Ⅱ型胶原短肽对胶原诱导性关节炎大鼠淋巴细胞体外增殖作用的研究

The proliferative effect of C Ⅱ 260-272 peptide on lymphocytes of collagen-induced arthritis rat

目的观察Ⅱ型胶原短肽（CⅡ260—272）对胶原诱导性关节炎（CIA）大鼠T淋巴细胞激活的影响。探讨其在类风湿关节炎（RA）发病中的作用。方法建立CIA大鼠模型，分别在建模1、2…8周后，体外分离、培养大鼠脾淋巴细胞，采用活细胞计数（CCK-8）法检测C1I260—272短肽对淋巴细胞增殖反应的影响，并分析CⅡ短肽特异性T淋巴细胞增殖反应与滑膜炎及血管翳评分、成模时间及抗CⅡ及其短肽抗体之间的相关性。用SPSS17．0软件进行数据分析。滑膜炎和血管翳评分结果比较用单因素方差分析，方差齐用LSD法，方差不齐用Games—Howell法；2组间增殖水平比较用Mann—WhitneyU检验；细胞增殖与其他指标的关系用Spearman相关分析；2组间抗体水平比较用t检验。结果免疫3周后，C11短肽特异性T细胞即出现明显的增殖反应[0．963（0．332-1．628）]，与0周组大鼠10．332（0．331N0．357）]比较，差异有统计学意义（P〈0．05）；分析发现，T细胞增殖反应与成模时间及抗bcⅡ抗体、抗Cit—bCⅡ抗体水平均呈正相关（r分别为：0．624，0．413，0．575，P均〈0．01）。免疫2周后模型组大鼠中抗CⅡ260—270抗体（0．827±0．155）与0周（0．043±0．009）比较，差异有统计学意义（P〈0．01），免疫4周后抗bcⅡ抗体（0．362±0．062）、抗cit—bcⅡ抗体（0．390±0．141）与0周相比，均有显著升高（P〈0．01）；同时，这3种抗体水平与血管翳评分也有显著相关性（P〈0．01或P〈0．05）。结论CⅡ短肽在建模早期即可刺激CIA大鼠脾淋巴细胞异常增殖，并在血清中检出针对CⅡ短肽的抗体反应。提示在利用CⅡ分子诱发CIA疾病发生过程中，该表位肽在驱动T、B淋巴细胞活化及引起自身免疫反应中发挥了重要作用。

Objective To examine the proliferative effect of synthetic CⅡ 260-272 peptide on T cells of collagen-induced arthritis （CIA） rat, and to explore the role of CⅡ 260-272 in RA. Methods The rat model of collagen-induced arthritis （CIA） was established. The T lymphocytes were isolated and incubated with CⅡ 260-272 peptide. Proliferation of T cells was determined by cell counting kit-8. The data were analyzed with SPSS 17.0. The Mann-Whitney U test was used for proliferation levels comparison between the two groups, and the relationship of cell proliferation with other indicators was analyzed with Spearman＇s correlation. Antibody levels between the two groups were compared using t test. Results Three weeks after the first immunization, T cell response to CⅡ 260-272 in the CIA groups [0.963 （0.332-1.628）] was more significant than in controls [0.332 （0.331-0.357）] （P〈0.05）, and the response was associated with disease course （r=0.624, P〈0.01 ）. Strong correlation between T cell proliferation and the antibodies, including anti- bCⅡ antibody and anti-Cit-bCⅡ antibody was also observed（P〈0.01 ）. Two weeks later, the levels of anti-CⅡ peptide antibody in each CIA group were significantly higher than those in the controls （P〈0.01）. The antibodies strongly correlated with pannus formation （P〈0.01, P〈0.05）. Conclusion The proliferative response of T cell to CⅡ 260-272 peptide can be found in the early stage of CIA rat, and the anti-CⅡ