Giant cell arteritis: lesional microbial sequences

巨细胞动脉炎：病变微生物序列

• 批准号: 6480403
• 负责人: Lynn K Gordon
• 金额: $ 15.94万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6480403
• 关键词: cerebrovascular disorder diagnosis; clinical research; diagnosis design /evaluation; disease /disorder etiology; human tissue; humoral immunity; immunocytochemistry; microorganism genetics; nucleic acid sequence; temporal arteritis

Giant cell arteritis (UCA) is a serious disease unique to the elderly that leads to loss of sight in up to 20% of affected individuals. The diagnosis of UCA is often nonspecific and requires a constellation of clinical findings or a positive arterial biopsy. While its etiology is unknown, the granulomatous inflammation and local immunologic features of GCA lesions suggest involvement of a cryptic microorganism in disease pathogenesis. The primary goal of this proposal is to identify gene segments of microbial origin physically localized to GCA lesions and targeted by a disease-specific immune response. The long-term objective and health relatedness of this work is to enhance the understanding of OCA pathogenesis and develop new diagnostic methods and/or therapeutic strategies. GCA lesion-associated DNA sequences will be isolated and identified. Experimentally, this proposal takes advantage of recent advances in the detection of cryptic microorganisms in chronic diseases. A subtractive cloning strategy (RDA) will be performed to isolate novel DNA sequences from areas of histopathologic involvement in GCA+ affected arterial specimens. The sequences will be cloned and prioritized based on homology to known microbial sequences. High priority (candidate) sequences will be further analyzed to obtain additional evidence for their association with GCA. The immunologic relevance of high-priority gene sequences will be tested and sequences will be further defined by their association with UCA. Open reading frames (ORFs) of candidate sequences will be identified and produced as fusion proteins. The fusion proteins will be used to detect evidence for a disease-associated cell mediated or humoral immune response. Lesional prevalence and location of the high-priority gene sequences or products will be evaluated. Amplification of candidate sequences from arterial biopsies will determine the prevalence and disease-relatedness of the sequences. Localization of candidate sequences by immunohistochemistry and/or in situ hybridization will further validate the disease-association of the high-priority candidates. Significantly preliminary work validates this approach. Initial studies identify several high-priority candidate microbial sequences from GCA lesions that appear to be correlated with GCA+ involved arteries and elicit a disease-associated humoral immune response. Detection of a disease-associated humoral immunity against a product of a candidate sequence could form the basis for development of a serological diagnostic test for GCA. Further analysis of these candidates and identification of other candidate sequences is required to achieve the goals of this proposal.

巨细胞炎（UCA）是老年人独有的严重疾病，导致多达20％的受影响的个体失去视力。 UCA的诊断通常是非特异性的，需要临床发现或动脉活检阳性。尽管其病因尚不清楚，但GCA病变的肉芽肿性炎症和局部免疫学特征表明，隐性微生物参与疾病发病机理。该提案的主要目的是鉴定微生物起源的基因段物理定位于GCA病变，并以疾病特异性免疫反应为目标。这项工作的长期目标和健康相关性是增强对OCA发病机理的理解，并制定新的诊断方法和/或治疗策略。 GCA病变相关的DNA序列将被分离和鉴定。在实验上，该提案利用了慢性疾病中隐性微生物检测的最新进展。将进行减法克隆策略（RDA），以将新型DNA序列与GCA+受影响的动脉标本的组织病理学区域分离出来。这些序列将基于与已知微生物序列的同源性克隆并确定优先级。将进一步分析高优先级（候选）序列，以获得其与GCA相关的其他证据。 将测试高优先级基因序列的免疫学相关性，并将通过与UCA的关联进一步定义序列。候选序列的开放式阅读帧（ORF）将被鉴定为融合蛋白。融合蛋白将用于检测与疾病相关的细胞介导或体液免疫反应的证据。 将评估高优先性基因序列或产物的病变患病率和位置。来自动脉活检的候选序列的扩增将决定序列的患病率和疾病相关性。通过免疫组织化学和/或原位杂交将候选序列定位将进一步验证高优先候选者的疾病关联。 显着的初步工作验证了这种方法。初步研究发现，GCA病变中似乎与GCA+相关的几个高优先级候选微生物序列涉及动脉，并引起与疾病相关的体液免疫反应。针对候选序列的产物检测与疾病相关的体液免疫可以为开发GCA的血清学诊断测试构成基础。需要进一步分析这些候选人，并确定其他候选序列以实现该提案的目标。