Hepatocyte differentiation from human ES cells using the simple embryoid body formation method and the staged-additional cocktail

To induce hepatocytes from human embryonic stem (hES) cells easily and effectively, a simple suspension culture method that separates ES colonies with a scraper and transfers them into newly developed, nonadherent MPC (2-methacryloyloxyethyl phosphorylcholine) plates, and the staged-additional cockt...

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Bibliographic Details
Published in:TheScientificWorld 2009-01, Vol.9, p.884-890
Main Authors: Sasaki, Katsunori, Ichikawa, Hinako, Takei, Shunsuke, No, Hee Sung, Tomotsune, Daihachiro, Kano, Yoshiya, Yokoyama, Tadayuki, Sirasawa, Sakiko, Mogi, Akimi, Yoshie, Susumu, Sasaki, Shujiro, Yamada, Satoshi, Matsumoto, Ken, Mizuguchi, Masahiro, Yue, Fengming, Tanaka, Yoshiki
Format: Article
Language:English
Subjects:
Base Sequence
Cell differentiation
Cell Line
DNA Primers
Embryonic development
Embryonic stem cells
Embryonic Stem Cells - cytology
Embryonic Stem Cells - ultrastructure
Health aspects
Hepatocytes - cytology
Humans
Liver cells
Microscopy, Electron, Transmission
Physiological aspects
Reverse Transcriptase Polymerase Chain Reaction
Short Communication
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Summary:To induce hepatocytes from human embryonic stem (hES) cells easily and effectively, a simple suspension culture method that separates ES colonies with a scraper and transfers them into newly developed, nonadherent MPC (2-methacryloyloxyethyl phosphorylcholine) plates, and the staged-additional cocktail method, including growth factors, cytokines, and Lanford serum-free medium, were developed and evaluated mainly by morphological analysis. The formed embryoid bodies (EBs) showed compact cellular agglomeration until day 4 and later formed coeloms in their interior. RT-PCR (reverse transcriptase-polymerase chain reaction) analysis showed that they are gene markers of the three germ layers. Mesenchymal cells with rough endoplasmic reticulum (rER) and extracellular matrix (ECM), and without junctions, were recognized in the interior of the EBs by transmission electron microscopy (TEM) in addition to epithelial cells. When they were stimulated by the staged-additional cocktail, they expressed albumin-positive immunoreactivity, indocyanine green (ICG) uptake, and typical ultrastructures of the hepatocytes, including bile canaliculi. These results indicate that these combined methods promote EB formation and hepatocyte differentiation from hES cells.
ISSN:1537-744X
2356-6140
1537-744X
DOI:10.1100/tsw.2009.97