Inflammatory macrophages in colonic mucosa are the leading drivers of the pathology associated with inflammatory bowel disease (IBD). Here we examined whether gadolinium chloride (GdCl3), a macrophage selective inhibitor, would improve the course of 2,4,6-trinitro benzene sulfonic acid (TNBS) and dextran sodium sulfate (DSS)-induced colitis in mice and the potential mechanisms were investigated. By giving GdCl3 to colitis mice through intravenous or intrarectal route, we found that GdCl3 markedly ameliorated the colitis severity, including less weight loss, decreased disease activity index scores, and improved mucosal damage. To investigate the potential mechanisms, flow-cytometric analysis was performed to detect the proportion of mucosal macrophages in colon. The results showed that GdCl3 had no macrophage depletion effect in colonic mucosa, but significantly suppressed TNBS and DSS-induced TNFα, IL-1β and IL-6 secretions. Also, Western blotting analysis indicated that NF-κB p65 expression was significantly attenuated in the mucosa in colitis mice with GdCl3 treatment. Then, the anti-inflammatory activity of GdCl3 was confirmed in LPS-stimulated RAW 264.7 cells that GdCl3 might down-regulate the production of proinflammatory cytokines by macrophages through inhibition of the NF-κB signaling pathway. Therefore, intervention with mucosal inflammatory macrophages may be a promising therapeutic target in IBD.