Mice harboring pathobiont-free microbiota do not develop intestinal inflammation that normally results from an innate immune deficiency

Background
Inability to maintain a stable and beneficial microbiota is associated with chronic gut inflammation, which classically manifests as colitis but may more commonly exist as low-grade inflammation that promotes metabolic syndrome. Alterations in microbiota, and associated inflammation, can originate from dysfunction in host proteins that manage the microbiota, such as the flagellin receptor TLR5. That the complete absence of a microbiota (i.e. germfree conditions) eliminates all evidence of inflammation in TLR5-deficient mice demonstrates that this model of gut inflammation is microbiota-dependent. We hypothesize that such microbiota dependency reflects an inability to manage pathobionts, such as Adherent-Invasive E. coli (AIEC). Herein, we examined the extent to which microbiota mismanagement and associated inflammation in TLR5-deficient mice would manifest in a limited and pathobiont-free microbiota. For this purpose, WT and TLR5-deficient mice were generated and maintained with the 8-member consortium of bacteria referred to as “Altered Schaedler Flora” (ASF). Such ASF animals were subsequently inoculated with AIEC reference strain LF82. Feces were assayed for bacterial loads, fecal lipopolysaccharide and flagellin loads, fecal inflammatory marker lipocalin-2 and microbiota composition.
Results
Relative to similarly maintained WT mice, mice lacking TLR5 (T5KO) did not display low-grade intestinal inflammation nor metabolic syndrome under ASF conditions. Concomitantly, the ASF microbial community was similar between WT and T5KO mice, while inoculation with AIEC strain LF82 resulted in alteration of the ASF community in T5KO mice compared to WT control animals. AIEC LF82 inoculation in ASF T5KO mice resulted in microbiota components having elevated levels of bioactive lipopolysaccharide and flagellin, a modest level of low-grade inflammation and increased adiposity.
Conclusions
In a limited-complexity pathobiont-free microbiota, loss of the flagellin receptor TLR5 does not impact microbiota composition nor its ability to promote inflammation. Addition of AIEC to this ecosystem perturbs microbiota composition, increases levels of lipopolysaccharide and flagellin, but only modestly promotes gut inflammation and adiposity, suggesting that the phenotypes previously associated with loss of this innate immune receptor require disruption of complex microbiota.

Background
无法维持稳定且有益的微生物群（microbiota）与慢性肠道炎症相关，这类炎症典型表现为结肠炎，但更常见的是以低度炎症形式存在，进而促进代谢综合征（metabolic syndrome）的发生。微生物群改变及伴随的炎症可源于宿主调控微生物群的蛋白功能异常，例如鞭毛蛋白受体TLR5（flagellin receptor TLR5）。完全清除微生物群（即无菌环境，germfree conditions）可消除TLR5缺陷小鼠的所有炎症迹象，这证明该肠道炎症模型是微生物群依赖性的。我们提出如下假说：这种微生物群依赖性反映了机体无法管控致病共生菌（pathobionts），例如黏附侵袭性大肠杆菌（Adherent-Invasive E. coli, AIEC）。本研究旨在探究，在有限且不含致病共生菌的微生物群环境中，TLR5缺陷小鼠的微生物群失调及相关炎症会呈现何种表现。为此，我们构建并饲养了野生型（wild type, WT）与TLR5缺陷小鼠，采用被称为"改良谢德勒菌群（Altered Schaedler Flora, ASF）"的8种细菌联合体进行饲养。随后，对这类ASF饲养的小鼠接种AIEC标准菌株LF82。通过粪便检测分析细菌载量、粪便脂多糖（lipopolysaccharide）和鞭毛蛋白载量、粪便炎症标志物脂钙蛋白-2（lipocalin-2）以及微生物群组成。
Results
与同样以ASF饲养的野生型小鼠相比，TLR5缺陷小鼠（T5KO）在ASF饲养环境下未出现低度肠道炎症，也未发生代谢综合征。与此同时，野生型小鼠与T5KO小鼠的ASF微生物群落结构相似；但与野生型对照组相比，T5KO小鼠接种AIEC菌株LF82后，其ASF群落结构发生了改变。对ASF饲养的T5KO小鼠接种AIEC LF82后，其微生物群组分的生物活性脂多糖和鞭毛蛋白水平升高，出现轻度低度炎症，且脂肪量增加。
Conclusions
在有限复杂度且不含致病共生菌的微生物群环境中，鞭毛蛋白受体TLR5的缺失不会影响微生物群组成，也不会影响其促炎症能力。向该生态系统中加入AIEC会扰乱微生物群组成，提升脂多糖和鞭毛蛋白水平，但仅会轻度促进肠道炎症和脂肪堆积，这表明此前与该先天免疫受体缺失相关的表型，需要复杂微生物群的紊乱作为前提。