Department of Gastroenterology, Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, China.
MP Biochemicals, Auckland, New Zealand.
BMC Immunol. 2019 Jun 26;20(1):21. doi: 10.1186/s12865-019-0304-1.
Autoimmune diseases result from aberrant immune attacks by the body itself. It is mysterious how autoantigens, a large cohort of seemingly unconnected molecules expressed in different parts of the body, can induce similar autoimmune responses. We have previously found that dermatan sulfate (DS) can form complexes with molecules of apoptotic cells and stimulate autoreactive CD5+ B cells to produce autoantibodies. Hence, autoantigenic molecules share a unique biochemical property in their affinity to DS. This study sought to further test this uniform principle of autoantigenicity.
Proteomes were extracted from freshly collected mouse livers. They were loaded onto columns packed with DS-Sepharose resins. Proteins were eluted with step gradients of increasing salt strength. Proteins that bound to DS with weak, moderate, or strong affinity were eluted with 0.4, 0.6, and 1.0 M NaCl, respectively. After desalting, trypsin digestion, and gel electrophoresis, proteins were sequenced by mass spectrometry. To validate whether these proteins have been previously identified as autoantigens, an extensive literature search was conducted using the protein name or its alternative names as keywords. Of the 41 proteins identified from the strong DS-affinity fraction, 33 (80%) were verified autoantigens. Of the 46 proteins with moderate DS-affinity, 27 (59%) were verified autoantigens. Of the 125 proteins with weak DS-affinity, 44 (35%) were known autoantigens. Strikingly, these autoantigens fell into the classical autoantibody categories of autoimmune liver diseases: ANA (anti-nuclear autoantibodies), SMA (anti-smooth muscle autoantibodies), AMA (anti-mitochondrial autoantibodies), and LKM (liver-kidney microsomal autoantigens).
This study of DS-affinity enrichment of liver proteins establishes a comprehensive autoantigen-ome for autoimmune liver diseases, yielding 104 verified and 108 potential autoantigens. The liver autoantigen-ome sheds light on the molecular origins of autoimmune liver diseases and further supports the notion of a unifying biochemical principle of autoantigenicity.
自身免疫性疾病是由机体自身异常免疫攻击引起的。令人费解的是，在身体不同部位表达的大量看似不相关的分子——自身抗原，如何能诱导相似的自身免疫反应。我们之前发现，硫酸皮肤素 (DS) 可以与凋亡细胞的分子形成复合物，并刺激自身反应性 CD5+ B 细胞产生自身抗体。因此，自身抗原分子在与 DS 的亲和力方面具有独特的生化特性。本研究旨在进一步检验这种自身抗原性的统一原则。
从新鲜采集的小鼠肝脏中提取蛋白质组。将其加载到填充有 DS-琼脂糖树脂的柱子上。用增加盐浓度的分步梯度洗脱蛋白质。与 DS 结合亲和力弱、中或强的蛋白质分别用 0.4、0.6 和 1.0 M NaCl 洗脱。脱盐、胰蛋白酶消化和凝胶电泳后，通过质谱对蛋白质进行测序。为了验证这些蛋白质是否以前被鉴定为自身抗原，我们使用蛋白质名称或其替代名称作为关键字进行了广泛的文献检索。在强 DS 亲和力级分中鉴定出的 41 种蛋白质中，有 33 种（80%）被验证为自身抗原。在中等 DS 亲和力的 46 种蛋白质中，有 27 种（59%）被验证为自身抗原。在弱 DS 亲和力的 125 种蛋白质中，有 44 种（35%）为已知的自身抗原。引人注目的是，这些自身抗原属于自身免疫性肝病的经典自身抗体类别：ANA（抗核自身抗体）、SMA（抗平滑肌自身抗体）、AMA（抗线粒体自身抗体）和 LKM（肝-肾微粒体自身抗原）。
本研究通过 DS 亲和富集肝脏蛋白质，建立了一个全面的自身免疫性肝病自身抗原组，产生了 104 种已验证和 108 种潜在自身抗原。肝脏自身抗原组揭示了自身免疫性肝病的分子起源，并进一步支持了自身抗原性的统一生化原理的观点。
