Open AccessSelective enrichment of hepatocytes from mouse embryonic stem cells with a culture system containing cholestatic serum 1
Author(s) -
MIN Jun,
SHANG Changzhen,
CHEN Yajin,
ZHANG Lei,
LIU Lu,
DENG Xiaogeng,
YANG Mei,
CHEN Dongping,
CAO Jun,
SONG Erwei,
CHEN Jisheng
Publication year - 2007
Publication title -
acta pharmacologica sinica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.514
H-Index - 90
eISSN - 1745-7254
pISSN - 1671-4083
DOI - 10.1111/j.1745-7254.2007.00715.x
Subject(s) - hepatocyte , sodium butyrate , biology , cellular differentiation , microbiology and biotechnology , immunocytochemistry , stem cell , amniotic epithelial cells , embryonic stem cell , cell culture , cytokeratin , in vitro , biochemistry , adult stem cell , immunology , immunohistochemistry , endocrinology , genetics , gene
Aim : There is increasing evidence indicating that embryonic stem (ES) cells are capable of differentiating into hepatocyte‐like cells in vitro. However, it is necessary to improve the differentiation efficiency so as to promote the clinical application. Here, we report an efficient culture system to support hepatocyte differentiation from ES cells by utilizing cholestatic serum. Methods : One week after the induction of E14 mouse ES cells into hepatocytes with sodium butyrate, cholestatic serum was added into the culture system at various concentrations and hepatocyte‐like cells were induced to proliferate. The morphological and phenotypic markers of hepatocytes were characterized using light microscopy, immunocytochemistry, and RT‐PCR, respectively. The function of glycogen storage of the differentiated cells was detected by Periodic acid‐Schiff (PAS) reaction, and the ratio of hepatic differentiation was determined by counting the albumin and PAS‐positive cells. Results : In the presence of conditional selective medium containing cholestatic serum, numerous epithelial cells resembling hepatocytes were observed. The RT‐PCR analysis showed that undifferentiated ES cells did not express any hepatic‐specific markers; however, in the presence of sodium butyrate and conditional selective medium containing cholestatic serum, hepatic differentiation markers were detected. Immunofluorescence staining showed that those ES‐derived hepatocytes were α‐fetoprotein, albumin, and cytokeratin 18 positive, with the ability of storing glycogen. Further determination of the hepatic differentiation ratio showed that the application of cholestatic serum efficiently enriched ES‐derived hepatocyte‐like cells by inducing lineage differentiation and enhancing lineage proliferation. Conclusion : The conditional selective medium containing cholestatic serum is optimal to selectively enrich hepatocyte‐like cells from mixed differentiated ES cells, which may provide a novel method to improve the hepatic differentiation ratio of ES cells.