溃结灵IV号对溃疡性结肠炎大鼠结肠粘膜Claudin-2、ZO-1、ICAM-1、TLR2基因表达的影响

Effect of Kuijieling IV on the Expression of Claudin-2, ZO-1, ICAM-1 and TLR2 Genes in Colonic Mucosa of Rats with Ulcerative Colitis

摘要: 目的：观察溃结灵IV号保留灌肠对三硝基苯磺酸(TNBS)诱导的溃疡性结肠炎(UC)大鼠结肠黏膜claudin-2、ZO-1、ICAM-1、TLR2基因表达的影响，探讨其治疗UC的可能机制。方法：实验于2015年6~8月在哈尔滨工业大学生命科学与工程学院实验室进行。SD大鼠60只，留取10只作为空白对照(BC)组，其余大鼠利用TNBS溶液灌肠法诱导制作UC大鼠模型，造模后随机分为模型对照(MC)组，溃结灵IV号低剂量(LD)组、中剂量(MD)组、高剂量(HD)组和阳性对照组美沙拉嗓(PC)组。HD、MD、LD组分別采用溃结灵IV号高(40 g/kg)、中(20 g/kg)、低(10 g/kg) 3个剂量保留灌肠给药，PC组使用美沙拉嗪(0.4 g/kg)进行灌肠给药，BC组及MC组给等体积的蒸馏水，检测大鼠结肠黏膜claudin-2、ZO-1、ICAM-1、TLR2基因表达，观察溃结灵IV号保留灌肠对TNBS诱导的UC大鼠结肠组织的影响。结果：1) 闭合蛋白-2 (claudin-2)，mRNA的表达：经PCR扩增、琼脂糖凝胶电泳后，在200 bp至250 bp位置可见电泳条带(Cldn2 244 bp)。结果表明溃结灵IV号高剂量、中剂量组及美沙拉嗪组claudin-2的mRNA表达明显增强，与空白组的表达接近，甚至略强于空白组的表达，模型组、溃结灵IV号高低剂量组claudin-2的mRNA略有的表达，表达不明显。2) 细胞粘附分子-1 (intercellular adhesion molecule, ICAM-1) mRNA的表达：经PCR扩增、琼脂糖凝胶电泳后，在200 bp至250 bp位置可见电泳条带(ICAM-1 211 bp)。结果表明正常组ICAM-1的mRNA表达不明，美沙拉嗪组ICAM-1的mRNA表达较正常组弱，溃结灵IV号高剂量、中剂量、低剂量及模型组ICAM-1的mRNA表达明显增强，明显高于空白组的表达。3) TLR2 (toll样受体2, Toll-like receptors 2) mRNA的表达：经PCR扩增、琼脂糖凝胶电泳后，在200 bp至250 bp位置可见电泳条带(TLR2 246 bp)。结果表明正常组TLR2的mRNA表达较弱，美沙拉嗪组、溃结灵IV号高剂量、中剂量组TLR2的mRNA表达较模型组和溃结灵IV号低剂量组明显减弱，模型组和溃结灵IV号低剂量组TLR2的mRNA表达明显增强，明显高于正常组的表达。

Abstract: Objective: To observe the gene expression of claudin-2, ZO-1, ICAM-1 and TLR2 of Kuijieling IV retention enema p-trinitrobenzenesulfonic acid (TNBS)-induced ulcerative colitis (UC) rats and explore the possible mechanism of its treatment of UC. Methods: The experiment was conducted in the laboratory of School of Life Science and Engineering, Harbin Institute of Technology from June to August 2015. Sixty SD rats were used, and 10 were used as the blank control (BC) group. The remaining rats were induced by TNBS solution enema to make UC rat models. After modeling, they were randomly divided into model control (MC) group and Kuijieling IV No. low-dose (LD) group, middle-dose (MD) group, high-dose (HD) group and positive control group Mesalah (PC) group. HD, MD, and LD groups were administered with Kuijieling IV high (40 g/kg), medium (20 g/kg), and low (10 g/kg) three-dose retention enema, and PC group used mesalazine (0.4 g/kg) for enema administration, BC group and MC group were given equal volume of distilled water to detect the gene expression of claudin-2, ZO-1, ICAM-1, and TLR2 in rat colonic mucosa. Observe Kuijieling IV retention enema for TNBS Effect of induced colonic tissue in UC rats. Results: 1) Expression of claudin-2 (claudin-2) mRNA: After PCR amplification and agarose gel electrophoresis, electrophoretic bands (Cldn2 244 bp) were visible from 200 bp to 250 bp. The results showed that the expression of claudin-2 mRNA of Kuijieling IV high-dose, middle-dose group and mesalazine group was significantly enhanced, which was close to the expression of the blank group, and even slightly stronger than that of the blank group. The model group, Kuijieling IV The mRNA of claudin-2 in the high and low dose group was slightly expressed, but the expression was not obvious. 2) mRNA expression of intercellular adhesion molecule-1 (intercellular adhesion molecule, ICAM-1): after PCR amplification and agarose gel electrophoresis, an electrophoretic band (ICAM-1 211 bp) can be seen at 200 bp to 250 bp. The results showed that the mRNA expression of ICAM-1 in the normal group was unknown, and the mRNA expression of ICAM-1 in the mesalazine group was weaker than that in the normal group. The high-, medium-, and low-dose Kuijieling IV was significantly expressed in the model group, obviously higher than the expression of the blank group. 3) TLR2 (toll-like receptors 2, Toll-like receptors 2) mRNA expression: After PCR amplification and agarose gel electrophoresis, the electrophoretic band (TLR2 246 bp) can be seen at 200 bp to 250 bp. The results showed that the mRNA expression of TLR2 in the normal group was weak. The mRNA expression of TLR2 in the mesalazine group, Kuijieling IV high-dose, and medium-dose group was significantly weaker than that in the model group and Kuijieling IV low-dose group. The expression of TLR2 mRNA in the low-dose group of Kuijieling IV was significantly enhanced, which was significantly higher than that in the normal group.