Extracellular matrix (stroma) regulation of mucosal T-cell function is incompletely understood. In this study, we uncovered a role for intestinal stromal products in the innate regulation of effector T cells. Stroma-conditioned media (S-CM) derived from the normal human intestinal stroma (transforming growth factor-Β (TGF-Β) hi/interleukin (IL)-6 lo/IL-1Β lo) significantly downregulated T-cell proliferation and interferon-γ (IFN-γ) production compared with S-CM derived from the inflamed Crohn's mucosa (TGF-Β hi/IL-6 hi/IL-1Β hi). Antibody neutralization studies showed that TGF-Β in normal S-CM inhibited T-cell proliferation and IFN-γ production, whereas IL-6 plus IL-1Β in Crohn's S-CM promoted T-cell proliferation, and IL-1Β alone promoted IFN-γ and IL-17 release. Importantly, normal S-CM inhibited T-bet expression, whereas Crohn's S-CM activated signal transducer and activator of transcription 3, suggesting that discordant T-cell responses are regulated at the transcription factor and signaling levels. These findings implicate stromal TGF-Β in the downregulation of T-cell 2responses in the normal intestinal mucosa, and stromal IL-6 and IL-1Β in the promotion of Th1 and Th17 responses in the inflamed Crohn's mucosa, suggesting an innate regulatory function for the intestinal extracellular matrix. © 2011 Society for Mucosal Immunology.