树突状细胞介导孕妇肠道菌群进入胎盘和羊水的研究

Healthy pregnant women placenta and amniotic fluid contain a certain number of microorganisms. These findings indicated that the colonization of human gut microbiota was from fetal period. However, till now the sources and transfer pathway of placenta and amniotic fluid microbita were not fully understood. Our previous studies indicated that gut permeability significantly increased in late period of pregnant rats. We speculate that late pregnancy period may be the critical period for the transfer of gut microbiota; gut microbiota may be the source of placenta and amniotic fluid microbiota. Based on the previous studies, in order to illustrate the gut microbiota is the source of placenta and amniotic fluid microbiota, this study will collect pregnant women feces, placenta and amniotic fluid samples, analyze the correlation of gut microbiota and placenta microbiota. To clarify the “window period”of gut microbiota transfer, intestinal wall tight junction proteins were examined in different pregnancy period of pregnant rats. To reveal the process that gut microbiota enters into placenta and amniotic fluid, GFP labeled E. cloacae were used for rat gavage in different pregnancy phase. Observe GFP labeled E. cloacae in intestinal wall, mesenteric lymph nodes, blood, placenta and amniotic fluid by fluorescence microscope. Immunofluorescence test was used to understand the assistant role of CD11c+ dendritic cells in the process of gut mictobiota transfer. The implementation of this study is of great significance in revealing the colonization mechanism of microbiota in fetal growth environment, and laying theoretical basis for regulation of pregnant women and fetal flora and promoting maternal and child health.

健康孕妇胎盘、羊水中含有微生物，此项发现将人类肠道菌群定植的窗口期提前至胎儿期。胎盘、羊水中菌群的来源及转移过程尚不明确。我们的前期研究显示妊娠晚期孕鼠肠壁的通透性增加，推测妊娠晚期为肠道菌群定位转移的关键时期，肠道菌群为胎盘羊水菌群的来源。本研究拟从前期工作基础出发，采集孕妇粪便、胎盘、羊水样本进行菌群相关性分析，明确肠道菌群是胎盘、羊水的菌群来源；通过不同孕期母鼠肠壁紧密连接蛋白的检测，分析肠道菌群定位转移的窗口期；应用GFP标记的肠杆菌（E. cloacae）进行不同孕期灌胃，荧光显微镜观察肠壁、肠系膜下淋巴结、血液、胎盘、羊水中GFP-肠杆菌的情况，以期揭示肠道菌群进入胎盘、羊水的过程；免疫荧光检测CD11c+树突状细胞在GFP-肠杆菌定位转移过程中的协助作用。本研究的实施对于揭示生命早期胎儿生长环境中菌群的定植机制具有重要意义，为未来调控孕妇、胎儿菌群，促进母婴健康奠定理轮基础。

本研究探讨孕妇肠道菌群与婴儿胎便菌群、羊水菌群的相关性，并应用绿色荧光蛋白标记的大肠杆菌干预孕鼠模型，研究羊水菌群的来源及菌群定位转移的相关机制。按照研究计划，初步证明了肠道菌群是羊水菌群的来源之一，血液中的树突状细胞及其它单核细胞携带肠道菌群能够协助肠道菌群的定位转移。研究为探索机体生命早期菌群来源及定位转移相关机制，促进母婴健康奠定了基础，达到了预先的目标。已发表SCI论文2篇，核心期刊论文3篇，在投SCI论文1篇。 .本研究通过16s-rDNA 测序技术分析了不同代谢状态的孕妇粪便菌群及所产婴儿胎便菌群，证实了孕妇的肠道菌群直接影响婴儿的胎便菌群。孕妇肠道菌群与婴儿胎便菌群存在明显的相关关系，但同时胎便中尚存在不同于孕妇肠道菌群的特有微生物，提示孕妇肠道菌群并非婴儿生命早期菌群建立的唯一来源。进一步针对孕妇孕中期及孕晚期的羊水菌群分析，证实孕中期即有可检测到的菌群的DNA进入羊水中，且孕中期与孕期晚期羊水中菌群并不完全相同，提示机体体内存在精细的调控机制，调控进入羊水的菌群。.本研究利用荧光标记的大肠杆菌给孕鼠灌胃，追踪肠道菌群定位转移的机制，证实了肠道菌群能够进入到羊水中，通过对血液中树突细胞，单核细胞分离检测，证实树突状细胞能够携带GFP-标记的大肠杆菌。此外，血液中的单核细胞也可以携带GFP-标记的大肠杆菌，参与肠道菌群的定位转移。本研究对于羊水菌群来源及相关机制的探讨，拓展了目前关于羊水菌群的认识，也为未来深入进行生命早期菌群建立的研究提供了新的思路。