Toll样受体阻断剂对内毒素血症小鼠肠黏膜损伤的影响

Effects of Toll-like receptor blockers on intestinal mucosal injury in mice with endotoxemia

摘要:

目的 探究Toll样受体（TLR）阻断剂对小鼠肠黏膜上皮细胞间紧密连接蛋白ZO-1的影响及对核转录因子-κB （NF-κB）和肿瘤坏死因子-α（TNF-α）的影响。方法 将32只BALB/C小鼠分为对照组、模型组、TLR4处理组、TLR2处理组（n=8），腹腔注射LPS建立内毒素血症小鼠模型，TLR4处理组、TLR2处理组在腹腔注射LPS同时分别给予TLR4抗体和TLR2抗体（10 μg/只）腹腔注射，对照组以生理盐水替代。取各组小鼠远端小肠组织，采用RT-PCR及免疫组化法检测ZO-1、NF-κBp65及TNF-α的mRNA和蛋白定位表达。结果 模型组ZO-1 mRNA及蛋白表达明显低于对照组（P < 0.05），NF-κBp65、TNF-α mRNA及蛋白表达明显高于对照组（P < 0.05）；TLR4处理组及TLR2处理组ZO-1 mRNA及蛋白表达明显高于模型组（P < 0.05），NF-κBp65、TNF-α mRNA及蛋白表达明显低于模型组（P < 0.05）；TLR4处理组与TLR2处理组ZO-1、NF-κBp65、TNF-α mRNA及蛋白表达比较差异无统计学意义（P > 0.05）。结论 抗TLR2、TLR4单克隆抗体可减少核转录因子的激活，抑制炎症因子大量分泌，保护紧密连接蛋白，有望对治疗肠源性感染疾病提供新的思路。

Abstract:

Objective To investigate the effects of Toll-like receptor blockers TLR2-Ab and TLR4-Ab on the tight junction protein ZO-1 in intestinal epithelial cells in mice, as well as their effects on nuclear factor-kappa B (NF-κB) and tumor necrosis factor-α (TNF-α).Methods A total of 32 BALB/C mice were divided into control group, model group, TLR4 treatment group, and TLR2 treatment group, with 8 mice in each group. A mouse model of endotoxemia was established by intraperitoneal injection of lipopolysaccharide. The mice in the TLR4 treatment group and the TLR2 treatment group were given intraperitoneal injection of TLR4 antibody and TLR2 antibody (10 μg each mouse), respectively, and those in the control group were given normal saline. The distal small intestinal tissue was collected, and RT-PCR and immunohistochemistry were used to measure the mRNA and protein expression of ZO-1, NF-κBp65, and TNF-α.Results Compared with the control group, the model group had significantly lower mRNA and protein expression of ZO-1 and significantly higher mRNA expression of NF-κBp65 and TNF-α (P < 0.05). Compared with the model group, the TLR4 treatment group and the TLR2 treatment group had significantly higher mRNA and protein expression of ZO-1 and significantly lower mRNA and protein expression of NF-κBp65 and TNF-α (P < 0.05). There were no significant differences in the mRNA and protein expression of ZO-1, NF-κBp65, and TNF-α between the TLR4 treatment group and the TLR2 treatment group (P > 0.05).Conclusions Anti-TLR2 and anti-TLR4 monoclonal antibodies can reduce the activation of nuclear transcription factors, inhibit the secretion of inflammatory factors, and protect tight junction protein, which is expected to provide new ideas for the treatment of enterogenous infectious diseases.