Inflammatory bowel disease (IBD) has been gradually considered as a public health challenge worldwide. This study determined the protective effect of Lachnum polysaccharide (LEP) on dextran sulfate sodium (DSS)-induced experimental colitis in mice and explored the underlying mechanism. Results showed that dietary LEP reduced DSS-induced disease activity index (DAI), colon shortening and colonic tissue damage. LEP treatment restored intestinal barrier integrity by regulating the expression of tight junction proteins and mucus layer protecting proteins. Moreover, pro-inflammatory cytokine production was inhibited by LEP through regulating PPARγ/NF-κB and IL-6/STAT3 pathways and inhibiting inflammatory cell infiltration. In addition, LEP also inhibited (NOD)-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation, endoplasmic reticulum (ER) stress and oxidative/nitrosative stress induced by DSS. These results provided a scientific basis for LEP as a potential natural agent for protecting mice from DSS-induced IBD.