Generation of marmoset primordial germ cell-like cells under chemically defined conditions

Primordial germ cells (PGCs) are the embryonic precursors of sperm and oocytes and are responsible for the transmission of genetic and epigenetic information across generations. The development of mouse gametes from PGCs can be reconstituted from pluripotent stem cells in vitro, opening up new avenues for the study of germ cell biology in biomedical research. However, the development of germ cells exhibits molecular and morphological differences between rodents and humans. Therefore, to establish an in vitro model system that is closely related to humans, we studied PGC development in vivo and in vitro in the common marmoset Callithrix jacchus (cj), a New World monkey. Post-migratory cjPGCs in gonads at embryonic day 74 express the germ cell-related genes SOX17, AP2?, BLIMP1 and the pluripotency markers NANOG and OCT4A (POU5F1), which is reminiscent of human PGCs. We established transgene-free induced pluripotent stem cell (cjiPSC) lines from foetal and postnatal fibroblasts. These cjiPSCs, cultured in a defined medium and in feeder-free conditions, can be differentiated into precursors of mesendoderm and subsequently into cjPGC-like cells (cjPGCLCs) with a transcriptome similar to human PGCs/PGCLCs. Our results not only pave the way to study PGC development in a non-human primate in vitro under experimentally controlled conditions without foetal calf serum and feeder cells, but also provide the opportunity to derive functional marmoset gametes in vitro in future studies. Overall design: Two biological replicates of each cell type analyzed: iPSCs cultured in different media (TESR and cjPSCM), Precursor of mesendoderm induced with or without Activin A, CXCR4&INTalpha6 double-positive and double-negative cells from day 4 embryoid bodies