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Identification of human cord blood-derived hepatic stem cells, and establishment of cell therapy for lethal liver disease

人脐带血肝干细胞的鉴定及致命性肝病细胞疗法的建立

基本信息

批准号：
16590581
负责人：
KAKINUMA Sei
金额：
$ 2.24万
依托单位：
The University of Tokyo (2005)Tokyo Medical and Dental University (2004)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
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

项目摘要

In this grant in aid for scientific research, we carried out a research project on development of cell therapy using human umbilical cord blood. In the project, we planned that identification of hepatic stem cells from cord blood through in vitro and in vivo assay, and establishment of therapeutic animal models for lethal liver disease using cord blood cell after the treatment of differentiation into functional hepatocytes. The outcome of the research was as follows.At first, we purified the cells derived from cord blood using fluorescence-activated cell sorter. The positive and negative fractions of cell surface markers (i.e., CD45, CD34, c-met, lineage markers, and so on) were sorted, and were cultured in our original medium supplemented with FGF-1, FGF-2, HGF, SCF and LIF. In this culture condition, total cord blood cells were able to produce albumin and metabolite ammonia. The in vitro assay revealed that the purified CD45-positive cells and lineage markers-negative cells derived f … More rom cord blood can express albumin mRNA. Further examination showed that purified CD34-positive cells were able to express albumin mRNA in our original culture condition.Next, we performed a transplantation assay using purified cord blood cells. Cord blood cells were transplanted into liver-injured recipient mice, and their liver and the sera were analyzed. After the cell transplantation, the liver of recipient mice transplanted with CD34-positive cells contained human albumin-producing cells, indicating that cord blood CD34-positive cells engrafted in the liver and gave rise to hepatocytes. Furthermore, in the case of chronically liver-injured recipient mice, transplanted total cord blood cells were able to express marker-genes of functionally mature hepatocytes. These results showed that cord blood cells contain the hepatic progenitor cells, which can give rise to functionally mature hepatocytes.In conclusion, our data implied that human umbilical cord blood could be a source of hepatic progenitor cells for liver disease. The outcome of the research project will be a basis of liver regenerative medicine, and we think the purpose of this grant in aid for scientific research was accomplished. Less

在这笔资助科学研究的赠款中，我们开展了一项利用人脐带血开发细胞疗法的研究项目。在该项目中，我们计划通过体外和体内实验从脐带血中鉴定出肝干细胞，并在诱导分化为有功能的肝细胞后利用脐带血建立致死性肝病的治疗性动物模型。研究结果如下：首先，利用荧光激活细胞分选机对脐带血来源的细胞进行纯化。对细胞表面标志(CD45、CD34、c-met、谱系标志等)阳性和阴性的部分进行分选，并在添加了成纤维细胞生长因子-1、成纤维细胞生长因子、肝细胞生长因子、干细胞因子和激光诱导因子的原始培养液中进行培养。在此培养条件下，脐带血总细胞能够产生白蛋白和代谢物氨。体外实验表明，纯化的CD45阳性细胞和谱系标志阴性的细胞来源于…。更多的脐带血可以表达白蛋白mRNA。进一步的检测表明，纯化的CD34阳性细胞在我们最初的培养条件下能够表达白蛋白mRNA。接下来，我们用纯化的脐血细胞进行了移植实验。将脐带血移植到肝损伤受体小鼠体内，并对其肝脏和血清进行分析。细胞移植后，移植CD34阳性细胞的受体小鼠肝脏中含有人白蛋白产生细胞，表明脐带血CD34阳性细胞植入肝脏并长出肝细胞。此外，在慢性肝损伤受体小鼠中，移植的全脐血细胞能够表达功能成熟的肝细胞的标记基因。这些结果表明，脐带血中含有肝祖细胞，可以诱导出功能成熟的肝细胞，提示人脐血可能是肝病的肝祖细胞来源。该研究项目的成果将成为肝脏再生医学的基础，我们认为这笔资助科学研究的目的已经实现。较少