IDENTIFICATION OF TREATMENT-RESISTANT LYME ARTHRITIS AUTOANTIGENS

耐药性莱姆关节炎自身抗原的鉴定

• 批准号: 7602033
• 负责人: Brigitte T. Huber
• 金额: $ 0.86万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-03 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7602033
• 关键词: Affect; Affinity; Antibiotic Therapy; Antibodies; Autoantibodies; Autoantigens; B-Lymphocytes; Borrelia burgdorferi; Cells; Computer Retrieval of Information on Scientific Projects Database; Cytokeratin; Digestion; Funding; Gel; Grant; In Vitro; Infection; Institution; Joints; Lyme Arthritis; MALDI-TOF Mass Spectrometry; Manuscripts; Peptides; Plasma Cells; Production; Proteins; Publishing; Research; Research Personnel; Resistance; Resources; Sampling; Sepharose; Slice; Source; Synovial Fluid; T-Lymphocyte; Tissues; Trypsin; United States National Institutes of Health; Western Blotting; Work

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Antibiotic treatment-resistant Lyme arthritis is characterized by B-T cell aggregates and plasma cell clusters in synovial tissue from affected joints, after apparent eradication of Borrelia burgdorferi infection. It is suspected that this cellular activity in the joint is a due to activation of self-reactive B cells with subsequent production of autoantibodies. This work focuses on identification of potential self antigenic proteins from synovial tissue. Microdissected plasma cells from affected joints were cultured in vitro and the antibodies were purified from the cell supernates. These potential autoantibodies were conjugated to agarose beads and used to affinity purify antigenic proteins from synovial fluid. Proteins were eluted from the beads and separated by SDS-PAGE. Protein bands were excised from gels and subjected to in-gel trypsin digestion. Peptides extracted from the in-gel digests were analyzed by MALDI-TOF mass spectrometry for protein identification. We have identified a cytokeratin that was present in one sample, but not in control blank gel slices. We have confirmed the finding by Western blot and continue to search for other proteins in additional samples. A manuscript describing our results has recently been published.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 抗生素治疗耐药的莱姆关节炎的特点是，在伯氏疏螺旋体感染明显根除后，受影响关节的滑膜组织中出现 B-T 细胞聚集体和浆细胞簇。人们怀疑关节中的这种细胞活动是由于自身反应性 B 细胞的激活以及随后产生的自身抗体所致。这项工作的重点是从滑膜组织中鉴定潜在的自身抗原蛋白。在体外培养来自受影响关节的显微解剖浆细胞，并从细胞上清液中纯化抗体。这些潜在的自身抗体与琼脂糖珠缀合，用于亲和纯化滑液中的抗原蛋白。将蛋白质从珠子上洗脱下来并通过 SDS-PAGE 进行分离。从凝胶上切下蛋白质条带并进行凝胶内胰蛋白酶消化。通过 MALDI-TOF 质谱分析从凝胶内消化物中提取的肽，以进行蛋白质鉴定。我们已经鉴定出一种细胞角蛋白存在于一个样品中，但不存在于对照空白凝胶切片中。我们已通过蛋白质印迹证实了这一发现，并继续在其他样本中寻找其他蛋白质。描述我们结果的手稿最近已发表。