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Development of expanding and differentiation-promoting system ex vivo for liver regeneration

肝再生体外扩增和分化促进系统的开发

基本信息

批准号：
16390209
负责人：
SHIOTA Goshi
金额：
$ 9.15万
依托单位：
National University Corporation Tottori University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16390209/
关键词：
stem cell bone marrow mesenchymal stem cell hepatocytet cytokine liver regeneration treatment demethylation agent HDAC inhibitor TERT 骨髄間葉系細胞臍帯血幹細胞 HDAC抑制剤アルブミン

项目摘要

For the purpose of development of liver regeneration therapy, we attempted to develop the novel methods, by which human mesenchymal stem cells (MSCs) can be differentiated to mature hepatocytes. We used bone marrow-derived MSCs and cord blood-derived MSCs.1.Study by using bone marrow-derived MSCsUsing UE7T-13 cells, human bone marrow-derived MSCs which were retrovirally infected with human papilloma virus E7 and human telomerase reverse transcriptase (hTERT), 40 conditions of several cytokines, 5-azacytidine and type IV collagen coating were examined on hepatic differentiation of UE7T-13 cells. As a result, the optical condition for differentiation is type IV collagen-coating/aFGF/bFGF/HGF. By this condition, expression of hepatocyte-specific genes was highest. In addition, glycogen synthesis and urea synthesis was also greatly promoted by this treatment. Under this circumstance, WISP1 and WISP2 were down-regulated. Knockdown of WISP1 and WISP2 by siRNA also greatly enhanced hepatic differentiation of of MSCs.2.Study by using cord blood-derived MSCsUsing UCBTERT-21 cells, human cord blood-derived MSCs which were retrovirally infected with human telomerase reverse transcriptase (hTERT), 8 conditions of several cytokines, 5-azacytidine, HDAC inhibitor were examined on hepatic differentiation of UCBTERT-21 cells. As a result, the optical condition for differentiation is 5-azacytidine/HGF/aFGF/OSM. By this condition, expression of hepatocyte-specific genes was highest. In addition, glycogen syntheisis and urea syntheisis was also greatly promoted by this treatment. Under this circumstance, Wnt signal was down-regulated. Knockdown of fzd8, the receptor of Wnt, WISP1 also greatly enhanced hepatic differentiation of of MSCs.These results indicated new findings of hepatic differentiation of MSCs ; and contribute to the development of expanding and differentiation-promoting system ex vivo for liver regeneration.

为了开发肝再生疗法，我们试图开发新的方法，使人间充质干细胞(MSCs)能够分化为成熟的肝细胞。利用UE7T-13细胞、逆转录病毒感染的人骨髓间充质干细胞和人端粒酶逆转录酶(HTERT)、多种细胞因子、5-氮胞苷和IV型胶原涂层等40种条件对UE7T-13细胞的肝分化进行了研究。结果表明，诱导分化的最佳条件为IV型胶原涂层/aFGF/bFGF/HGF。在此条件下，肝细胞特异性基因的表达最高。此外，对糖原合成和尿素合成也有明显的促进作用。在这种情况下，WISP1和WISP2表达下调。2.以脐血MSCs为研究对象，观察了逆转录病毒感染人端粒酶逆转录酶(HTERT)、8种细胞因子、5-氮胞苷、HDAC抑制剂对脐血MSCs向UCBTERT-21细胞分化的影响。结果表明，分化的最佳条件为5-氮杂胞苷/HGF/aFGF/OSM。在此条件下，肝细胞特异性基因的表达最高。此外，对糖原合成和尿素合成也有明显的促进作用。在这种情况下，Wnt信号被下调。Wnt、WISP1受体FZD8基因的敲除也显著促进了MSCs的肝脏分化，这些结果提示了MSCs肝脏分化的新发现，并有助于建立体外扩增和分化促进系统以促进肝脏再生。