In Vitro Differentiation of Liver Progenitor Cells Derived from Healthy Dog Livers
In Vitro Differentiation of Liver Progenitor Cells Derived from Healthy Dog Livers
复制标题
DOI：
10.1089/scd.2008.0043
复制
发表时间：
2009-03-01
影响因子：
4
通讯作者：
Rothuizen, Jan
中科院分区：
文献类型：
--
作者：
Arends, Brigitte;Spee, Bart;Rothuizen, Jan
犬中自然发生的肝病与人类肝病非常相似;包括急性和慢性肝病中肝祖细胞（LPC）的激活。本研究的目的是分离、培养和鉴定来自健康成熟狗肝脏的祖细胞。分离富含小肝细胞的非实质细胞组分，并在无肝酶血清培养基（SFM）中培养，以刺激集落形成小上皮细胞的生长。培养2周后，在肝细胞单层中出现角蛋白7（K7）免疫阳性小细胞的克隆扩增，具有大的核/质比。这些集落表达几种肝细胞（CYP 1A 1、ALB和KRT 18）、胆管细胞/LPC（KRT 7和KRT 19）和祖细胞标志物（甲胎蛋白、CD 44、CD 44蛋白1、KIT、THY 1和神经细胞粘附分子1）的基因，表明其不成熟和双能性。基因表达谱表明肝酶-SFM中的肝分化比威廉培养基E（WME）中的肝分化更明显。此外，集落形成细胞向中间肝细胞样细胞分化，具有更明显的膜K7免疫染色。总之，长期犬肝细胞培养中的集落形成小上皮细胞表达LPC标志物，并具有分化能力。因此，这些细胞可以被认为是肝脏的祖细胞。
Naturally occurring liver disease in dogs resemble human liver disease in great detail; including the activation of liver progenitor cells (LPC) in acute and chronic liver disease. The aim of the present study was to isolate, culture, and characterize progenitor cells derived from healthy mature dog livers. A nonparenchymal cell fraction enriched with small hepatocytes was isolated and cultured in Hepatozyme-serum-free media (SFM) to stimulate the growth of colony-forming small epithelial cells. After 2 weeks of culturing, clonal expansion of keratin 7 (K7) immunopositive small cells with a large nucleus/cytoplasm ratio emerged in the hepatocyte monolayer. These colonies expressed genes of several hepatocyte (CYP1A1, ALB, and KRT18), cholangiocyte/LPC (KRT7 and KRT19), and progenitor cell markers (alpha-fetoprotein, CD44, prominin1, KIT, THY1, and neural cell adhesion molecule 1), indicating their immature and bipotential nature. Gene-expression profiles indicated a more pronounced hepatic differentiation in Hepatozyme-SFM compared to William's Medium E (WME). Furthermore, colony-forming cells differentiated toward intermediate hepatocyte-like cells with a more pronounced membranous K7 immunostaining. In conclusion, colony-forming small epithelial cells in long-term canine liver cell cultures express LPC markers and have differentiating capacities. These cells may therefore be considered as progenitor cells of the liver.