RHEUMATOID FACTOR SPECIFICITY

类风湿因子特异性

• 批准号: 3160810
• 负责人: RALPH C WILLIAMS
• 金额: $ 15.13万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-08-01 至 1995-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3160810
• 关键词: RNA splicing; autoantigens; chimeric proteins; human subject; immunoglobulin G; molecular cloning; rheumatoid factor; site directed mutagenesis

DESCRIPTION (adapted from applicant's abstract): The investigators propose to provide more direct information on the specificity of human IgM rheumatoid factors (RFs) and particularly those with heteroclitic reactivity. Thus, the investigators will explore the immunochemical basis for RF anti-Gm(a) specificity in a GM(a) negative RA patient, as well as why GM(a) negative RA patients RF react with GM(g)(+) IgG. Since most human RFs probably react at a relatively small 4-8A0 site between the C-gamma 2 - C-gamma 3 interface, changes in either conformation or selected residues in this region will be studied with particular respect to the individual Gm specificities of RFs isolated from serum by monomeric IgG affinity columns or with high avidity human monoclonal RFs produced by EBV stimulation of CD5 restricted B-cells. Quantitative studies of Gm phenotypes of IgG produced during short-term five to seven day pokeweed cultures of normals and RA patients will be completed using newly developed mouse MAbs to Gm(a) and Gm(f) determinants. These studies will attempt to examine whether lymphocytes from Gm(a) negative RA patients and normal controls will produce small amounts of Gm(a)(+) IgG when cultured in vitro with pokeweed mitogen. In parallel Gm(a) oligonucleotide-specific probes will be used with DNA in Gm(a) negative subjects to determine through polymerase chain reaction amplification the possible occurrence of mutational Gm(a) coding nucleotide sequences occurring in cultured PBMC from normals and RA subjects. Since three histidines are clustered together in residues 433, 435, and 310 at the C-gamma 2 - C-gamma 3 cleft where RF binds, chemical alteration of these histidines using diethylpyrocarbonate, platinum chromophores or spin-labeling will be attempted to study the effects both on Gm-related antigens as well as RF binding. Moreover, myeloma proteins of known Gm phenotypes will be studied after chemical histidine, tyrosine or lysine modifications to determine relative contributions of these residues to conformations or single residues responsible for Gm(a), (b), (g), or (f) allotypic specificities.

描述(改编自申请人的摘要)：调查人员建议 提供有关人免疫球蛋白M特异性的更直接信息 类风湿因子(RF)，特别是具有异质性的因子 反应性。因此，研究人员将探索免疫化学基础 对于GM(A)阴性RA患者的RF抗GM(A)特异性，以及 GM(A)阴性RA患者RF与GM(G)(+)免疫球蛋白反应的原因因为大多数人 人的RFs可能在相对较小的4-8A0部位发生反应 C-伽马2-C-伽马3界面，构象或选择的变化 该区域的残留物将进行研究，特别是 用单体免疫球蛋白从血清中分离RFS的个体GM特异性 EBV制备的亲和柱或高亲和力人源单抗RFs 刺激CD5限制性B细胞。转基因生物的定量研究 短期5~7天商陆产生的免疫球蛋白表型 正常人和类风湿关节炎患者的培养将使用新开发的 小鼠单抗对GM(A)和GM(F)决定簇。这些研究将试图 检测GM(A)阴性的类风湿关节炎患者和正常人的淋巴细胞 对照组在体外培养时会产生少量GM(A)(+)IgG 含有商陆有丝分裂原。平行GM(A)寡核苷酸特异性探针 将与GM(A)阴性受试者的DNA一起用于确定通过 聚合酶链式反应扩增可能发生的 外周血单核细胞中出现的GM(A)编码突变核苷酸序列 来自正常人和类风湿关节炎受试者。因为三个组氨酸聚集在一起 在C-Gamma 2-C-Gamma 3裂隙处的残基433、435和310中一起 在RF结合的地方，这些组氨酸使用 二乙基焦碳酸酯、铂生色团或自旋标记 试图研究其对GM相关抗原和RF的影响 有约束力的。此外，还将研究已知GM表型的骨髓瘤蛋白。 经化学组氨酸、酪氨酸或赖氨酸修饰后测定 这些残基对构象或单一结构的相对贡献 负责GM(A)、(B)、(G)或(F)异型特异性的残基。