One of the earliest signs of mucosal immune activation in celiac disease (CD) is an increase in the intraepithelial lymphocyte (IEL) count in the small intestinal epithelium. Though most of those IELs express T cell receptor (TcR)-alphabeta chains, CD is characterized by an increase in TcR-gammadelta+ IELs and by the loss of CD3- IELs. There is currently little evidence that these changes in IEL subset distribution are of relevance in the pathogenesis of CD. We aimed to determine the pattern of cytokine production by IEL subsets isolated from duodenal biopsy specimens from control subjects and CD patients at different stages of the disease. We quantified the capacity of IEL subsets to produce IFN-gamma, TNF-alpha, IL-2, IL-4, and IL-10 by intracellular staining by flow cytometry. All IEL subsets studied displayed a type I cytokine profile in both CD and control subjects, with TcR-alphabeta+ IELs being the main IFN-y producers. Untreated CD exhibited a trend toward a superior accumulation of IFN-gamma per cell but a reduced proportion of INF-gamma+ cells in vitro in association with a significantly increased apoptotic rate of IELs. IL-4 was almost undetectable in all cases and IL-10 showed a tendency to increase in treated and "silent" celiac patients. IEL subsets have a similar Th1 profile in controls and CD patients, and the superior in vitro apoptosis of IELs from CD patients may reflect their superior in vivo activation. The induction of IL-10-dependent regulatory Tr1 responses may be of potential clinical significance in this disease and merits further investigation.