Abstract Dendritic cells play a critical role in the initiation and maintenance of inflammatory responses. Galectin-3, a protein found in DCs, is a carbohydrate-binding protein implicated in several cellular processes. In mouse models of asthma and atopic dermatitis, galectin-3-deficient (gal3-/-) mice had significantly fewer infiltrating eosinophils and displayed lower Th2 but higher Th1 responses compared to wild-type mice, suggesting that galectin-3 plays a key role in allergic inflammation. Given the ability of DCs to direct the T lymphocyte response, we hypothesized that galectin-3 may affect immune responses by altering DC functions and tested the hypothesis by comparing wild-type and gal3-/- DCs. OT-II CD4+ cells cultured with OVA-pulsed gal3-/- DCs generated higher Th1 responses relative to gal3+/+ DCs, and the differences were diminished following IL-12 neutralization. Moreover, gal3-/- DCs expressed more IL-12p35 mRNA than gal3+/+ DCs, indicating that gal3 may modulate IL-12 at the transcriptional level or through upstream signaling pathways. T cells co-cultured with gal3-/- DCs also secreted more IL-17 than cells cultured with gal3+/+ DCs, suggesting that gal3 may also negatively regulate Th17 responses. Furthermore, we observed higher IL-6 secretion and IL-23p19 expression in gal3-/- DCs, which may contribute to the enhanced Th17 polarization induced by these cells. We conclude that gal3 may regulate DC cytokine expression, thereby modulating T helper responses.