ANALOG PEPTIDES IN COLLAGEN-INDUCED ARTHRITIS

胶原诱导关节炎（CIA）是一种通过免疫ⅱ型胶原（CII）易感菌株引起的自身免疫性关节炎的实验模型。我们最近对表达与类风湿关节炎（RA）易感性相关的HLA-DR转基因（Tg）小鼠的研究表明，DR1 （DRB1*0101）和DR4 （DRB1*0401）可以结合并呈递源自人(h) CII的肽，并且携带这些转基因DR分子的小鼠对CIA易感。当与hCII同时免疫时，CII、A12、CII 256-276类似肽（F263->N, E266->D）可下调hCII和关节炎的免疫应答。与用野生型CII肽（CII 256-276）培养的细胞相比，用Al2培养的DR Tg小鼠的人CII致敏脾和淋巴结细胞产生的IL-4和IL-10的数量增加。这些观察结果表明，在这些DR Tg小鼠中，A12对CII和CIA免疫应答的抑制作用是通过细胞因子谱的转变介导的，从Thl到Th2。然而，导致这种转变的机制尚不清楚。与野生型CII肽相比，Al2含有两个氨基酸取代。取代有两种类型，a)残基263参与与DR分子的肽结合，b)残基266与TCR相互作用。在本应用中要测试的假设是，A12对CIA的作用是通过对主要Th2谱的特异性免疫反应的极化介导的，这种极化是由DR和肽之间的亲和力变化或DR/肽与TCR的相互作用引起的，导致T细胞信号传导的改变，从而导致细胞因子的产生。因此，我们计划：1)确定介导DR Tg小鼠对CIIl和CIA免疫应答调节的类似肽Al2的结构特征；2)确定在三螺旋CII分子内编码Al2取代是否增加了抑制CIA自身免疫反应的效果；3)确定给药A12对CIA的抑制是否依赖于il - 10或IL-4的分泌；4)通过分析参与T细胞活化的信号通路，确定Al2改变T细胞功能的机制。从这些研究中获得的信息将为设计新的治疗方法提供重要的见解，这些治疗方法可能对人类自身免疫性关节炎的治疗有益。

Collagen-induced arthritis (CIA) is an experimental model of autoimmune arthritis induced by immunization of susceptible strains of mice with type II collagen (CII). Our recent studies using HLA-DR transgenic (Tg) mice expressing HLA class II molecules associated with susceptibility to rheumatoid arthritis (RA) reveal that DR1 (DRB1*0101) and DR4 (DRB1*0401) can bind and present peptides derived from human (h) CII, and that mice bearing these transgenic DR molecules are susceptible to CIA. The immune response to hCII and arthritis can be down regulated by an analog peptide of CII, A12, CII 256-276 (F263->N, E266->D), when it is co-administered at the time of immunization with hCII. Human CII- sensitized spleen and lymph node cells from DR Tg mice produce increased amounts of IL-4 and IL-10 when cultured with Al2 in comparison to cells cultured with the wild-type CII peptide (CII 256-276). These observations suggest that the suppressive effect of A12 on immune response to CII and CIA in these DR Tg mice is mediated by a shift in the cytokine profile, from that of a Thl to a Th2. However, the mechanism(s) whereby the shift is brought about is not known. Al2 contains two amino acid substitutions as compared to the wild-type CII peptide. The substitutions are of two types, a) residue 263 that participates in peptide binding to the DR molecule, and b) residue 266 that interacts with the TCR. The hypothesis to be tested in this application is that the A12 effect on CIA is mediated through polarization of the specific immune response to a predominant Th2 profile, and that this polarization is caused by changes in the affinity between either DR and peptide or DR/peptide interaction with the TCR leading to altered T cell signaling and, consequently, the production of cytokines. We, therefore, plan to: 1) Identify the structural characteristics of the analog peptide Al2 that mediate its modulation of the immune response to CIIl and CIA in DR Tg mice; 2) Determine whether encoding the Al2 substitutions within the triple helical CII molecule increases the efficacy in inhibiting the autoimmune response in CIA; 3) Determine whether the suppression of CIA produced by A12 administration is dependent on IL-l0 or IL-4 secretion; and 4) Determine the mechanism by which Al2 alters T cell function by analyzing signaling pathways involved in T cell activation. Information gained from these studies will provide important insight for the design of novel therapeutic approaches that may prove beneficial in the treatment of autoimmune arthritis in humans.