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| ID:
49900.0,
MPI für Neurobiologie / Neuroimmunology |
| Clonal tracking of autoaggressive T cells in polymyositis by combining laser microdissection, single-cell PCR, and CDR3-spectratype analysis |
| Authors: | Hofbauer, Monika; Wiesener, S.; Babbe, H.; Roers, A.; Wekerle, Hartmut; Dornmair, Klaus; Goebels, Norbert | | Language: | English | | Date of Publication (YYYY-MM-DD): | 2003-04-01 | | Title of Journal: | Proceedings of the National Academy of Sciences U S A | | Journal Abbrev.: | PNAS | | Volume: | 100 | | Issue / Number: | 7 | | Start Page: | 4090 | | End Page: | 4095 | | Review Status: | not specified | | Audience: | Not Specified | | Abstract / Description: | Clonal expansions of CD8+ T cells have been identified in muscle and blood of polymyositis patients by PCR techniques, including T cell receptor (TCR) complementarity-determining region (CDR)3 length analysis (spectratyping). To examine a possible pathogenic role of these clonally expanded T cells, we combined CDR3 spectratyping with laser microdissection and single-cell PCR of individual myocytotoxic T cells that contact, invade, and destroy a skeletal muscle fiber. First, we screened cDNA from muscle biopsy specimens by CDR3 spectratyping for expanded TCR chain variable region (BV) sequences. To pinpoint the corresponding T cells in tissue, we stained cryostat sections with appropriate anti-TCR BV mAbs, isolated single BV+ T cells that directly contacted or invaded a muscle fiber by laser-assisted microdissection, and amplified their TCR BV chain sequences from rearranged genomic DNA. In this way, we could relate the oligoclonal peaks identified by CDR3-spectratype screening to morphologically characterized microdissected T cells. In one patient, a large fraction of the microdissected T cells carried a common TCR-BV amino acid CDR3 motif and conservative nucleotide exchanges in the CDR3 region, suggesting an antigen-driven response. In several cases, we tracked these T cell clones for several years in CD8+ (but not CD4+) blood lymphocytes and in two patients also in consecutive muscle biopsy specimens. During immunosuppressive therapy, oligoclonal CDR3-spectratype patterns tended to revert to more polyclonal Gaussian distribution-like patterns. Our findings demonstrate that CDR3 spectratyping and single-cell analysis can be combined to identify and track autoaggressive T cell clones in blood and target tissue. This approach should be applicable to other inflammatory and autoimmune disorders. | | External Publication Status: | published | | Document Type: | Article |
| Affiliations: | MPI für Neurobiologie/Neuroimmunology (Wekerle)/Clinical Neuroimmunology (Hohlfeld)
| | External Affiliations: | Institute for Clinical Neuroimmunology, Ludwig Maximilians University, D-81377 Munich, Germany; Department of Neuroimmunology, Max Planck Institute for Neurobiology, 82152 Martinsried, Germany; and Institute for Genetics and Department of Dermatology, University of Cologne, 50931 Cologne, Germany
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