Embryonic stem cells can form germ cells in vitro

Abstract

Knock-in embryonic stem (ES) cells, in which GFP or lacZ was expressed from the endogenous mouse vasa homolog (Mvh), which is specifically expressed in differentiating germ cells, were used to visualize germ cell production during in vitro differentiation. The appearance of MVH-positive germ cells depended on embryoid body formation and was greatly enhanced by the inductive effects of bone morphogenic protein 4-producing cells. The ES-derived MVH-positive cells could participate in spermatogenesis when transplanted into reconstituted testicular tubules, demonstrating that ES cells can produce functional germ cells in vitro. In vitro germ cell differentiation provides a paradigm for studying the molecular basis of germ line establishment, as well as for developing new approaches to reproductive engineering. G erm-line cells are responsible for transmitting genetic information and for reproducing totipotency from generation to generation. Pluripotent stem cell lines, embryonic stem (ES) cells, and embryonic germ cells are established from cells of the germ cell lineage. Therefore, germ cell specification must be linked to the maintenance of pluripotency, as well as to cell fate commitment leading to gametogenesis.Unlike many animal species, in which the germ line is predetermined by maternal factors, germ cell specification in mammals takes place at the onset of gastrulation, after implantation of the embryo. In the mouse, primordial germ cells (PGCs) are first distinguished at the base of the allantois in gastrulating embryos at embryonic day (E) 7.25 (1). Lineage studies of epiblast cells show that mouse PGCs are specified by inductive interactions at the onset of gastrulation (2, 3). Genetic analyses using targeted mutations have revealed that bone morphogenic protein (BMP) 4 and -8b, soluble growth factors belonging to the transforming growth factor ␤ superfamily that are produced by extraembryonic ectoderm close to the boundary with the proximal epiblast, are required for the generation of PGCs from epiblast cells (4,5). Moreover, primary cultures of epiblast fragments from embryos at E5.5-E6.0 generate migrating PGCs when they are cocultured with extraembryonic ectoderm (6), and culturing of whole epiblasts from E6.0 embryos on feeder cells expressing both BMP4 and BMP8b gives rise to PGCs (7). These results reveal that BMPs derived from the extraembryonic ectoderm play crucial roles in PGC determination in the proximal epiblast.Despite such developments, it is not yet known how the founder population of PGCs is segregated from other pluripotent epiblast cells that form somatic cells. To approach this question, we examined the production of germ cells by an established pluripotent ES cell line. ES cells can form all cell lineages when introduced into host blastocysts and give rise to various somatic cell lineages in culture. However, it is not known whether they can generate the germ cell lineage in culture in the absence of the morphogenetic events associated with gastrulation. It has been difficult to addr...

In vitro production of fertile sperm from murine spermatogonial stem cell lines

“…In this regard, germ cells or primordial germ cells reported or claimed to have been derived from ES/iPS cells and epiblast cells [34][35][36][37] could be tested regarding their spermatogenic activity with our in vitro system. This particular application of our method would break the boundary between germ and somatic cells, and open a new phase in germ cell research.…”

Amniotic fluid stem cell-based models to study the effects of gene mutations and toxicants on male germ cell formation

“…The efficiency of differentiation from ES or iPS cells into haploid male germ cells is generally reported to be low. [14][15][16][17][18][19][20][21][22][23][24] Although there is no evidence for the speculation that this efficiency might be higher using AFS cells, to our opinion, the here reviewed knowledge warrants testing this new stem cell type for their usability for the in vitro investigation of sperm production (Figure 1).…”

“…Since spontaneous induction of meiosis is low, overexpression of genes of the DAZ gene family can be performed to induce meiosis for haploid spermatids 5,27 ( Figure 1). Finally, in mice it is possible to test the biological function of so-obtained haploid male gametes either by transplantation into busulfan-mediated sperm-depleted testes to check in vivo differentiation, or by intracytoplasmic injection into recipient oocytes to test whether this supports cleavage [14][15][16] (Figure 1).…”

Can we grow sperm? A translational perspective on the current animal and human spermatogenesis models

“…Toyooka and colleagues 53 have reported finding mature germ cells from their cultured ESCs injected into a mouse testis. In a landmark paper, Geijsen et al 54 reported the derivation of mouse PGCs by adding retinoic acid to an ESC culture media.…”