Intestinal barrier dysfunction plays a pivotal role in multiorgan dysfunction during heatstroke (HS). Neutrophils are involved in intestinal inflammation and thus dampen the mucosal integrity. Regulatory T cells (Tregs) have been shown to orchestrate neutrophils and thus sustain mucosal integrity in miscellaneous inflammation-related diseases. However， whether Tregs are involved in HS-induced intestinal barrier dysfunction remains unknown. Thus， we investigated whether Tregs could alleviate intestinal barrier dysfunction in mice. We found that HS could induce intestinal injury and mucosal barrier dysfunction 0， 24， and 72?h after heat stress. Flow cytometry revealed an increase of neutrophil infiltration and a decrease of Treg frequencies in the small intestinal epithelium 72?h after heat stress. Treg depletion starting 2?days before HS exacerbated intestinal damage and mucosal barrier dysfunction. Adoptive transfer of Tregs at 0?h improved intestinal injury and mucosal barrier dysfunction at 72?h. The manipulation of Tregs affected the neutrophil frequencies in the small intestinal epithelium 72?h after heat stress. Our study demonstrated that Tregs could improve HS-induced intestinal barrier dysfunction， probably via modulation of neutrophils in the intestine of mice during HS.