摘要
目的 基于Toll样受体4(TLR4)-核苷酸结合寡聚化结构域样受体(NOD)信号通路探究溃结灵对溃疡性结肠炎(UC)大鼠肠黏膜修复和肠道自噬的影响.方法 将 60 只大鼠分为正常组、UC组(模型组)和溃结灵低、中、高剂量组,每组12 只,除正常组外其余4 组给予2,4,6-三硝基苯磺酸和乙醇溶液灌肠制备UC模型.正常组和UC组灌胃等量生理盐水,溃结灵低、中、高剂量组灌胃1.85、3.70、7.40 g/kg溃结灵,每日1 次,连续12 周.分析各组大鼠病变活动指数(DAI)评分、结肠长度、结肠黏膜损伤指数(CMDI)评分;检测血清 D-乳酸(D-LAC)、二胺氧化酶(DAO)、血清和结肠组织肿瘤坏死因子-α(TNF-α);检测尿乳果糖(L)、甘露醇(M),用于计算尿L/M;采用Western blot法检测结肠组织自噬相关蛋白1(Beclin1)、自噬相关基因 5(ATG5)、TLR4、TLR2、NOD样受体蛋白 3(NLPR3)蛋白表达.结果 与正常组比较,治疗1、2、6 和 12 周UC组DAI评分升高;与UC组比较,治疗 1、2、6 和 12 周溃结灵低、中、高剂量组DAI评分降低(P＜0.05).与正常组比较,UC 组结肠长度明显缩短,CMDI 评分、血清 D-LAC 和DAO、尿L/M、血清和结肠组织 TNF-α及结肠组织 TLR4、TLR2、NLPR3 蛋白表达升高或增加,结肠组织 Beclin1 和ATG5 表达减少;与UC组比较,溃结灵低、中、高剂量组结肠长度延长,结肠组织Beclin1、ATG5 表达增加,CMDI评分、血清D-LAC和DAO、尿L/M、血清和结肠组织 TNF-α 及结肠组织 TLR4、TLR2、NLPR3 蛋白表达降低或减少(P＜0.05).溃结灵低、中、高剂量组上述指标比较差异有统计学意义(P＜0.05),呈剂量效应关系.结论 溃结灵可通过调控TLR4-NOD信号通路改善UC大鼠肠道自噬,减轻结肠组织炎症反应,有助于肠黏膜修复,而高剂量组效果较优.
Abstract
Objective To investigate the effects of Kuijieling on intestinal mucosal repair and intestinal autophagy in ulcerative colitis(UC)rats based on Toll-like receptor 4(TLR4)-nucleotide-bound oligomeric domain-like receptor(NOD)signaling pathway.Methods A total of 60 rats were divided into normal group,UC group(model group),and low-dose,medium-dose and high-dose Kuijieling groups,with 12 rats in each group.Except the normal group,the other four groups were given 2,4,6-trinitrobenzene sulfonic acid and ethanol solution enema to prepare UC model.The normal group and UC group were given the same amount of normal saline,and the low-,medium-and high-dose Kuijieling groups were given the same amount of Kuijieling 1.85 g/kg,3.70 g/kg and 7.40 g/kg,respectively,once a day for 12 consecutive weeks.Disease activity index(DAI)score,colon length,and colon mucosal damage index(CMDI)score in each group were analyzed.Ser-um D-lactic acid(D-LAC),diamine oxidase(DAO),serum and colon tissue tumor necrosis factor-α(TNF-α)were detected using detection kit.Lactulose(L)and mannitol(M)were detected,for calculating urine L/M.Western blot assay was used to detect the expressions of autophagy-related protein Beclin1,autophagy-related gene 5(ATG5),TLR4,TLR2 and NOD-like receptor protein 3(NLPR3)proteins in colon tissue.Results Compared with normal group,DAI scores in the UC group was higher at 1,2,6 and 12 weeks after treatment,and compared with UC group,DAI scores in low-,medium-and high-dose Kuijieling groups was significantly decreased at 1,2,6 and 12 weeks after treatment(P＜0.05).Compared with normal group,colon length in UC group was significantly shortened,CMDI score,serum D-LAC and DAO,urinary L/M values,TNF-α in serum and colon tissue,and TLR4,TLR2,NLPR3 protein expressions in colon tissue were significantly increased,and Beclin1 and ATG5 in colon tissues were significantly decreased(P＜0.05).Compared with UC group,colonic length was significantly prolonged in low-,medium-and high-dose Kuijieling groups,Beclin1 and ATG5 protein expression was signifi-cantly increased.CMDI score,serum D-LAC and DAO,urinary L/M values,TNF-α in serum and colon tissue,and TLR4,TLR2,NLPR3 protein expressions were significantly decreased(P＜0.05).There were statistically significant differences in the above indexes among low-,medium-and high-dose Kuijieling groups in a dose-effect manner(P＜0.05).Conclusion Kuijieling can improve intestinal autophagy in UC rats by regulating TLR4-NOD signaling pathway,alleviate colon tissue in-flammation,and facilitate intestinal mucosal repair,and the effect of high-dose group was better.
基金项目
南通市基础研究和民生科技计划指导性项目(2021)(JCZ21130)
维普
万方数据