Park Yong, Lee Seung Jin, Choi In Young, Lee Se Ryeon, Sung Hwa Jung, Kim Jong Hoon, Yoo Young Do, Geum Dong Ho, Kim Sun Haeng, Kim Byung Soo
Institute of Stem Cell Research, Korea University, Seoul, Korea.
Cell Reprogram. 2010 Jun;12(3):315-28. doi: 10.1089/cell.2009.0113.
The use of a mouse embryonic fibroblast (MEF) feeder for culture of embryonic stem cells (ESCs) is a widely accepted method, regardless of the ESCs' origin and type. In this study, we performed the undifferentiated propagation of human ES cell lines (hESCs, H1, and HSF6) and mouse ES cell lines (mESCs, D3, and CE3), which were previously maintained on an MEF feeder, using human placenta-derived fibroblast-like cell (HPC) feeders originated from chorionic villi of women who had undergone therapeutic abortion due to known maternal disease that is aggravated by pregnancy. Moreover, we tried to introduce the HPC feeder for the establishment of inducible pluripotent stem cells (iPSCs) from human placental mesenchymal stem cells (MSCs). On the HPC feeder we were able to propagate ESCs and iPSCs colonies as an undifferentiated state up to the 50th passage and 20th passage, respectively. Maintenance of undifferentiated ESCs was identified by the expression of ALP, SSEA-1, SSEA-4, TRA-81, TRA-60, Oct-4, Nanog, or Rex-1. Also, addition of leukemia inhibitory factor was not required for undifferentiated propagation of mESCs on the HPC feeder. The efficiency and expression of three germ layer markers of embryoid bodies (EBs) from ESCs were satisfactory in both the MEF and HPC group. EBs formed from iPSCs were scant, and differentiation to the three germ layers was identifiable by reverst transcription-polymerase chain reactio (RT-PCR) only in the HPC group. In conclusion, the HPC feeder can efficiently support the undifferentiated propagation of hESCs, mESCs, and iPSCs, suggesting that human placenta may be a useful source of universal feeder cells for hESC, mESC, and iPSC culture.
使用小鼠胚胎成纤维细胞(MEF)饲养层培养胚胎干细胞(ESC)是一种被广泛接受的方法,无论ESC的来源和类型如何。在本研究中,我们使用源自因已知妊娠会加重的母体疾病而接受治疗性流产的妇女绒毛膜绒毛的人胎盘来源的成纤维样细胞(HPC)饲养层,对先前在MEF饲养层上维持的人胚胎干细胞系(hESC,H1和HSF6)和小鼠胚胎干细胞系(mESC,D3和CE3)进行了未分化增殖。此外,我们尝试引入HPC饲养层,用于从人胎盘间充质干细胞(MSC)建立诱导多能干细胞(iPSC)。在HPC饲养层上,我们能够分别将ESC和iPSC集落维持在未分化状态直至第50代和第20代。通过碱性磷酸酶(ALP)、阶段特异性胚胎抗原-1(SSEA-1)、阶段特异性胚胎抗原-4(SSEA-4)、肿瘤相关抗原81(TRA-81)、肿瘤相关抗原60(TRA-60)、八聚体结合转录因子4(Oct-4)、Nanog或Rex-1的表达来鉴定未分化ESC群体的维持情况。此外,在HPC饲养层上对mESC进行未分化增殖不需要添加白血病抑制因子。在MEF组和HPC组中,ESC来源的胚状体(EB)的三个胚层标志物的效率和表达均令人满意。iPSC形成的EB很少,并且仅在HPC组中通过逆转录-聚合酶链反应(RT-PCR)可鉴定到向三个胚层的分化。总之,HPC饲养层可以有效地支持hESC、mESC和iPSC的未分化增殖,这表明人胎盘可能是用于hESC、mESC和iPSC培养的通用饲养细胞的有用来源。