Generating human primordial germ cell-like cells from pluripotent stem cells: a scoping review of in vitro methods

STUDY QUESTION: How do the methods and outcomes of established protocols to specify human primordial germ cell-like cells (hPGCLCs) in vitro compare to each other? SUMMARY ANSWER: All analyzed protocols were successful in generating hPGCLCs, and a few were able to induce further germ cell maturation. WHAT IS KNOWN ALREADY: There are a variety of protocols for generating hPGCLCs in vitro, each with its own advantages and disadvantages. To date no comparison has been made, hindering the practical application of in vitro-derived hPGCLCs in research and the advancement toward generating mature germ cells. STUDY DESIGN, SIZE, DURATION: For this scoping review, a systematic search for protocols was conducted in the databases Scopus and Web of Science, including publications since 2010. Search terms included human, differentiation/specification/induction, germ cell/oogonia/spermatogonia, and primordial. PARTICIPANTS/MATERIALS, SETTING, METHODS: Two separate authors performed the database search according to the inclusion/exclusion criteria. The data regarding the materials and methods as well as results of the included articles were extracted and organized based on protocol (cell type and culture system) and outcome. MAIN RESULTS AND THE ROLE OF CHANCE: A systematic search revealed 32 articles describing the generation of hPGCLCs. Of these, 24 articles contained an original hPGCLC differentiation protocol and 8 articles provided an extension of a previously published protocol. The extension protocols focused either on extending hPGCLC culture or maturing hPGCLCs further. The articles were compared regarding protocol methods and differentiation outcomes. The data showed that differentiation in 2D or 3D, in the presence of bone morphogenetic protein 4 (BMP4) (or retinoic acid), activated the WNT and NODAL signaling pathways to induce hPGCLCs. Further maturation (based on gene expression) was also achieved, depending on the inclusion of subsequent differentiation steps. The 2D culture systems showed high efficiency and scalability, while the 3D culture systems were more suitable for germ cell maturation purposes. Further improvements would include a deeper assessment of epigenetic and gene expression, functional analyses, and use of multiple cell lines to reflect protocol versatility. LIMITATIONS, REASONS FOR CAUTION: Only literature has been compared; no extensive experimental comparison or a meta-analysis was performed due to the heterogeneity in outcome measurements. WIDER IMPLICATIONS OF THE FINDINGS: This review offers a comparison of hPGCLC differentiation protocols and aims to aid researchers in selecting appropriate protocols and making informed modifications to the culture conditions to achieve efficient germ cell differentiation.