The role of TLR4/MyD88/NF-κB in the protective effect of ulinastatin on intestinal mucosal barrier in mice with sepsis

Abstract

Objective To investigate the effect of TLＲ4 /MyD88 /NF-κB signaling pathway in the protective effect of ulinastatin on intestinal mucosal barrier in mice with sepsis. Methods The mouse model of sepsis was established by classical cecal ligation and perforation. Thirty-two SPF C57BL/6 mice were randomly divided into four groups with 8 in each: control group (Con group), ulinastatin group (Uti group), Uti +TLR4 agonist LPS group (Uti+LPS group) and LPS group. Mice in the Con group received the same volume of saline injected into the tail vein 2 hours after modeling; Mice in the Uti group received 10000 U/kg ulinastatin injected into tail vein 2 hours after modeling; Mice in the Uti+LPS group received LPS (5mg/kg) injected into tail vein at 0h after modeling, other procedures were the same as in the Uti group; Mice in the LPS group received LPS (5mg/kg) injected into tail vein 0h after modeling. The weight loss ratio of mice was calculated at 48h after surgery. All animals were sacrificed at 48h after surgery to assess the injury of jejunal mucosa, the levels of TNF-α, IL-6 and IL-1 β in tail vein, and the expression of TLR4, MyD88 and NF-κ B mRNA in small intestinal mucosa tissues using ELASA and RT-PCR. Results The weight change of mice in Uti at 48h after operation was significantly reduced than that of the Con (p=0. 008 vs. Con); while the weight change of mice in Uti+LPS group and LPS group was significantly increased than that of the Uti (P=0. 020, P=0. 036 vs. Uti). The scores of intestinal mucosal injury at 48 h of the Uti were significantly lower than that of the Con (p＜0.001 vs. Con)；while the scores of intestinal mucosal injuryat 48 h of the Uti+LPS were significantly higher than that of the Uti ( P=0.044 vs. Uti). The expression of TNF-α, IL-6 and IL-1 β in the Uti decreased significantly at 48h after surgery than that in the Con（P=0.001，P=0.014，P=0.004 vs. Con), while the expression of TNF-α, IL-6 and IL-1β in Uti+LPS increased significantly at 48h after surgery than that in the Uti（P=0.026，P=0.040，P=0.039 vs. Uti）. The expression of TLR4, MyD88 and NF-κB mRNA in Uti decreased significantly than that in the Con(P=0.001，P=0.021，P=0.007 vs. Con); while the expression of TLR4, MyD88 and NF-κB mRNA in Uti+LPS was higher than that in Uti (P=0.023，P=0.040，P=0.045 vs. Uti). Conclusion These findings indicate that the intestinal mucosal barrier protective effect against sepsis of ulinastatin may be mediated through the TLR4/MyD88/NF-κB pathway.