2005 Fiscal Year Final Research Report Summary
Treatment of liver failure by transplantation of hepatocytes derived from mouse and monkey ES cells and human umbilical cord blood
| Project/Area Number |
16390357
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General surgery
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| Research Institution | Tokyo medical and Dental University |
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Principal Investigator |
TERAMOTO Kenichi Tokyo medical and Dental University, surgery, Assistant professor, 大学院・医歯学総合研究科, 助教授 (80197813)
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| Co-Investigator(Kenkyū-buntansha) |
ARII Shigeki Tokyo medical and Dental University, surgery, professor, 大学院・医歯学総合研究科, 教授 (50151171)
TANAKA Yujiro Tokyo medical and Dental University, professor, 医学部付属病院, 教授 (70236644)
SAITO Keiko Tokyo medical and Dental University, instructor, 難治疾患研究所, 教務職員 (50178969)
ASAHINA Kinji Tokyo medical and Dental University, Assistant, 難治疾患研究所, 助手 (40345294)
TERAOKA Hirobumi Tokyo medical and Dental University, M.R.I, professor, 難治疾患研究所, 教授 (30019137)
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| Project Period (FY) |
2004 – 2005
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| Keywords | Cynomolgus / Embryonic stem cells / Hepatocyte differentiation / Transgenic mouse / umbilical cord blood / liver failure / hepatocyte transplantation / teratoma |
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| Research Abstract |
1) Monkey embryonic stem (ES) cells have characteristics that are similar to human ES cells, and might be useful as a substitute model for preclinical research. When embryoid bodies (EBs) formed from monkey ES cells were cultured, expression of many hepatocyte-related genes including cytochrome P450 (Cyp) 3a and Cyp7a1 was observed. Hepatocytes were immunocytochemically observed using antibodies against albumin (ALB), cytokeratin-8/18, and a1-antitrypsin in the developing EBs. The in vitro differentiation potential of monkey ES cells into the hepatic lineage prompted us to examine the transplantability of monkey EB cells. As an initial approach to assess the repopulation potential, we transplanted EB cells into immunodeficient urokinasetype plasminogen activator transgenic mice that undergo liver failure. After transplantation, the hepatocyte colonies expressing monkey ALB were observed in the mouse liver. Fluorescence in-situ hybridization revealed that the repopulating hepatocytes ar
… More
ise from cell fusion between transplanted monkey EB cells and recipient mouse hepatocytes. In contrast, neither cell fusion nor repopulation of hepatocytes was observed in the recipient liver after undifferentiated ES cell transplantation. These results indicate that the differentiated cells in developing monkey EBs, but not contaminating ES cells, generate functional hepatocytes by cell fusion with recipient mouse hepatocytes, and repopulate injured mouse liver.
2) We previously reported that hepatocytes can be differentiated from embryonic stem (ES) cells by way of embryoid body (EB) formation and are transplantable into the mouse liver. However, the transplantation of EB-derived cells frequently resulted in teratoma formation in the recipient liver. In the present study, we eliminated the tumorigenic cells from EB outgrowths and examined the effects of enriched ES-cell-derived hepatocyte transplantation into an injured liver. On day 15 in culture, the EBs were partially disaggregated and subcultured. Hepatocytes in the subcultured cells were examined by the expression of hepatocyte markers. Undifferentiated cells contaminating in the EB-derived cells were eliminated by Percoll discontinuous gradient centrifugation. Furthermore, undifferentiated cells, endothelial cells, and macrophages were eliminated by magnetic cell sorting using platelet/endothelial cell adhesion molecule (PECAM)-1 and Mac-1 antibodies. These enriched ES-cell-derived hepatocytes were then transplanted into the injured mouse liver. Percoll centrifugation and PECAM-1 antibodies eliminated the undifferentiated cells expressing Oct-3/4 from the EB-derived cells. ES-cell-derived hepatocytes showed expression of liver-related genes, synthesis of urea and glycogen, and structural characteristics during subculture. A transplantation study showed that the enriched ES-cell-derived hepatocytes integrated into the injured mouse liver and produced no teratomas. When the ES-cell-derived hepatocytes were transplanted into a CCl4-injured liver, the liver function was subsequently improved. Functional hepatocytes can be differentiated from mouse ES cells by way of EB formation. The elimination of undifferentiated cells from the EBs provides transplantable cells for liver failure without tumorigenicity. Less
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Research Products
(10 results)
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[Journal Article] Enrichment of Hepatocytes Differentiated from Mouse Embryonic Stem Cells as a Transplantable Source.2005
Author(s)
Yuji Kumashiro, Kimji Asahina, Rie Ozeki, Keiko Shimizu-Saito, Yujiro Tanaka, Yujiro Kida, Kouji Inoue, Michinari Kaneko, Tetsuji Sato, Kenichi Teramoto, Shigeki Arii, Hirobumi Teraoka
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Journal Title
Transplantation 79(5)
Pages: 550-557
Description
「研究成果報告書概要(和文)」より
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[Journal Article] Generation of hybrid hepatocytes by cell fusion from monkey embryoid body cells in the injured mouse liver.2005
Author(s)
Kentaro Okamura, Kinji Asahina, Hiroaki Fujimori, Rie Ozeki, Keiko Shimizu-Saito, Yujiro Tanaka, Kenichi Teramoto, Shigeki Arii, Kozo Takase, Miho Kataoka, Yoshinori Soeno, Chise Tateno, Katsutoshi Yoshizato, Hirobumi Teraoka
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Journal Title
Histochemistry and Cell Biology 125(3)
Pages: 245-257
Description
「研究成果報告書概要(和文)」より
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