Notenboom Robbert G E, van den Bergh Weerman Marius A, Dingemans Koert P, Vermeulen Jacqueline L M, van den Eijnde Stefan, Reutelingsperger Chris P, Hut Hans, Willemsen Rob, Offerhaus G Johan A, Lamers Wouter H
AMC Liver Center, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
Hepatology. 2003 Sep;38(3):683-91. doi: 10.1053/jhep.2003.50365.
To study the differentiation of hepatocytes along the biliary epithelial lineage in vivo, embryonic day 14 (E14) rat hepatocytes were isolated by differential centrifugation and transplanted as single-cell suspensions into the spleen of adult syngeneic rats. Hepatocytes and cholangiocytes were identified and their maturation characterized by the level of expression of alpha-fetoprotein (AFP), glutamate dehydrogenase (GDH), and carbamoyl phosphate synthetase I (CPS); annexin IV, annexin V, cytokeratin 19 (CK-19), and cystic fibrosis transmembrane conductance regulator (CFTR); and electron microscopy. By correlating morphologic changes with the timing in the expression of these markers, we show that the organization of the transplanted E14 hepatocytes into lobular structures is accompanied by the formation and maturation of bile ducts around these developing lobules. Morphologic differentiation of the emerging bile ducts was accompanied by a gradual loss of hepatocyte markers and a gradual acquisition of cholangiocyte markers, with markers identifying a large-cholangiocyte phenotype appearing latest. Once fully differentiated, the intrasplenic liver lobules developed cholestatic features. The accompanying proliferation of bile ducts was due to cholangiocyte proliferation, but ductular transformation of hepatocytes was also observed. In conclusion, (1) bile duct formation at the interface between hepatocytes and connective tissue is an inherent component of liver development and (2) the susceptibility of developing hepatocytes to bile duct-inducing signals is highest in the fetal liver but that (3) this capacity is not irreversibly lost in otherwise mature hepatocytes.
为了研究体内肝细胞沿胆管上皮谱系的分化情况，通过差速离心法分离出胚胎第14天（E14）大鼠的肝细胞，并将其作为单细胞悬液移植到同基因成年大鼠的脾脏中。通过甲胎蛋白（AFP）、谷氨酸脱氢酶（GDH）和氨甲酰磷酸合成酶I（CPS）的表达水平；膜联蛋白IV、膜联蛋白V、细胞角蛋白19（CK - 19）和囊性纤维化跨膜传导调节因子（CFTR）；以及电子显微镜鉴定肝细胞和胆管细胞，并对其成熟情况进行表征。通过将形态学变化与这些标志物表达的时间相关联，我们发现移植的E14肝细胞组织成小叶结构的过程伴随着这些发育中小叶周围胆管的形成和成熟。新生胆管的形态学分化伴随着肝细胞标志物的逐渐丧失和胆管细胞标志物的逐渐获得，识别大胆管细胞表型的标志物出现最晚。一旦完全分化，脾内肝小叶就会出现胆汁淤积特征。伴随的胆管增殖是由于胆管细胞增殖，但也观察到肝细胞的小胆管化生。总之，（1）肝细胞与结缔组织界面处的胆管形成是肝脏发育的固有组成部分，（2）发育中的肝细胞对胆管诱导信号的敏感性在胎儿肝脏中最高，但（3）在其他方面成熟的肝细胞中这种能力不会不可逆转地丧失。
