Derivation of Primordial Germ Cells from Human Embryonic and Induced Pluripotent Stem Cells Is Significantly Improved by Coculture with Human Fetal Gonadal Cells
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Open Access
- 22 January 2009
- journal article
- research article
- Published by Oxford University Press (OUP) in The International Journal of Cell Cloning
- Vol. 27 (4), 783-795
- https://doi.org/10.1002/stem.13
Abstract
The derivation of germ cells from human embryonic stem cells (hESCs) or human induced pluripotent stem (hIPS) cells represents a desirable experimental model and potential strategy for treating infertility. In the current study, we developed a triple biomarker assay for identifying and isolating human primordial germ cells (PGCs) by first evaluating human PGC formation during the first trimester in vivo. Next, we applied this technology to characterizing in vitro derived PGCs (iPGCs) from pluripotent cells. Our results show that codifferentiation of hESCs on human fetal gonadal stromal cells significantly improves the efficiency of generating iPGCs. Furthermore, the efficiency was comparable between various pluripotent cell lines regardless of origin from the inner cell mass of human blastocysts (hESCs), or reprogramming of human skin fibroblasts (hIPS). To better characterize the iPGCs, we performed Real-time polymerase chain reaction, microarray, and bisulfite sequencing. Our results show that iPGCs at day 7 of differentiation are transcriptionally distinct from the somatic cells, expressing genes associated with pluripotency and germ cell development while repressing genes associated with somatic differentiation (specifically multiple HOX genes). Using bisulfite sequencing, we show that iPGCs initiate imprint erasure from differentially methylated imprinted regions by day 7 of differentiation. However, iPGCs derived from hIPS cells do not initiate imprint erasure as efficiently. In conclusion, our results indicate that triple positive iPGCs derived from pluripotent cells differentiated on hFGS cells correspond to committed first trimester germ cells (before 9 weeks) that have initiated the process of imprint erasure. Disclosure of potential conflicts of interest is found at the end of this article.Keywords
Funding Information
- UCLA Department of Molecular Cell and Developmental Biology
- STOP Cancer Foundation
- NIH (P01 GM081621-01A1)
This publication has 46 references indexed in Scilit:
- The promoter of the oocyte-specific gene, Gdf9, is active in population of cultured mouse embryonic stem cells with an oocyte-like phenotypeMethods, 2008
- Generation of human induced pluripotent stem cells from dermal fibroblastsProceedings of the National Academy of Sciences of the United States of America, 2008
- Proximal visceral endoderm and extraembryonic ectoderm regulate the formation of primordial germ cell precursorsBMC Developmental Biology, 2007
- Gene-specific vulnerability to imprinting variability in human embryonic stem cell linesGenome Research, 2007
- Status of genomic imprinting in human embryonic stem cells as revealed by a large cohort of independently derived and maintained linesHuman Molecular Genetics, 2007
- Bone Morphogenetic Proteins Induce Germ Cell Differentiation from Human Embryonic Stem CellsStem Cells and Development, 2006
- Testicular Cell Conditioned Medium Supports Differentiation of Embryonic Stem Cells into Ovarian Structures Containing OocytesThe International Journal of Cell Cloning, 2005
- BMP signaling mediated by ALK2 in the visceral endoderm is necessary for the generation of primordial germ cells in the mouse embryoGenes & Development, 2004
- Derivation of Oocytes from Mouse Embryonic Stem CellsScience, 2003
- Structural Evidence of Functional Divergence in Human Alkaline PhosphatasesOnline Journal of Public Health Informatics, 2002