以斑马鱼为模式生物探究消化系统早期发育中的细胞命运调控机制

Digestive system includes digestive tract and digestive gland, which are derived from endodermal cells. In the digestive system, the extrahepatic bile duct joins with the pancreatic duct to form a common duct connecting the digestive tract, and the secretions from liver and pancreas are transported to the digestive tract through the common duct. At present, little research has been done on how the digestive glands form a common duct and connect to the digestive tract accurately. .In our previous study, we found that Hhex functions as an important transcriptional repressor and represses the expression of cdx1b and pdx1 to safeguard the development of the bile duct system. In the hhex mutant, cdx1b and pdx1 are expressed ectopically in the bile duct progenitor cells. This ectopic expression changes the cell fate of bile duct progenitor cells, which results into loss of bile duct system and the formation of an “intrahepatic intestinal tube” in the mutant..In our recently study, we found that the liver, bile duct, duodenum and pancreas may originate from the endodermal precursor cells in the same region. We suppose that transcription factors such as Hhex, Cdx1b and Pdx1 form regulatory networks in this region, which determine the cell fate of different cells and developmental direction of these organs. Recently, we also found that this process may be directly regulated by BMP and other mesodermal signals..The formation of an organism relies on the generation of diverse cell types. Within the field of developmental biology one goal is to understand how a cell's fate is determined. This project aims to reveal the molecular mechanism of cell fate determination in endodermal precursor cells development by means of cell lineage tracing, immunofluorescence labeling and ChIP. Our studies on cell lineage and cell fate regulators will contribute to refine the protocols for stem cells and cell induction in vivo and in vitro, which is of great significance for the treatment of diseases in liver, intestine and other digestive organs.

在消化系统形态发生过程中，肝脏，胆管，前肠和外分泌胰腺细胞起源于同一区域的内胚层前体细胞。目前对于决定这片前体细胞命运的直接调控机制研究较少。我们前期的研究发现转录因子Hhex通过直接抑制转录因子Pdx1和Cdx1b的表达来决定胆管细胞命运。hhex突变体中胆管和胆囊缺失，Pdx1和Cdx1b在原胆管位置异位表达，从而导致肝脏内发育出一个具肠道特性的导管结构。我们推测Hhex, Cdx1b和Pdx1等转录因子形成有序的调控网络决定这区域细胞的命运及发育方向，且这一过程受到BMP等信号的直接调控。本项目拟构建相关的遗传材料和转基因鱼品系，通过细胞谱系示踪、染色质共沉淀等方法揭示直接调控肝胆胰导管细胞命运决定的分子机制，研究结果将有利于我们认识消化系统形态发生的细胞和分子行为，同时将有助于指导体外和体内细胞诱导研究，进而为消化系统疾病的治疗提供理论基础。

在之前的研究中，我们发现Hhex蛋白发挥转录抑制功能，进而保证胆管细胞的正常分化和发育。.在本项目的研究中，我们又进一步发现cdx1b基因缺失会导致肠道细胞出现肝样化。利用探针原位技术证明，cdx1b突变体的肠道中出现了Fabp10a等肝脏特异基因的异位表达。通过精细的解剖取样，我们对野生型和突变体斑马鱼肠道进行了取样及RNA测序和单细胞测序，分析显示突变体斑马鱼肠道细胞中有大量肝实质细胞特异基因的表达上调。ATAC测序证明cdx1b基因缺失导致肝脏富集基因的染色体位置趋于开放。进一步的分子机制研究证明该肠样化表型受到BMP和WNT等信号通路调控，且可以被prox1a敲除拯救。我们的研究显示Hhex和Sox9b、Cdx1b及Pdx1可能组成了一个调控胆管-肠道组织形态发生和细胞命运决定的信号通路。.此外，依托本课题，我们还建立了实验室自主的微孔板技术平台。