Objective：To investigate the effects and mechanisms of 1，25（OH）2D3 on ameliorating dextran sulfate sodium（DSS）-induced colitis in mice. Methods：With DSS-induced colitis mice model established，inflammatory phenotype and goblet cells were detected and analyzed among model groups.The expression levels of tight junction proteins（TJs）among colonic tissues from different model groups were detected by RT-qPCR，Western blot and immunofluorescence staining. Using Caco-2 monolayer cell model，the protection of 1，25（OH）2D3 for TJs was detected in vitro. The mechanism of 1，25（OH）2D3 regulating expression of Claudin 4 was investigated via Chromatin immuno-precipitation（ChIP）and luciferase reporter assay. Results：1，25（OH）2D3 could enhance the intestinal epithelial barrier by promoting the expression of TJs，maintaining the structure of goblet cells and decreasing the intestinal permeability，thus relieving inflammation in DSS-induced acute colitis in mice. Conclusion：1，25（OH）2D3 plays a protective role in acute colitis mouse model via preserving intestinal permeability，promoting the expression levels of TJs and regulating the expression of Claudin4 through Vitamin Dreceptor.