重组绵羊肺表面活性物质结合蛋白A的体外生物学活性检测  

Biological activity of recombinant ovine pulmonary surfactant-associated protein A in vitro

作　　者：李增强 赵宁[1] 张彦兵[1] 李珂儿 魏立翔 孙延鸣[1] LI Zengqiang;ZHAO Ning;ZHANG Yanbing;LI Keer;WEI Lixiang;SUN Yanming(College of Animal Science and Technology,Shihezi University,Shehezi 832003,China)

机构地区：[1]石河子大学动物科技学院,新疆石河子832003

出　　处：《畜牧与兽医》2021年第9期89-94,共6页Animal Husbandry & Veterinary Medicine

基　　金：兵团重大科技项目(2017AA006-04)。

摘　　要：旨在研究重组绵羊肺表面活性物质结合蛋白A(rSP-A)对中性粒细胞弹性蛋白酶(NE)活性及淋巴细胞体外增殖的影响,初步分析rSP-A的抑菌活性。使用绵羊肺炎支原体(Mo)感染体外培养的绵羊外周血中性粒细胞以刺激其释放NE,然后应用底物显色法对比分析不同浓度rSP-A对NE活性的影响;使用MTT比色法测定rSP-A对体外培养的绵羊外周血淋巴细胞增殖活性的影响;使用酸化法和平板菌落计数法测定rSP-A对Mo代谢与增殖的影响;应用平板菌落计数法对rSP-A的抑菌活性进行初步分析。NE活性检测结果表明,rSP-A可显著增强Mo刺激绵羊外周血中性粒细胞释放的NE活性(P<0.05),且呈剂量效应关系;淋巴细胞增殖试验显示,rSP-A能够显著抑制淋巴细胞的增殖(P<0.05);代谢试验证实rSP-A对体外培养的Mo的代谢具有明显的抑制作用(P<0.01);抑菌试验显示,rSP-A对致病性肺炎链球菌和巴氏杆菌的增殖具有明显的抑制作用(P<0.05)。研究结果说明,真核表达的rSP-A具有明显的生物学活性,这为进一步研究绵羊体内肺表面活性物质结合蛋白A(SP-A)的生物学特性奠定了基础。In order to determine the effect of the recombinant ovine pulmonary surfactant-associated protein A(rSP-A) on the activity of neutrophil elastase(NE) activity and the proliferation of lymphocyte, in vitro, and to make a preliminary analysis of the antimicrobial activity of rSP-A in vitro, Mycoplasma ovipneumoniae(Mo) was used, in this study, to stimulate the release of NE by ovine peripheral blood neutrophils in vitro, and then the effects of rSP-A at different concentrations on NE activity was analyzed by chromogenic reaction of the substrates. MTT colorimetric assay was used to determine the effect of rSP-A on the proliferation of lymphocytes isolated from ovine peripheral blood. Biological acidification and the standard plate count method were used to determine the effect of rSP-A on the metabolism and proliferation of Mo. The standard plate count method was used to preliminarily analyze the antibacterial activity of rSP-A. The results of the NE activity test showed that rSP-A could significantly enhance the activity of NE released by Mo-stimulated ovine peripheral blood neutrophils in vitro in a dose-dependent manner(P<0.05). The lymphocyte proliferation test showed that rSP-A could significantly inhibit the proliferation of lymphocytes(P<0.05). The metabolic experiments confirmed that rSP-A could inhibit the metabolism of Mo in vitro(P<0.01). The antibacterial test showed that rSP-A had a significant inhibitory effect on the proliferation of the pathogenic Streptococcus pneumoniae and Pasteurella(P<0.05). The present study indicated that the recombinant ovine SP-A expressed by the eukaryotic expression system had obvious biological activity, which laid a foundation for further research on the biological characteristics of ovine SP-A.

关 键 词：绵羊 重组绵羊肺表面活性物质结合蛋白A 中性粒细胞弹性蛋白酶 淋巴细胞 生物学活性 绵羊肺炎支原体 

分 类 号：S852.4[农业科学—基础兽医学]