粪菌移植调控PAR2-TRPV1通路对肠易激综合征模型大鼠内脏高敏感性、肥大细胞活化及肠道菌群的影响

The Effects of Fecal Microbiota Transplantation Regulating the PAR2-TRPV1 Pathway on Visceral Hypersensitivity, Mast Cell Activation,and Gut Microbiota in a Rat Model of Irritable Bowel Syndrome

目的:探究粪菌移植调控PAR2-TRPV1通路对肠易激综合征模型大鼠内脏高敏感性、肥大细胞活化及肠道菌群的影响。方法:将大鼠随机分为Control组、IBS组、FMT组和FMT+SLIGRL-NH2组。测定粪便含水量,结直肠扩张(CRD)及行为学观察评估内脏敏感性,甲苯胺蓝染色检测结肠肥大细胞(MC)活性,16S-rDNA肠道菌群测序,蛋白质印迹法检测结肠组织中PAR2、TRPV1、SP、CGRP蛋白表达。结果:IBS组大鼠AWR评分、MC数量及结肠组织中PAR2、TRPV1、SP、CGRP蛋白表达明显高于Control组,OTU数、Chao1和Shannon指数明显低于Control组(P<0.05);FMT组大鼠AWR评分、MC数量及结肠组织中PAR2、TRPV1、SP、CGRP蛋白表达明显低于IBS组,OTU数、Chao1和Shannon指数明显高于IBS组(P<0.05);和FMT组相比,FMT+SLIGRL-NH2组大鼠AWR评分、MC数量及结肠组织中PAR2、TRPV1、SP、CGRP蛋白表达明显升高,OTU数、Chao1和Shannon指数明显降低(P<0.05)。和Control组相比,IBS组厚壁菌门、拟杆菌门和乳酸菌属菌落相对丰度明显降低,螺旋体菌门、变形菌门和普雷沃菌属菌落相对丰度明显升高(P<0.05);和IBS组相比,FMT组厚壁菌门和拟杆菌门和乳酸菌属菌落相对丰度明显升高,螺旋体菌门和变形菌门和普雷沃菌属菌落相对丰度明显降低(P<0.05);和FMT组相比,FMT+SLIGRL-NH2组厚壁菌门、拟杆菌门和乳酸菌属菌落相对丰度明显降低,螺旋体菌门、变形菌门和普雷沃菌属菌落相对丰度明显升高(P<0.05)。结论:粪菌移植可抑制IBS大鼠内脏高敏性和肥大细胞活化,调节肠道微生物平衡,其作用机制可能和抑制PAR2-TRPV1信号通路激活有关。

Objective: To investigate the effects of fecal microbiota transplantation regulating the PAR2-TRPV1 pathway on visceral hypersensitivity, mast cell activation, and gut microbiota in a rat model of irritable bowel syndrome. Methods: Rats were randomly divided into Control group, IBS group, FMT group, and FMT+SLIGRL-NH2 group. Measurement of fecal water content, colorectal distension(CRD) and behavioral observation to evaluate visceral sensitivity, detection of colon mast cell(MC) activity using toluidine blue staining, 16S r DNA sequencing of gut microbiota, and protein blotting to detect the expression of PAR2, TRPV1, SP, and CGRP proteins in colon tissue. Results: The results showed that the AWR score, number of MCs, and expression of PAR2, TRPV1, SP, and CGRP proteins in colon tissue of rats in the IBS group were significantly higher than those in the Control group, while the number of OTUs, Chao1,and Shannon index were significantly lower than those in the Control group(P<0.05);The AWR score, number of MCs, and protein expression of PAR2, TRPV1, SP, and CGRP in colon tissue of FMT group rats were significantly lower than those of IBS group, while the number of OTUs, Chao1, and Shannon index were significantly higher than those of IBS group(P<0.05);Compared with the FMT group,the FMT+SLIGRL-NH2 group showed a significant increase in AWR score, number of MCs, and expression of PAR2, TRPV1, SP, and CGRP proteins in colon tissue, while the number of OTUs, Chao1, and Shannon index decreased significantly(P<0.05). Compared with the Control group, the relative abundance of Firmicutes, Bacteroidetes, and Lactobacillus in the IBS group decreased significantly, while the relative abundance of Spirochetes, Proteobacteria and Prevotella increased significantly(P<0.05);Compared with the IBS group, the relative abundance of Firmicutes, Bacteroidetes, and Lactobacillus in the FMT group significantly increased, while the relative abundance of Spirochetes, Proteobacteria, and Prevotella decreased significantly(P<0.05);Compared with the FMT group, the relative abundance of Firmicutes, Bacteroidetes, and Lactobacillus in the FMT+SLIGRL-NH2 group decreased significantly, while the relative abundance of Spirochetes, Proteobacteria, and Prevotella increased significantly(P<0.05). Conclusion: Fecal microbiota transplantation can inhibit visceral hypersensitivity and mast cell activation in IBS rats, regulate intestinal microbiota balance, and its mechanism of action may be related to the inhibition of PAR2-TRPV1 signaling pathway activation.