Abstract
The ability to generate germ cells from pluripotent stem cells (PSCs) is valuable for human regenerative medicine and animal breeding. Germ cell-like cells (GCLCs) have been differentiated from mouse and human PSCs, but not from porcine PSCs, which are considered an ideal model for stem cell applications. Here, we developed a defined culture system for the induction of primordial germ cell-like cells (PGCLCs) from porcine induced PSCs (piPSCs). The identity of the PGCLCs was characterized by observing cell morphology, detecting germ cell marker gene expression and evaluating epigenetic properties. PGCLCs could further differentiate into spermatogonial stem cell-like cells (SSCLCs) in vitro. Importantly, meiosis occurred during SSCLC induction. Xenotransplantation of GCLCs into seminiferous tubules of infertile immunodeficient mice resulted in immunohistochemically identifiable germ cells in vivo. Overall, our study provides a feasible strategy for directing piPSCs to the germ cell fate and lays a foundation for exploring germ cell development mechanisms.
Highlights
The germ cell lineage, which is the source of totipotency, is capable of differentiating into all types of cells
After approximately 15 days of culture in the neurogenesis condition, porcine induced PSCs (piPSCs) had differentiated into NESTIN+ neural progenitor cells, TUJ1+ neurons, or GFAP+ astrocytes, which were detected by immunofluorescence staining (Fig. 1g)
Mouse primordial germ celllike cells (PGCLCs) are induced via an epiblast stem cell-like cell (EpiLC) state[1], while human PGCLCs are derived through a mesoderm-like cell population[13,14]
Summary
The germ cell lineage, which is the source of totipotency, is capable of differentiating into all types of cells. Human PSCs can be induced into SSCLCs, enter meiosis, and differentiate into haploid spermatogenic cells in vitro by overexpressing Dazl and other related genes or adding growth factors such as RA to the culture systems[15,16,17,18]. Transplantation of human iPSCs directly into mouse seminiferous tubules, which provide a germ cell niche, can direct the germ cell differentiation in vivo[19]. PGCLCs were induced from piPSCs and further differentiated into SSCLCs. Interestingly, the induced PGCLCs proliferated and developed for more than 6 weeks in vivo and exhibited germ cell features after injection into the seminiferous tubules of immunodeficient mice that lacked endogenous germ cells
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