Aims Leucine supplementation promotes intestinal health, but the mechanism is largely unknown. This study aimed to elucidate the mechanisms underlying the beneficial effects of leucine on intestinal homeostasis. Methods and Results Female ICR mice (6-week-old) were randomly assigned into three groups: (i) mice received a basal diet; (ii) mice received a dietary 0 center dot 5% crystalline l-leucine supplementation; and (iii) mice received a dietary 1 center dot 0% crystalline l-leucine supplementation. Our results showed that leucine supplementation stimulated the secretion of SIgA in mice ileum and expression of cytokines related to SIgA production. Moreover, leucine supplementation improved the expression of mTOR and p70S6K1 expression. Further study showed that leucine supplementation markedly decreased microbiota richness and induced a shift in the Firmicutes : Bacteroidetes ratio in favour of Firmicutes. Conclusions Therefore, our data suggested that leucine supplementation could enhance intestinal health through the regulation of mTOR pathway and promoting SIgA secretion in the mouse intestine, which might be associated with intestinal microbiota. Significance and Impact of the Study The present study found that dietary leucine supplementation of mice could improve intestinal health by enhancing intestinal SIgA secretion via a nonexclusive mechanism, which might include T cell-dependent pathway, T cell-independent pathway and gut microbiota.