基础医学与临床 ›› 2017, Vol. 37 ›› Issue (2): 145-149.

• 研究论文 •    下一篇

伴侣素包含T复合蛋白1ε亚基(CCT5)是与人γδT细胞相关的自身抗原

陈慧,滕达,游红琴,高运安,张建民,何维   

  1. 中国医学科学院基础医学研究所
  • 收稿日期:2016-03-30 修回日期:2016-07-01 出版日期:2017-02-05 发布日期:2017-01-16
  • 通讯作者: 何维 E-mail:mianyi333@163.com
  • 基金资助:
    国家科技部资助的国家重点基础研究发展规划(973)项目;国家自然基金委面上项目;国家自然科学基金-青年项目;北京协和医学院-协和青年基金

Chaperonin containing T-complex protein 1 subunit epsilon (CCT5) serves as γδ T cell-related autoantigen

  • Received:2016-03-30 Revised:2016-07-01 Online:2017-02-05 Published:2017-01-16
  • Contact: HE WEI E-mail:mianyi333@163.com

摘要: 目的 探讨CCT5与γδ T细胞及自身免疫病的关系。方法 PCR扩增人CCT5的基因,表达并纯化CCT5重组蛋白;用3条CCT5蛋白的肽段免疫小鼠,制备并筛选抗CCT5的单克隆抗体;固相化CCT5重组蛋白,体外扩增人外周血单个核细胞中的γδ T细胞;用制备的单抗通过ELISA检测正常人、类风湿性关节炎(RA)及系统性红斑狼疮(SLE)病人血浆中CCT5蛋白的含量;用流式细胞技术分析不同亚型γδ T细胞的比例,并与血浆CCT5含量做相关性分析。结果 PCR扩增获得CCT5基因,目的片段长度为1 750 bp;经SDS-PAGE分析可见重组CCT5蛋白分子量约为65ku,并获得了纯化。通过筛选,获得两株抗CCT5单克隆抗体对。重组CCT5蛋白能在体外扩增外周血γδ T细胞;而血浆CCT5浓度与Vγ9和Vδ2 T细胞的比例呈显著正相关。在RA及SLE患者血浆中,可溶性CCT5的浓度显著低于正常对照(p<0.05)。结论 CCT5是与人Vγ9δ2 γδ T细胞相关的自身抗原,有助于我们深入了解Vγ9δ2 γδ T细胞在自身免疫病中的作用。

关键词: CCT5, 自身免疫病, γδ T细胞, 自身抗原

Abstract: Objective To examine the relationship between CCT5, γδ T cell and autoimmune diseases. Methods Recombinant CCT5 protein was cloned, expressed and purified in E.coli. Three peptides of CCT5 protein was used to prepare for anti-CCT5 monoclonal antibodies. Purified CCT5 protein was used to expand γδ T cells from peripheral blood mononuclear cells (PBMC). Plasma levels of CCT5 in healthy donors, patients with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) were detected by ELISA assays. The correlation analysis between plasma CCT5 concentration and the percentage of different subtypes of γδ T cells measured by flow cytometry was made. Results The CCT5 gene was amplified by PCR and the length of the target fragment was 1 750 bp. The expressed 65ku CCT5 protein was purified and validated by SDS-PAGE. Two paired monoclonal anti-CCT5 antibodies were screened to detect CCT5 protein in plasma. Immobilized recombinant CCT5 protein was able to induce specific significant amplification of peripheral γδ T cells. Correlation analysis of 10 healthy donors indicated significant correlation between the plasma CCT5 concentration and the proportion of Vγ9 and Vδ2 γδ T cells. The plasma CCT5 concentration was significantly decreased in autoimmune diseases patients, including RA and SLE. Conclusion These data suggest that CCT5 could be a novel Vγ9δ2 γδ T cell-related factor in autoimmune diseases, which deepen our understanding of Vγ9δ2 γδ T cell function in autoimmune diseases.

Key words: CCT5, autoimmune diseases, γδ T cells, autoantigen

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