Giant cell arteritis: lesional microbial sequences

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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.