类风湿关节炎患者外周血及滑膜液Ⅹ型胶原反应性B细胞检测及其特异性分析

Detection and specificity analysis of B cell response to human collagen type in peripheral blood and synovial fluid of patients with rheumatoid arthritis

目的：探讨类风湿关节炎（ＲＡ）患者体内是否存在抗Ⅹ型胶原Ｂ细胞反应。方法：以ＥＬ４ＯＵｒＢＵｒ／ｒＩＬ２／ＰＭＡ作为刺激系统，通过酶联斑点技术检测抗Ⅹ型胶原免疫球蛋白分泌Ｂ细胞（ａｎｔｉＨＣⅩＩＳＣ）频率。结果：２８例ＲＡ患者滑膜中１１例出现ａｎｔｉＨＣⅩＩＳＣ，而对照组骨性关节炎（ＯＡ）患者滑膜液中则未检测到ａｎｔｉＨＣⅩＩＳＣ。ＲＡ患者外周血及滑膜液ＣＤ１９＋及ＣＤ２０＋细胞百分率于刺激前后无明显变化，但ＣＤ５＋Ｂ细胞百分率却明显增加，外周血刺激前后分别为１７．０±６．０，２７．０±９．０，滑膜液分别为８．０±３．２，１６．０±６．０。结论：（１）ＲＡ患者体内存在抗Ⅹ型胶原Ｂ细胞反应，（２）ＣＤ４＋ＥＬ４ＯＵｒＢＵｒ／ｒＩＬ２／ＰＭＡ是激活ＣＤ５＋自身抗体产生Ｂ细胞的有效方法。

Objective: To detect the frequencies of B cell reaction to human collagen type (HC) in synovial fluid and peripheral blood in patients with rheumatoid arthritis (RA) and osteoarthritis (OA). ZMethods: The enzymelinked immunospot (ELISPOT) assay was used for enumerating antiHC immunoglobulin secreting cells (antiHC ISC) after highly efficient polyclonal B cell stimulation with the thymoma cells line (clone OUr BUr) in conjunction with PMA and recombinant human IL2. Results:TBZA low level, but significantly of human collagen specific B cell reaction was found in SFMC of RA patients after stimulation with EL4 thymoma cells together with PMA and rIL2, however, this polyclonal activation was inefficiency in SFMC of OA patients. Similar ratios were found for cells that expressed CD19 or CD20 in both RA and OA patients after stimulation of PBMC and SFMC with EL4/rIL2/PMA, however, the number of CD5+ B cell was increased in both RA synovial fluid and peripheral blood. Specificity analysis demonstrated that B cell reaction with HC in SFMC of RA patients was strongly inhibited by antiHC antibody but not by isotype matched antiHCII antibody in both ELISA and ELISPOT assays. WTHZConclusion: (1)antibody against type collagen in RA patients have pathogenic potential, however, this collagen specific B cell response only can be found in a very sensitive assay and a very sufficient stimulation. (2) CD4+EL4 OUr BUr/rIL2/PMA is efficient for activation of CD5+ autoantibody producing cells.