Complete Meiosis from Embryonic Stem Cell-Derived Germ Cells In Vitro

Abstract

In vitro generation of functional gametes is a promising approach for treating infertility, although faithful replication of meiosis has proven to be a substantial obstacle to deriving haploid gamete cells in culture. Here we report complete in vitro meiosis from embryonic stem cell (ESC)-derived primordial germ cells (PGCLCs). Co-culture of PGCLCs with neonatal testicular somatic cells and sequential exposure to morphogens and sex hormones reproduced key hallmarks of meiosis, including erasure of genetic imprinting, chromosomal synapsis and recombination, and correct nuclear DNA and chromosomal content in the resulting haploid cells. Intracytoplasmic injection of the resulting spermatid-like cells into oocytes produced viable and fertile offspring, showing that this robust stepwise approach can functionally recapitulate male gametogenesis in vitro. These findings provide a platform for investigating meiotic mechanisms and the potential generation of human haploid spermatids in vitro.

Stem cells, in vitro gametogenesis and male fertility

“…However, the resultant offspring were apparently not healthy, as they were either smaller or larger than controls and showed premature death between 5 days and 5 months after birth. Recently, Zhou and coworkers reported a more robust system, in which haploid round spermatid could be derived from mouse ESCs (Zhou et al 2016). In this culture system, which was based on those in the previous reports (Hayashi et al 2011(Hayashi et al , 2012, PGCLCs were derived from ESCs and then cultured with W/Wv germ cell-deficient testicular cells.…”

“…It is also a concern that the differentiation process is accelerated in all the culture systems. It usually takes about 6 weeks to differentiate mature sperm from PGCs via prospermatogonia in vivo, whereas it requires only 2-3 weeks to progress from PGCLCs to haploid spermatid in culture (Zhou et al 2016), and thus, the latter process is unlikely to recapitulate faithfully gametogenesis in vivo. Similarity between in vivo and in vitro gametogenesis or so-called 'biological reproducibility', seems particularly important, as evidence shows that prospermatogonia undergo dynamic epigenetic rearrangement (Davis et al 2000, Ueda et al 2000, Ly et al 2015 and are likely sensitive to manipulation in culture , Ishikura et al 2016.…”

Male germline stem cells in non-human primates

“…Mouse induced pluripotent stem cells (iPSCs) can be reprogrammed into PGC-like cells. These in vitro generated cells can be further developed into oocyte-like or haploid male gametes with fertilization competence once an in vivo gonadal environment is provided (Zhou et al, 2016;Ishikura et al, 2016).…”

Derivation and application of pluripotent stem cells for regenerative medicine

“…In addition, human oocyte morphology classification and therapeutic cloning by xenotransplanted oocytes (Riaz et al, 2011) were also tried. Recently, fertile offspring from Kit w /Kit wv infertile male mice was generated through in vivo differentiation of gene corrected ntESCs , and spermatid-like cells from mouse ESCs in vitro gametogenesis (Zhou et al, 2016) were derived, exploring new paths to rescue male infertility caused by genetic mutations .…”