Quantification and characterization of myosin peptide-specific CD4 + T cells in autoimmune myocarditis

• Published in: Journal of immunological methods 2005-09, Vol.304 (1), p.117-125
• Main Authors: Maier, Reinhard, Miller, Simone, Kurrer, Michael, Krebs, Philippe, de Giuli, Rita, Kremer, Marcel, Scandella, Elke, Ludewig, Burkhard
• Format: Article
• Language: English
• Subjects: Animals; Autoantigens - chemistry; Autoantigens - immunology; Autoimmune Diseases - immunology; Autoimmune Diseases - metabolism; Autoimmune myocarditis; Biological and medical sciences; CD4-Positive T-Lymphocytes - immunology; CD4-Positive T-Lymphocytes - metabolism; Cytokines; ELISPOT; Enzyme-Linked Immunosorbent Assay; Epitopes, T-Lymphocyte - chemistry; Epitopes, T-Lymphocyte - immunology; Female; Flow Cytometry - methods; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Immunophenotyping - methods; Kinetics; Mice; Mice, Inbred BALB C; Molecular immunology; Myocarditis - immunology; Myosin Heavy Chains - chemistry; Myosin Heavy Chains - immunology; Peptide Fragments - chemistry; Peptide Fragments - immunology; T helper cells; Techniques; Th1 Cells - immunology
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2005.06.013

Characterization of autoantigen-specific CD4 + T cells at the single cell level is crucial for understanding the immunopathological mechanisms underlying autoimmune diseases. Cardiac myosin heavy chain (myhca) is the major autoantigen associated with autoimmune myocarditis both in humans and in experimental autoimmune myocarditis (EAM) in mice. In the current study, we evaluated two methods for the enumeration and phenotypic characterization of myhca-specific CD4 + T cells during the course of EAM. Both enzyme-linked immunospot (ELISPOT) and cytokine flow cytometry (CFC) assays were suitable for the detection and characterization of myhca-specific Th cells during acute myocardial inflammation and the late healing phase of the disease. Cytokine production of myhca-specific Th cells was restricted to interferon-γ (IFNγ). Only trace amounts of the Th2 cytokines IL-4 and IL-5 could be detected. Concomitant surface marker analysis in the CFC assay revealed the prototypical effector phenotype of myhca-specific Th1 cells during the acute phase of the disease. Taken together, the combination of both methods appears to be most appropriate for a comprehensive ex vivo single cell analysis of Th cells in heart-specific autoimmune disorders.