Iron deficiency is the most common comorbidity of inflammatory bowel disease (IBD), but the effect of iron supplementation on the repair processes of intestinal injury in weaned mice is unknown. This study aimed to evaluate the potential mechanism of dietary iron on intestinal injury and intestinal regeneration in the dextran sodium sulfate (DSS)-induced colitis of the weaned mouse model. The mice were fed either a control diet containing (45.00 mg/kg Fe) or iron supplemental (448.30 mg/kg Fe) diet for 14 days, followed by a 7-day oral administration of 2.5% DSS to all mice. The result showed that at day 0 of the recovery period (0 DRP), the impact of iron on the gut index and intestinal morphology was found to be more significant in weaned mice compared to adult mice. At 3 DRP, the iron diet alleviated inflammation-induced weight loss, shortening of colon length, thickening of the muscle layer, and disruption of gut morphology. At 0, 3, and 7 DRP, we found that an iron diet increased intestinal stem cell (ISC) viability and protected epithelial integrity. Furthermore, FeSO(4) significantly enhanced organoid viability and increased mRNA expression of differentiation, ISC, and retinol metabolism-related marker genes in the organoids compared with the control group. Overall, this study demonstrates that the iron diet accelerates intestinal regeneration after intestinal injury in weaned mice by activating the retinol metabolic pathway to regulate the proliferation and differentiation of ISCs.