NDLI: Clonal tracking of autoaggressive T cells in polymyositis by combining laser microdissection, single-cell PCR, and CDR3-spectratype analysis.

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Clonal tracking of autoaggressive T cells in polymyositis by combining laser microdissection, single-cell PCR, and CDR3-spectratype analysis.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Wiesener, Solveigh Hohlfeld, Reinhard Hofbauer, Monika Goebels, Norbert Roers, Axel Dornmair, Klaus Babbe, Holger Wekerle, Hartmut
Description	Author Affiliation: Hofbauer M ( Institute for Clinical Neuroimmunology, Ludwig Maximilians University, D-81377 Munich, Germany.);
Abstract	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 beta 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.
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	7
Volume Number	100
Language	English
Publisher	National Academy of Sciences
Publisher Date	2003-04-01
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Gene Rearrangement, T-Lymphocyte Genetics Polymyositis Immunology Receptor-CD3 Complex, Antigen, T-Cell T-Lymphocytes Amino Acid Sequence Antigens, CD Blood Biopsy DNA Primers Dissection Molecular Sequence Data Muscle, Skeletal Pathology Polymerase Chain Reaction Chemistry Research Support, Non-U.S. Gov't Multidisciplinary
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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