摘要
【目的】探讨脆弱拟杆菌(Bacteroides fragilis)中六型蛋白分泌系统(T6SS)对肠道屏障的影响及作用机制。【方法】采用自杀载体构建B.fragilis T6SS缺陷株。建立葡聚糖硫酸钠(DSS)诱导的肠炎小鼠模型,并分别补充PBS,B.fragilis WT和B.fragilisΔT6SS,随后比较三组小鼠疾病特征、肠道屏障完整性的差异。利用荧光定量PCR和免疫组化检测小鼠紧密连接蛋白的表达。采用非靶向代谢组学比较各组小鼠肠道差异代谢物。【结果】T6SS缺失不影响B.fragilis的生物活性。与PBS对照组相比,B.fragilis WT明显改善小鼠的体重丢失、结肠长度等疾病指标,表现出对DSS诱导的肠炎的保护性,而B.fragilisΔT6SS随着T6SS的敲除丧失了对DSS诱导的肠炎的保护性。小鼠血清中异硫氰酸荧光素葡聚糖含量和肠道病理切片均表明B.fragilis T6SS能改善小鼠肠道屏障的完整性。荧光定量PCR和免疫组化均表明B.fragilis T6SS影响肠道紧密连接蛋白的表达。非靶向代谢组学分析表明,与B.fragilisΔT6SS组相比,B.fragilis WT组显著上调96个肠道差异代谢产物,其中多个代谢产物富集到胆碱能突触代谢和甘油磷脂代谢相关通路。【结论】拟杆菌T6SS改变肠道代谢组并提高肠道细胞紧密连接蛋白的表达,改善肠道屏障的通透性,参与到拟杆菌对肠道屏障的保护性。
【Objective】To investigate the effect and mechanism of Bacteroides fragilis type VI protein secretion system(T6SS)on the intestinal barrier.【Method】Suicide vector was used to construct B.fragilis T6SS mutant strain.The dextran sulfate sodium(DSS)saltinduced colitis mouse model was constructed,and supplemented mice with PBS,B.fragilis WT,and B.fragilisΔT6SS,respectively.The disease characters and intestinal barrier integrity were compared among the three groups.Quantitative PCR and immunohistochemistry were used to detect the expressions of tight junction proteins.Untargeted metabolomics was used to compare gut differential metabolites in each group of mice.【Result】The deletion of T6SS did not affect bio-viability of B.fragilis.B.fragilis WT showed the improvements in body weight loss and colon length compared to PBS controls,demonstrating the protection to DSS-induced colitis;while this protection was lost with the absence of T6SS.The fluorescein isothiocyanate dextran concentration in the mouse serum and histological examination in intestinal pathological sections indicated that B.fragilis T6SS improved the integrity of the intestinal barrier in mice.T6SS mutation affected intestinal tight junction protein expression,confirmed by quantitative PCR and immunohistochemistry.Untargeted metabolomics analysis revealed 96 up-regulated differentially expressed metabolites in the B.fragilis WT group,compared to the B.fragilisΔT6SS group,with multiple metabolites enriched in cholinergic synaptic metabolism and glycerophospholipid metabolism-related pathways.【Conclusion】B.fragilis T6SS alters intestinal metabolome and increases the expression of tight junction protein in intestinal cells,thus improving the permeability of intestinal barrier,being involved in the protection of B.fragilis to the intestinal barrier.
作者
雷棋怡
徐杨
李鹏飞
LEI Qi-yi;XU Yang;LI Peng-fei(Department of Hematology,The First Affiliated Hospital of Soochow University,Suzhou 215000;Institute of Blood and Marrow Transplantation,Soochow University,Suzhou 215000)
出处
《生物技术通报》
CAS
CSCD
北大核心
2024年第3期286-295,共10页
Biotechnology Bulletin
基金
国家自然科学基金项目(82100231)
江苏省双创博士项目(JSSCBS20211562)
姑苏卫生人才科研项目(S2022021)。
