Berger Sven, Bannantine John P, Griffin J Frank T
Disease Research Laboratory, Department of Microbiology and Immunology, University of Otago, PO Box 56, Dunedin NZ 1, New Zealand.
Microbes Infect. 2007 Jul;9(8):963-70. doi: 10.1016/j.micinf.2007.03.016. Epub 2007 Apr 7.
Mycobacterial infection has been linked to the generation of autoantibodies, including anti-neutrophil cytoplasmic autoantibodies (ANCA), in clinical studies and small animal models. In an attempt to identify antibodies that react with both self and mycobacterial antigens in naturally infected ruminants, we generated a phage display library comprising single chain antibody fragments (scFv) from sheep with Johne's disease (JD). The library was screened simultaneously against ovine small intestinal tissue and Mycobacterium avium subsp. paratuberculosis (MAP). A clone (termed AutoH1) reacted strongly with host tissue and MAP, recognizing a proteinase-susceptible 32 kDa determinant in ovine gut tissues and lymphatics, and in blood granulocytes but not mononuclear cells. In granulocytes, binding was to cytoplasmic granules and cell membranes; in MAP, AutoH1 bound bacterial cell wall determinants. We further identified a synthetic peptide sequence recognized by AutoH1, using this to generate a carrier:peptide fusion protein (paH1). Sera of normal and JD sheep were screened for AutoH1-like autoantibody activity; 7/11 JD animals showed autoreactivity that could be blocked by paH1, while 0/21 normal animals showed no such serological reactivity. It is possible that the severe pathology observed in ruminant JD may have an autoimmune component, possibly due to ANCA-type binding; this remains to be further investigated.
在临床研究和小型动物模型中，分枝杆菌感染已与自身抗体的产生相关联，包括抗中性粒细胞胞浆自身抗体（ANCA）。为了鉴定在自然感染反刍动物中与自身和分枝杆菌抗原均发生反应的抗体，我们构建了一个噬菌体展示文库，该文库包含患有副结核病（JD）的绵羊的单链抗体片段（scFv）。该文库同时针对绵羊小肠组织和鸟分枝杆菌副结核亚种（MAP）进行筛选。一个克隆（称为AutoH1）与宿主组织和MAP强烈反应，识别绵羊肠道组织、淋巴管以及血液粒细胞（而非单核细胞）中一种对蛋白酶敏感的32 kDa决定簇。在粒细胞中，结合发生在细胞质颗粒和细胞膜上；在MAP中，AutoH1结合细菌细胞壁决定簇。我们进一步鉴定了AutoH1识别的一个合成肽序列，并利用该序列生成了一种载体：肽融合蛋白（paH1）。对正常绵羊和患JD绵羊的血清进行AutoH1样自身抗体活性筛选；11只患JD动物中有7只表现出可被paH1阻断的自身反应性，而21只正常动物中没有一只表现出这种血清学反应性。反刍动物JD中观察到的严重病理可能有自身免疫成分，可能是由于ANCA型结合；这仍有待进一步研究。
