Liaskos Christos, Spyrou Vassiliki, Athanasiou Labrini V, Orfanidou Timoklia, Mavropoulos Athanasios, Rigopoulou Eirini I, Amiridis Georgios S, Shoenfeld Yehuda, Billinis Charalambos, Bogdanos Dimitrios P
Cellular Immunotherapy and Molecular Immunodiagnostics, Biomedical Section, Institute for Research and Technology Thessaly (IReTeTh), ITE, Larissa 41222, Greece; Department of Rheumatology, Faculty of Medicine, School of Health Sciences, University of Thessaly, Biopolis, 45000 Larissa, Greece.
Department of Animal Production, Technological Educational Institute of Thessaly, Larissa 41110, Greece.
Clin Res Hepatol Gastroenterol. 2015 Jun;39(3):384-90. doi: 10.1016/j.clinre.2014.12.001. Epub 2015 Jan 6.
Pancreatic autoantibodies (PABs) specifically recognizing GP2 and/or CUZD1 are present in more than 35% of patients with Crohn's disease (CrD). We have recently provided evidence of the presence of GP2-specific PABs in ruminants with paratuberculosis (ptb), a Mycobacterium avium paratuberculosis (MAP)-induced disease resembling CrD.
To assess whether anti-CUZD1 antibodies are also present in ruminants with ptb.
A total of 110 samples (73 cattle/37 sheep) were studied including 40 with ptb (24 cattle/16 sheep; 20 anti-GP2 antibody pos) and 70 without ptb (49 cattle/21 sheep; 10 anti-GP2 antibody pos). The samples were pre-characterized for anti-MAP and anti-GP2 antibodies by ELISA. Evidence of MAP was confirmed by PCR. Anti-CUZD1 antibody testing was performed by indirect immunofluorescence (IIF) based on transfected HEK293 cells expressing CUZD1. Anti-sheep or anti-cattle specific antisera were used as revealing antibodies.
None of the ruminant sera had anti-CUZD1 antibodies by IIF testing at dilutions varying from 1/10 to 1/160. Methodological flaws were prevented by a series of tests. Control sera from anti-CUZD1 positive CrD samples have shown anti-CUZD1 antibody reactivity at various concentrations. Antibody reactivity to GP2-expressing HEK293 cells has confirmed the reactivity to GP2 in ruminant sera found positive for anti-GP2 antibodies by ELISA.
The present study has found no evidence of anti-CUZD1 PABs in MAP-induced ptb. Our findings indicate that the induction of CUZD1-specific PABs is unrelated to MAP infection and that the mechanisms responsible for the loss of tolerance to GP2 and CUZD1 are probably quite distinct.
超过35%的克罗恩病（CrD）患者体内存在特异性识别GP2和/或CUZD1的胰腺自身抗体（PABs）。我们最近发现，患有副结核病（ptb）的反刍动物体内存在GP2特异性PABs，副结核病是一种由鸟分枝杆菌副结核亚种（MAP）引起的疾病，与CrD相似。
评估患有ptb的反刍动物体内是否也存在抗CUZD1抗体。
共研究了110份样本（73头牛/37只羊），其中40份患有ptb（24头牛/16只羊；20份抗GP2抗体阳性），70份未患ptb（49头牛/21只羊；10份抗GP2抗体阳性）。通过酶联免疫吸附测定（ELISA）对样本进行抗MAP和抗GP2抗体的预鉴定。通过聚合酶链反应（PCR）确认MAP的存在。基于转染表达CUZD1的人胚肾293（HEK293）细胞，采用间接免疫荧光法（IIF）进行抗CUZD1抗体检测。使用抗羊或抗牛特异性抗血清作为显色抗体。
通过IIF检测，在1/10至1/160的不同稀释度下，反刍动物血清中均未检测到抗CUZD1抗体。通过一系列测试避免了方法学缺陷。来自抗CUZD1阳性CrD样本的对照血清在不同浓度下均显示出抗CUZD1抗体反应性。对表达GP2的HEK293细胞的抗体反应性证实了ELISA检测中抗GP2抗体呈阳性的反刍动物血清中对GP2的反应性。
本研究未发现MAP诱导的ptb中存在抗CUZD1 PABs的证据。我们的研究结果表明，CUZD1特异性PABs的诱导与MAP感染无关，对GP2和CUZD1耐受性丧失的机制可能截然不同。
