Predominant cellular immune response to the cartilage autoantigenic G1 aggrecan in ankylosing spondylitis and rheumatoid arthritis.
OBJECTIVE
Based on their HLA association, both ankylosing spondylitis (AS) and rheumatoid arthritis (RA) seem to be T-cell-driven diseases in which the autoantigens remain to be defined. One possible autoantigen is the G1 domain of aggrecan, the major cartilage proteoglycan. In BALB/c mice immunized with this protein, spondylitis and erosive polyarthritis have been reported. Immune reactivity to the G1 has been described in patients with RA and AS in an earlier study. Using novel and more sensitive techniques and relevant controls we sought to define the role of G1 as an autoantigen more precisely and to extend the specific analyses to the peptide level.


METHODS
Peripheral blood (PB) mononuclear cells (MNC) from 47 AS patients, 22 RA patients and 20 healthy normal controls were exposed in vitro for 6 h to the cartilage-derived autoantigens G1, human cartilage (HC) gp-39 and collagen II. Synovial fluid (SF) MNC from seven AS and four RA patients were similarly analysed. Furthermore, PB MNC of 15 AS and 10 RA patients were examined with overlapping 18-mer peptides covering the whole G1 protein to identify the immunodominant epitopes. T cells were stained by monoclonal antibodies directed against the surface markers CD4, CD69 and against the intracellular cytokines interferon-gamma (IFN gamma), tumour necrosis factor-alpha (TNF alpha), interleukin 4 (IL-4) and IL-10. The percentage of reactive T cells was quantified by flow cytometry.


RESULTS
After antigen-specific stimulation with the G1 protein, the CD4+ T cells of 30 AS patients (61.7%) and of 12 RA patients (54.5%) secreted significant amounts of IFN gamma and TNF alpha, while, in contrast, only 10% of the normal controls showed a response (P < 0.05). The synovial CD4+ T cells of five AS (71.5%) and of all four RA patients showed antigen-specific responses to the G1. In contrast, stimulation with HC gp-39 and collagen II showed no significant IFN gamma and TNF alpha secretion of MNC in all groups. Several G1-derived T-cell epitopes were identified as immunodominant in PB MNC of AS and RA patients and were partly overlapping.


CONCLUSIONS
These data show that a cellular immune response to G1 is present in most AS and RA patients. G1-immunodominant epitopes were identified. The relevance of this finding for the pathogenesis of AS and RA remains to be established.
Department of Rheumatology, Klinikum Benjamin Franklin, Free University, Berlin, Germany.
J
J
Zou
Y
Y
Zhang
A
A
Thiel
M
M
Rudwaleit
S-L
SL
Shi
A
A
Radbruch
R
R
Poole
J
J
Braun
J
J
Sieper
D054392:Adipokines; D000328:Adult; D053674:Aggrecans; D001172:Arthritis, Rheumatoid / Q000276:immunology*; D001324:Autoantigens / Q000494:pharmacology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D002358:Cartilage, Articular / Q000276:immunology*; D016022:Case-Control Studies; D002478:Cells, Cultured; D000071451:Chitinase-3-Like Protein 1; D024043:Collagen Type II / Q000494:pharmacology; D000939:Epitopes / Q000032:analysis; D016326:Extracellular Matrix Proteins; D005434:Flow Cytometry; D006023:Glycoproteins / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D016898:Interferon-alpha / Q000276:immunology; D037102:Lectins; D037181:Lectins, C-Type; D008213:Lymphocyte Activation; D008875:Middle Aged; D017434:Protein Structure, Tertiary; D011509:Proteoglycans / Q000737:chemistry / Q000494:pharmacology*; D013167:Spondylitis, Ankylosing / Q000276:immunology*; D013582:Synovial Fluid / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
D054392:Adipokines; D053674:Aggrecans; D001324:Autoantigens; C084533:CHI3L1 protein, human; D000071451:Chitinase-3-Like Protein 1; D024043:Collagen Type II; D000939:Epitopes; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006133:Growth Substances; D016898:Interferon-alpha; D037102:Lectins; D037181:Lectins, C-Type; D011509:Proteoglycans; D014409:Tumor Necrosis Factor-alpha