Objective To investigate the expression of Toll-like receptor 4 (TLR4), NF-κB p65 and the negative regulating factor Toll-interacting protein (Tollip) in colonic mucosa of rats with 2，4，6-trinitrobenzene sulfonic acid (TNBS)-induced experimental colitis. MethodsForty healthy male SD rats were randomly divided into normal control group and model group. The expression of TLR4, NF-κBp65 and Tollip was detected and compared between the two groups by immunohistochemistry(IHC) and RT-PCR. ResultsRT-PCR revealed a significant higher expression of TLR4 and NF-κB p65 mRNA in the model group compared with the normal control group (TLR4: 0.376±0.029 vs 0.215±0.049, t= 2.731, P=0.013； NF-κB p65: 0.746±0.049 vs 0.206±0.063, t=6.055, P=0.000). The expression of TLR4 and NF-κB p65 was positively correlated with the general injury score (TLR4: r=0.754, P=0.000; NF-κB p65: r=0.548, P=0.012) and the histological injury score (TLR4: r=0.866, P=0.000; NF-κB p65: r=0.919, P=0.000). Expression of Tollip mRNA was not significantly different between the two groups as detected by RT-PCR (0.288±0.050 vs 0.140±0.046, t=1.993, P=0.061). IHC revealed that Tollip protein expression was mainly assembled in the colonic mucosal lamina propria, and the expression in the model group was higher than that in the normal control group. Tollip expression was negatively correlated with the general injury score (r=-0.497, P=0.026) and the histological injury score (r=-0.551，P=0.012). ConclusionThe unbalanced expression of TLR4 and the negative regulating factor Tollip participates in the pathogenesis of inflammatory bowel disease.