Efficient and scalable generation of primordial germ cells in 2D culture using basement membrane extract overlay

Overeem, Arend W.; CHANG, Yolanda W.; Moustakas, Ioannis; Roelse, Celine M.; Hillenius, Sanne; Helm, Talia Van Der; Schrier, Valerie F. van der; Gonçalves, Manuel A.F.V.; Mei, Hailiang; Freund, Christian; Lopes, Susana M. Chuva de Sousa

doi:10.1016/j.crmeth.2023.100488

Cited by 7 publications

(11 citation statements)

References 47 publications

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“…The generation of PGCLCs and subsequent in vitro -derived egg and sperm have rightly taken center stage in reproductive biology. Since Hayashi and Saitou's breakthrough generation of mouse PGCLCs and gametes ( Hayashi et al, 2011 , 2012 ), protocols have been developed to improve PGCLC survival and the efficiency of derivation ( Overeem et al, 2023 ; Jo et al, 2022 ; Gell et al, 2020 ). PGCLCs are key to achieving translational goals, such as the preservation of endangered species and reproductive medicine ( Hayashi et al, 2022 ; Wesevich et al, 2023 ).…”

Section: Conclusion Current Challenges and The Next Frontiermentioning

confidence: 99%

The journey of a generation: advances and promises in the study of primordial germ cell migration

Barton,

Roa-de la Cruz,

Lehmann

et al. 2024

Development

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The germline provides the genetic and non-genetic information that passes from one generation to the next. Given this important role in species propagation, egg and sperm precursors, called primordial germ cells (PGCs), are one of the first cell types specified during embryogenesis. In fact, PGCs form well before the bipotential somatic gonad is specified. This common feature of germline development necessitates that PGCs migrate through many tissues to reach the somatic gonad. During their journey, PGCs must respond to select environmental cues while ignoring others in a dynamically developing embryo. The complex multi-tissue, combinatorial nature of PGC migration is an excellent model for understanding how cells navigate complex environments in vivo. Here, we discuss recent findings on the migratory path, the somatic cells that shepherd PGCs, the guidance cues somatic cells provide, and the PGC response to these cues to reach the gonad and establish the germline pool for future generations. We end by discussing the fate of wayward PGCs that fail to reach the gonad in diverse species. Collectively, this field is poised to yield important insights into emerging reproductive technologies.

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Section: Conclusion Current Challenges and The Next Frontiermentioning

confidence: 99%

The journey of a generation: advances and promises in the study of primordial germ cell migration

Barton,

Roa-de la Cruz,

Lehmann

et al. 2024

Development

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“…In humans, it is possible to differentiate human PGCLCs (hPGCLCs) from human PSCs, but the efficiency of these protocols is highly variable depending on the PSC lines used [30][31][32]. Yamashiro and colleagues introduced a differentiation protocol whereby hPGCLCs matured further into meiosis but failed to progress through meiotic recombination [33].…”

Section: Ovarian Organoids To Study Oocyte Formation and Follicular A...mentioning

confidence: 99%

Bioengineered 3D Ovarian Models as Paramount Technology for Female Health Management and Reproduction

Valle

Lopes

2023

Bioengineering

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Ovarian dysfunction poses significant threats to the health of female individuals. Ovarian failure can lead to infertility due to the lack or inefficient production of fertilizable eggs. In addition, the ovary produces hormones, such as estrogen and progesterone, that play crucial roles not only during pregnancy, but also in maintaining cardiovascular, bone, and cognitive health. Decline in estrogen and progesterone production due to ovarian dysfunction can result in menopausal-associated syndromes and lead to conditions, such as osteoporosis, cardiovascular disease, and Alzheimer’s disease. Recent advances in the design of bioengineered three-dimensional (3D) ovarian models, such as ovarian organoids or artificial ovaries, have made it possible to mimic aspects of the cellular heterogeneity and functional characteristics of the ovary in vitro. These novel technologies are emerging as valuable tools for studying ovarian physiology and pathology and may provide alternatives for fertility preservation. Moreover, they may have the potential to restore aspects of ovarian function, improving the quality of life of the (aging) female population. This review focuses on the state of the art of 3D ovarian platforms, including the latest advances modeling female reproduction, female physiology, ovarian cancer, and drug screening.

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“…Additionally, we switched to a basement membrane extract overlay monolayer induction protocol which allowed higher cell seeding densities. 36 In some samples, we also tested the retinoic acid receptor agonist AM580, but we observed that this inhibited cell proliferation, and we did not include in in the final protocol out of concerns that it could induce premature differentiation of oogonia. After this second round of screening, we chose six promising factors and performed a fractional factorial screen, testing 16 combinations in two cell lines.…”

Section: An Improved Protocol Allows Production Of Iolcs With Higher ...mentioning

confidence: 99%

Efficient Human Germ Cell Specification from Stem Cells via Combinatorial Expression of Transcription Factors

Kramme

Smela

Wolf

et al. 2022

Preprint

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Germ cells are the vehicle of human reproduction, arising early in embryonic development and developing throughout adult life until menopause onset in women. Primordial germ cells are the common precursors of germline cells in both sexes, undergoing sexual specification into oogonia or gonocytes which further develop into oocytes or spermatocytes during development. Methods for recapitulation of primordial germ cell and oogonia formation have been developed extensively in recent decades, but fundamental technical limitations in their methodologies, throughput, and yield limit their utilization. Recently, transcription factor (TF)-based methods for human primordial germ cell-like cell (hPGCLC) formation, mouse meiotic entry, and mouse oocyte maturation have demonstrated the feasibility of gene overexpression screening in identifying potent regulators of germ cell development. Here we screened 47 folliculogenesis-regulating TFs for their role in hPGCLC and oogonia formation, identifying DLX5, HHEX, and FIGLA whose individual overexpression enhances hPGCLC formation from hiPSCs. Additionally, we identify a set of three TFs, ZNF281, LHX8, and SOHLH1 whose combinatorial overexpression drives direct oogonia-like formation from hiPSCs in a four-day, feeder-free monolayer culture condition with additional feeder-free culture capabilities post-isolation. We characterize these TF-based germ cells via gene and protein expression analyses, and demonstrate their broad similarity to in vivo germ cells. Together, these results identify novel regulators of human germ cell development and establish new TF-based tools for human in vitro oogenesis research.

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Efficient and scalable generation of primordial germ cells in 2D culture using basement membrane extract overlay

Cited by 7 publications

References 47 publications

The journey of a generation: advances and promises in the study of primordial germ cell migration

The journey of a generation: advances and promises in the study of primordial germ cell migration

Bioengineered 3D Ovarian Models as Paramount Technology for Female Health Management and Reproduction

Efficient Human Germ Cell Specification from Stem Cells via Combinatorial Expression of Transcription Factors

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