the respiratory burst activity of head-kidney phagocytes in Wuchang bream（Megalobrama amblycephala）.The head-kidney phagocytes of Wuchang bream were separated by 51% Percoll gradient and the morphological and ultrastructural features were observed under light and electron microscope.Staphylococcus aureus killed by formalin（F-SA）were incubated with phagocytes and observed.PP and PI were counted with Wright Giemsa staining under oil-immersion microscopy.The respiratory burst activity of phagocytes was measured by flow cytometry using dihydrorhodasmine 123（DHR123）as fluorescence probe after stimulation with phorbol 12-myristate 13-acetate（PMA）.The results indicated that the separated cells showed the morphological and functional characteristics of phagocytes.OCS（100μg/mL）,WSC（100μg/mL）and ASC（100μg/mL）were all able to stimulate the respiratory burst activity of head-kidney phagocytes after incubation for 30min.The percentage of fluoresce cell and the fluorescence intensity of fluoresce cell（two parameters of respiratory burst activity）in the experimental group were significantly different（P0.01）from the control group. However,the percentage of fluoresce cell of the phagocytes stimulated by WSC was the highest.But the strongest fluorescence intensity was observed in the phagocytes stimulated by ASC.

关 键 词：团头鲂 吞噬细胞 呼吸爆发 壳寡糖 水溶性壳聚糖 普通壳聚糖 吞噬作用 

分 类 号：S942.5[农业科学—水产养殖]