基于伪无菌实验与肠菌移植技术研究气滞证慢传输型便秘模型大鼠肠道菌群对肠道动力的影响  

Study on the Effect of Intestinal Flora on Intestinal Motility in Rats with Slow Transit Constipation of Qi Stagnation Pattern Based on Pseudo-Sterile Experiment and Fecal Microbiota Transplantation

作　　者：刘启鸿[1] 柯晓[1] 骆云丰[1] 胡露楠 任彦[1] 方文怡[1] 赵培琳[1] 严锦贤[1,2] LIU Qihong;KE Xiao;LUO Yunfeng;HU Lunan;REN Yan;FANG Wenyi;ZHAO Peilin;YAN Jinxian(Second People's Hospital Affiliated to Fujian University of Traditional Chinese Medicine,Fuzhou,350003;Fujian Provincial Key Laboratory of Chinese Medicine Preparattiionin Medical Institutions)

机构地区：[1]福建中医药大学附属第二人民医院,福建省福州市350003 [2]福建省医疗机构中药制剂重点实验室

出　　处：《中医杂志》2024年第9期943-948,共6页Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金(82274282);福建省自然科学基金(2021J01878);福建省科技计划(2023L3013);福建省科技平台建设项目(2021Y2010)。

摘　　要：目的运用伪无菌实验与肠菌移植技术,探讨气滞证慢传输型便秘(STC)大鼠的肠道菌群对肠道动力的影响以及二者的关系。方法24只Wistar大鼠随机分为正常组6只、气滞证STC组6只、伪无菌组12只。气滞证STC组采用洛哌丁胺混悬液3 mg/kg灌胃联合夹尾刺激建立气滞证STC大鼠模型,造模成功后分别采集气滞证STC组和正常组大鼠的新鲜粪便制备100 mg/ml粪菌混悬液。伪无菌组采用抗生素鸡尾酒法(给予含杆菌肽、硫酸链霉素、硫酸新霉素各20 mg/ml的混合抗生素混悬液灌胃)建立伪无菌大鼠模型,造模成功后将大鼠随机分为正常粪菌液组、气滞证STC粪菌液组各6只,再分别给予制备的相应大鼠粪菌混悬液10 ml/kg灌胃;气滞证STC组大鼠给予等体积无菌水灌胃,均每日1次、连续7天。比较气滞证STC组、正常粪菌液组、气滞证STC粪菌液组小肠推进率;ELISA法检测血清5-羟色胺(5-HT)水平;免疫组化法检测结肠组织5-羟色胺3受体(5-HT3R)、5-羟色胺4受体(5-HT4R)阳性表达水平;Western blot法检测结肠组织色氨酸羟化酶1(TPH1)、色氨酸羟化酶2(TPH2)、5-羟色胺转运体(SERT)、单胺氧化酶A(MAO-A)蛋白表达水平。结果与正常粪菌液组比较,气滞证STC组、气滞证STC粪菌液组大鼠小肠推进率、血清5-HT水平,结肠组织5-HT3R、5-HT4R阳性表达以及TPH1、TPH2、SERT、MAO-A蛋白表达均降低(P<0.05);气滞证STC组和气滞证STC粪菌液组各项指标比较差异均无统计学意义(P>0.05)。结论气滞证STC大鼠的肠道菌群紊乱可导致肠道传输功能减慢,其机制可能是通过影响5-HT的合成、转运、代谢等途径而导致肠道动力障碍。Objective To clarify the relationship between intestinal flora and intestinal motility in rats with slow transit constipation(STC)and qi stagnation syndrome by conducting a pseudo-sterile experiment and fecal microbiota transplantation(FMT)technology.Methods Twenty-four Wistar rats were randomly divided into normal group(n=6),STC with qi stagnation pattern group(n=6)and pseudo-sterile group(n=12).In the STC group with qi stagnation pattern,3 mg/kg of loperamide suspension by intragastric administration combined with tail clamping stimulation were performed to establish the rat model of STC with qi stagnation pattern.After successful modeling,fresh feces from the rats in the STC with qi stagnation pattern group and the normal group were collected to prepare 100 mg/ml of fecal bacterial suspension.In the pseudo-sterile group,the antibiotic cocktail method was used(a mixed antibiotic suspension containing bacitracin,streptomycin sulfate,and neomycin sulfate at 20 mg/ml each was administered intragastrically)to establish pseudo-sterile rats model.After successful modeling,the rats were randomly divided into normal fecal bacterial liquid group and STC with qi stagnation pattern fecal bacterial liquid group,with six rats in each group,and then were given 10 ml/kg of the prepared corresponding rat fecal bacterial suspension by gavage.Rats in STC with qi stagnation pattern group were given an equal volume of sterile water by gavage.All groups were administered once a day for 7 consecutive days.The small intestinal propulsion rate of the STC with qi stagnation pattern group,the normal fecal bacterial liquid group,and STC with qi stagnation pattern fecal bacterial liquid group were compared.ELISA method was used to detect serum 5-hydroxytryptamine(5-HT)levels.Immunohistochemistry was used to detect the positive expression levels of 5-hydroxytryptamine 3 receptor(5-HT3R)and 5-hydroxytryptamine 4 receptor(5-HT4R)in colon tissue.Western blot method was used to detect the protein expression levels of tryptophan hydroxylase 1(T

关 键 词：慢传输型便秘 气滞证 伪无菌实验 肠菌移植 肠道菌群 肠道动力 

分 类 号：R256.35[医药卫生—中医内科学]