Clinical and pathogenic significance and its therapeutic application of anti-calpastatin antibodies in rheumatoid arthritis

我们在类风湿关节炎（RA）患者中发现了新型的钙蛋白酶抑制剂（calpastatin）自身抗体。由于钙蛋白酶被认为是一种中性蛋白酶，可能参与关节破坏和炎症过程，其抑制蛋白calpastatin的自身抗体的产生可能与RA的发病机制有关。在本研究中，我们打算检查1）使用来自编码人钙蛋白酶抑制蛋白的全长cDNA的表达产物的自身抗体的结构域反应性，2）患者自身抗体对钙蛋白酶抑制蛋白的抑制活性，以及3）钙蛋白酶抑制剂对关节炎模型动物的作用。（结构域L、I、II、III、IV和V），并且反应模式倾向于与疾病相关。RA血清主要反应域I和II。81%的RA患者血清与至少一个结构域反应，而SLE、硬皮病和肌炎患者血清与至少一个结构域反应的比例分别为46%、32%和43%，含有抗calpastatin抗体的RA患者血清IgG以剂量依赖方式抑制calpastatin结构域融合蛋白的calpain抑制活性。5/11例（45%）RA血清IgG恢复了至少一个结构域的钙蛋白酶活性，而SLE和正常对照组IgG对钙蛋白酶抑制蛋白结构域的功能无抑制作用。Calpeptin治疗组大鼠的滑膜浸润和软骨破坏较未治疗组明显减轻，提示抗Calpastatin抗体可作为RA的一种新的标志物，并可能与RA的发病机制密切相关。此外，我们的研究结果导致了一种新的治疗策略。

We found novel autoantibodies to calpastatin, natural inhibitor for calcium-dependent cycteine proteinase (calpain), in patients with rheumatoid arthritis (RA). Since calpain is thought to be one of neutral proteinases that may be involved in joint destruction and inflammation process, the production of autoantibodies to its inhibitor protein, calpastatin, might be associated with pathogenic mechanisms of RA. In this study, we intended to examine 1) the domain reactivity of autoantibodies using the expression products from the full-length cDNA encoding for human calpastatin, 2) the inhibitory activity of calpastatin by patient autoantibodies, and 3) the effect of calpain inhibitors for arthritis model animals.Patient sera showed a variety of reactivity among the five domains of calpastatin (domains L, I, II, III, IV and V), and the reactive patterns tended to correlate to the diseases. RA sera predominantly reacted domains I and II. 81% of sera from RA patients reacted at least one domain, whereas 46%, 32% and 43% were reacted in sera from SLE, scleroderma and myositis, respectively.IgG from RA patient sera containing anti-calpastatin antibodies suppressed the calpain inhibitory activity of calpastatin domain fusion proteins in dose-dependent manner. 5 of 11 (45%) IgG from RA sera recovered more than 40% of calpain activity of at least one domain, whereas IgG from SLE and normal controls did not inhibited the function of calpastatin domains.Among the various calpain inhibitors, calpeptin suppressed significantly the development of type II collagen-induced rat arthritis. Calpeptin-treated rats showed less synovial infiltration and less cartilage destraction than untreated rats.These results suggest that anti-calpastatin antibodies may be useful as a novel marker of RA, and may be closely associated with pathogenic mechanisms of RA. Moreover, our results lead to a new strategy of treatment.