山羊小肠细菌定植与粘膜天然免疫应答的关联机制研究

Plenty of bacteria, which differ in number, diversity and function, inhabit in the digesta and mucosa of the ruminant small intestine. The small intestine (especially the jejunum and ileum) is the primary organ of innate immune responses, and colonization of most commensal bacteria can enhance intestinal barrier function and innate immune function, playing a key role in the prevention of diseases. Relationship between intestinal bacteria colonization and mucosal innate immune responses is an important part of host-bacteria interaction and this topic has been scarcely reported in ruminants. Therefore, in this project, we use goats at different development phases (non-rumination, transition and rumination phases) as experimental animals. The aims of this study were (1) to investigate age-related bacteria colonization process (diversity, distribution and function) in the jejunum and ileum, (2) to explore changes in jejunal and ileal mucosal innate immune function and related signaling pathway during different development phases, (3) to reveal correlation mechanism of bacteria colonization and mucosal innate immune responses in the small intestine of goats. These results will be helpful to provide new ideas to explore host-bacteria interaction, as well as to establish scientific basis for improving nutrition manipulation and keeping health of the intestinal tracts.

反刍家畜小肠的食糜及粘膜上皮中栖息着数量庞大的细菌，它们在数量、种类及功能上都存在显著差异。小肠（尤其是空肠、回肠）是机体发生天然免疫应答的重要器官，且很多共生菌的定植可以增强小肠粘膜的屏障功能及天然免疫功能，以防止疾病的发生。探讨小肠细菌的定植与宿主粘膜天然免疫应答的关联机制是研究宿主与细菌相互作用的一个重要方面，而关于这一科学问题的研究目前在反刍家畜上报道较少。本项目拟以山羊为试验动物，研究其在不同发育阶段（非反刍、过渡和反刍阶段），空肠和回肠细菌定植（从出生到反刍功能形成）、分布（食糜和粘膜上皮）与功能演变（代谢产物与代谢通路）规律；解析小肠粘膜天然免疫应答信号通路关键指标的变化规律；并通过整合分析及相关分析阐明小肠细菌定植与粘膜天然免疫应答的关联机制。旨在为研究细菌与宿主的相互作用提供思路，并为反刍家畜各阶段的饲养管理与肠道健康稳衡调控提供科学依据。

反刍家畜的小肠是机体发生天然免疫应答的重要器官，其食糜及粘膜上皮中栖息着的共生细菌的定植可以增强小肠粘膜的天然免疫功能。本项目采用营养学、微生物学、分子生物学以及多组学等研究手段，系统研究了山羊不同发育阶段，小肠细菌的定植、分布及功能演变规律；解析小肠粘膜天然免疫应答信号通路关键指标的变化规律；并通过整合分析阐明关联机制。取得的主要结果如下：. （1）幼龄山羊空肠的细菌组成在食靡和粘膜存在极大差异，在粘膜中显著富集的细菌主要是Faecalibacterium和Roseburia，而在食靡中显著富集的主要是Ruminococcus, Eubacterium, Clostridium_sensu_stricto, Lactobacillus, Streptococcus等。随着日龄的增长，空肠食靡的Lactobacillus和Butyricicoccus属的含量显著降低，而Ruminococcus, Eubacterium 和 Butyrivibrio的含量提高了5-10倍。空肠粘膜的Bifidobacterium, Corynebacterium, Faecalibacterium 和 Roseburia的含量增加了5-20倍。对于回肠食靡，Lactobacilllus和Streptococcus的丰度随日龄增长而下降。. （2）回肠细菌的代谢组学（d60 vs d10）筛选出了6个碳水化合物、37个氨基酸和128个脂质相关的差异代谢产物，表明回肠细菌的3条氨基酸代谢通路、淀粉及蔗糖代谢通路等均在随日龄增长而增强。宏基因组的结果表明反刍阶段回肠细菌的碳水化合物、氨基酸、脂质代谢、能量代谢等均显著增强。. （3）小肠的组织形态随日龄增长而逐渐完善，同时日龄对小肠粘膜天然免疫相关基因的表达有调控作用。在蛋白层面，紧密连接蛋白的表达量随日龄增长而降低。. （4）相关分析结果表明有可能的介导粘膜天然免疫应答的细菌属包括Shewanella、Eubacterium、Ruminococcus和Streptococcus。. （5）营养调控试验表明，大黄添加可以提高幼龄山羊回肠屏障功能，且这一功能的改善与粘膜细菌群落的改变有密切关系。. 此研究阐明了山羊小肠细菌定植与粘膜天然免疫应答的关联机制，并为反刍家畜各阶段的饲养管理与肠道健康稳衡调控提供了科学依据。