Enrichment of hepatocytes differentiated from mouse embryonic stem cells as a transplantable source
Enrichment of hepatocytes differentiated from mouse embryonic stem cells as a transplantable source
复制标题
DOI：
10.1097/01.tp.0000153637.44069.c6
复制
发表时间：
2005-03-15
期刊：
影响因子：
6.2
通讯作者：
Teraoka, H
中科院分区：
文献类型：
--
作者：
Kumashiro, Y;Asahina, K;Teraoka, H
背景。我们以前报道过肝细胞可以通过胚状体（EB）的形成从胚胎干细胞（ES）分化出来，并且可以移植到小鼠肝脏中。然而，eb来源的细胞移植经常导致受体肝脏形成畸胎瘤。在本研究中，我们从EB外生物中去除致瘤细胞，并检测了富集es细胞来源的肝细胞移植到损伤肝脏中的效果。在培养第15天，EBs部分分解并传代培养。通过肝细胞标志物的表达检测传代细胞中的肝细胞。通过Percoll不连续梯度离心去除eb源性细胞中的未分化细胞。此外，使用血小板/内皮细胞粘附分子(PECAM)-1和Mac-1抗体进行磁性细胞分选，消除未分化细胞、内皮细胞和巨噬细胞。然后将这些富集的es细胞来源的肝细胞移植到损伤小鼠的肝脏中。Percoll离心和PECAM-1抗体消除eb源性细胞中表达Oct-3/4的未分化细胞。es细胞衍生的肝细胞在传代培养过程中表现出肝脏相关基因的表达、尿素和糖原的合成以及结构特征。一项移植研究表明，富集的es细胞来源的肝细胞整合到受伤的小鼠肝脏中，没有产生畸胎瘤。将es细胞来源的肝细胞移植到ccl4损伤的肝脏后，肝功能得到改善。小鼠胚胎干细胞可通过EB形成分化为功能性肝细胞。从EBs中清除未分化的细胞为肝衰竭提供了可移植的细胞，而不会致瘤性。
Background. 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.Methods. 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.Results. 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.Conclusions. 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.