芬氏另枝菌促进炎症性肠病发展及其机制

Alistipes finegoldii promotes the development of inflammatory bowel disease: effect and mechanism

摘要:

目的 探究芬氏另枝菌(Alistipes finegoldii)对炎症性肠病(inflammatory bowel disease, IBD)的影响，并进一步阐明其潜在的相关致病机制。方法 6周龄的雄性C57BL/6J小鼠在饮用链霉素3 d后，被随机分为3组：对照组(Control组)、PBS组和给菌组(AF组)。通过灌胃给予芬氏另枝菌(每只1×10⁹ CFU/200 μL)或PBS，持续2周。之后，小鼠饮用含有2.5%葡聚糖硫酸钠(dextran sulfate sodium, DSS)的水溶液1周以诱导结肠炎模型。评估小鼠的体重下降分数、粪便性状、血便情况、结肠长度，对结肠组织进行HE染色并进行组织病理评分；于实验开始及结束时收集小鼠粪便，进行16S rRNA基因扩增子测序；用实时定量聚合酶链式反应(qPCR)检测结肠组织相关肠屏障蛋白和炎症因子的mRNA表达。结果 AF组小鼠的体重下降分数、疾病活动度指数评分、结肠缩短程度及组织病理评分均高于PBS组。肠屏障的紧密连接蛋白Occludin、Claudin 5的mRNA表达相较于PBS组下降，而炎症因子IL-17A的mRNA表达则升高。AF组相较于PBS组的肠道菌群α多样性降低；β多样性分析显示两组间肠道菌群多样性存在显著性差异。线性判别分析效应大小(linear discriminant analysis effect size, LEfSe)分析发现，给菌组与PBS组之间存在显著差异的细菌类群: 在纲水平上，芽孢杆菌纲 (Bacilli)；在目水平上，丹毒丝菌目(Erysipelotrichales)；在科水平上，丹毒丝菌科(Erysipelotrichaceae)、海生线状菌科(Marinifilaceae)；在属水平上，气杆菌属(Odoribacter)、杜博斯氏菌属(Dubosiella)；在种水平上，纽约杜博斯氏菌(Dubosiella newyorkensis)，在给菌组中显著高于PBS组。结论 芬氏另枝菌能够促进炎症因子的分泌，损害肠黏膜通透性，改变肠道菌群的结构和多样性，从而加剧肠炎的发展。

关键词:炎症性肠病;肠道菌群;芬氏另枝菌;肠道屏障

Abstract:

Objective To study the effect of Alistipes finegoldii (AF) on inflammatory bowel disease (IBD) and the underlying mechanism.Methods Six-week-old male C57BL/6J mice were administrated with streptomycin for three days and then randomly assigned into the control, phosphate buffered saline (PBS), and AF groups. Mice were administrated with AF suspension (1×10⁹ CFU, 200 μL per mouse) or PBS by gavage for two weeks, followed by drinking of the water containing 2.5% dextran sulfate sodium (DSS) for one week for the modeling of colitis. The weight loss fraction percentage, fecal characteristics, blood fecesstools, and colon length were determined. The colon tissue was stained with hematoxylin-eosin for the scoring of histopathological changes, and feces samples were collected at the beginning and end of the experiment for sequencing of 16S rRNA gene amplicons at the beginning and end of the experiment. The mRNA levels of colon tissue-associated intestinal barrier proteins and inflammatory mediators were determined by qPCR.Results The mice in the AF group had severer disease conditions than those in the PBS group regarding the weight loss percentage, disease activity index, colon shortening, and histopathological score. Compared with the PBS group, the AF group showed down-regulated mRNA levels of occludin and claudin 5 and up-regulated mRNA level of interleukin (IL)-17A. The AF group had lower alpha diversity of intestinal flora than the PBS group, and the beta diversity showed significant differences between AF and PBS groups. The linear discriminant analysis effect size (LEfSe) results revealed that the significantly differential bacteria between AF and PBS groups were Bacilli, Erysipelotrichales, Erysipelotichaceae, Odoribacter, Marinifilaceae, Dubosiella, and Dubosiella newyorkensis.Conclusion AF promotes the secretion of inflammatory mediators, impairs the permeability of the intestinal mucosa, and alters the structure and diversity of the intestinal flora, thereby promoting the development of IBD.

Key words:inflammatory bowel disease;intestinal flora;Alistipes finegoldii;intestinal barrier