Double-negative (CD4- CD8-) T (DNT) cells have been postulated to be potentially autoreactive. However, the role of DNT cells in rheumatoid arthritis (RA) has received limited attention. We investigated the distribution of DNT subsets in peripheral blood (PB) and synovial fluid (SF) from patients with active RA to determine whether these cells have relevance to RA. Two-colour flow cytometric analysis was performed to detect DNT cells in PB from 35 RA patients, 26 healthy controls and in SF aspirated from 19 inflamed rheumatoid joints. The subsets of DNT cells, i.e those expressing T cell receptor alphabeta (alphabeta DNT) or gammadelta (gammadelta DNT) were simultaneously examined. Our results showed that DNT cells constituted a very minor subset of PB lymphocytes. When expressed as a percentage of total lymphocytes, alphabeta DNT levels in normal individuals ranged from 0.27 to 2.08% (average 0.76%), while those of gammadelta DNT ranged from 1.02 to 11.42% (average 3.23%). Compared with normal individuals, RA patients had a similar distribution of alphabeta DNT cells in both PB and SF. However, RA patients had significantly lower levels of gammadelta DNT cells in PB than control subjects (1.38 +/- 1.08% vs 3.23 +/- 2.12%, p<0.05), while the levels of gammadelta DNT cells in SF of RA patients were higher than those in PB from RA patients and normal controls. The difference between PB and SF in RA was statistically significant (3.90 +/- 1.88% vs 1.38 +/- 1.08%, p<0.05). A higher level of gammadelta DNT in SF than their paired PB was consistently noted from nine available paired samples. Our findings suggest that gammadelta NT cells, but not alphabeta DNT cells, are probably relevant to RA. The lower percentage of circulating gammadelta DNT cells might have resulted from migration from the circulation into the synovium, suggesting a role for gammadelta DNT cells in the pathogenesis of rheumatoid synovitis.