Studies of regenerative medicine on differentiation into hepatic and bile duct cells for preclinical trial to surgery

尽管在严重损伤的肝脏中，肝内胆管被认为是肝祖细胞的来源，但关于肝外胆管中是否存在肝祖细胞的问题却鲜有人关注。我们检测了胆管结扎后小鼠EHBD的表型变化。胆管结扎后，EHBD内c-kit阳性的上皮细胞数量增加。结扎的EHBD表达肝祖细胞标志物的mRNA。在胆管结扎后，肝细胞标记物也在EHBD和胆囊中瞬时检测到。在培养的EHBD群体中发现c-kit和白蛋白阳性的肝祖细胞。这些结果表明，将肝干/祖细胞移植到胎肝来分析肝…中的未成熟细胞是可取的更多的发展环境。在这里，我们建立了一种新的监测工具，通过宫外手术将肝祖细胞移植到小鼠胎肝中。胚胎期(ED)14.5d胎肝内注射成肝细胞后，移植细胞表达丰富的白蛋白或弱表达α-胎儿蛋白，并含有糖原，表明移植的成肝细胞能与周围受体实质细胞一致地增殖和分化。移植的细胞在6周龄小鼠的肝脏中成熟。此外，该方法适用于小鼠胚胎干细胞来源的肝母细胞样细胞的移植。这些数据表明，这一独特的技术将为研究肝干/祖细胞的细胞命运和肝脏器官发生提供一个新的体内实验系统。我们揭示了VEGF参与了肝细胞样细胞的高效分化，未分化细胞标志基因DPPA4的生物学功能，以及成熟肝细胞标志基因MAT1A的表观遗传调控。较少

Although the intrahepatic bile duct has been suggested to be a source of hepatic progenitor cells in the severely damaged liver, little attention has been paid to the question of whether hepatic progenitor cells exist in the extrahepatic bile duct (EHBD). We examined the phenotypic changes of the mouse EHBD following the bile duct ligation. After bile duct ligation, the number of c-kit-positive epithelilal cells increased in the EHBD. The ligated EHBD expressed mRNA for hepatic progenitor cell markers. Hepatocyte markers were also transiently detected in the EHBD and gall bladder after a bile duct ligation. Hepatic progenitor cells positive for both c-kit and albumin were found in the cultured EHBD population. These results suggest a potential of the EHBD and gall bladder as useful transplantable sources for liver injury.Avoiding the limitations of the adult liver niche, transplantation of hepatic stem/progenitor cells into fetal liver is desirable to analyze immature cells in a hepati … More c developmental environment. Here, we established a new monitor tool for cell fate of hepatic progenitor cells transplanted into the mouse fetal liver by using ex utero surgery. When embryonic day (ED) 14.5 hepatoblasts were injected into the ED14.5 fetal liver, the transplanted cells expressed albumin abundantly or α-fetoprotein weakly, and contained glycogen in the neonatal liver, indicating that transplanted hepatoblasts can proliferate and differentiate in concord with surrounding recipient parenchymal cells. The transplanted cells became mature in the liver of 6-week-old mice. Furthermore, this method was applicable to transplantation of hepatoblast-like cells derived from mouse ES cells. These data indicate that this unique technique will provide a new in vivo experimental system for studying cell fate of hepatic stem/progenitor cells and liver organogenesis.We revealed involvement of VEGF in efficient differentiation into hepatocyte-like cells, biological function of the undifferentiated cell marker gene DPPA4, and epigenetic regulation of the mature hepatocyte marker gene MAT1A. Less