Establishment of a Cell Line Derived from a Mouse Fetal Liver That Has the Characteristic to Promote the Hepatic Maturation of Mouse Embryonic Stem Cells by a Coculture Method
Establishment of a Cell Line Derived from a Mouse Fetal Liver That Has the Characteristic to Promote the Hepatic Maturation of Mouse Embryonic Stem Cells by a Coculture Method
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DOI：
10.1089/ten.tea.2009.0357
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发表时间：
2009-12-01
影响因子：
4.1
通讯作者：
Uemoto, Shinji
中科院分区：
文献类型：
--
作者：
Fukumitsu, Ken;Ishii, Takamichi;Uemoto, Shinji
小鼠胎肝中的基质细胞在体外具有促进小鼠胚胎干细胞(ESCs)和肝祖细胞(HPC)成熟的能力。分离的基质细胞为CD49f(+/-)CD45(-)Thy1(+)gp38(+)细胞组分。本研究建立了一种小鼠胎肝基质细胞系，可诱导小鼠胚胎干细胞和造血祖细胞的肝脏成熟。将含有温度敏感型SV40大T抗原的转基因载体导入原代胎肝基质细胞。这些永生化细胞被命名为gp38阳性和Thy1阳性的小鼠肝基质细胞(MLSGT)，通过共培养的方法诱导小鼠ESCs和HPC分化为成熟的肝细胞样细胞。由于MLSGT不是克隆细胞，因此选择了一株与其亲本基质细胞相似的、具有诱导肝分化特性的克隆MLSGt20。与MLSGt20细胞共培养的ESC来源的内胚层细胞表达成熟的肝细胞特异性基因标记物，包括葡萄糖-6-磷酸酶、酪氨酸氨基转移酶、色氨酸2，3-二加氧酶和细胞色素P450(细胞色素P450，细胞色素P1a1，细胞色素P1B1，细胞色素P1A2，细胞色素P450)。此外，这些细胞还表现出肝脏功能，如糖原储存和氨代谢。透射电子显微镜显示，共培养的ESCs具有成熟肝细胞的形态特征。综上所述，通过共培养的方法建立了一株具有促进小鼠胚胎干细胞和造血干细胞肝脏成熟特性的细胞系。
Stromal cells residing in murine fetal livers have the ability to promote the hepatic maturation of murine embryonic stem cells (ESCs) and hepatic progenitor cells (HPCs) in vitro. These stromal cells were isolated as the CD49f(+/-) CD45(-) Thy1(+) gp38(+) cell fraction. The present study established a murine fetal liver stromal cell line that induced hepatic maturation in mouse ESCs and HPCs. A transgene containing a temperature-sensitive SV40 large T antigen was transfected into the primary fetal liver stromal cells. These immortalized cells, which were named as the gp38-positive and Thy1-positive murine liver stromal (MLSgt) cells, induced both mouse ESCs and HPCs to differentiate into mature hepatocyte-like cells using a coculture method. Since MLSgt is not a cloned cell line, one clone, MLSgt20, was selected as a line with the characteristic to induce hepatic differentiation, which was comparable to its parental stromal cells. The ESC-derived endoderm cells cocultured with the MLSgt20 cells expressed mature hepatocyte-specific gene markers, including glucose-6-phosphatase, tyrosine aminotransferase, tryptophan 2,3-dioxgenase, and cytochrome P450 (CYP1a1, Cyp1b1, Cyp1a2, and Cyp3a11). In addition, these cells also exhibited hepatic functions, such as glycogen storage and ammonia metabolism. Transmission electron microscopy showed that the cocultured ESCs expressed the morphologic features of mature hepatocytes. In conclusion, a cell line was established that has the characteristic to promote the hepatic maturation of mouse ESCs and HPCs by a coculture method.