Sex-Specific Isolation and Propagation of Human Premeiotic Fetal Germ Cells and Germ Cell-Like Cells

Overview

Journal Cells

Publisher MDPI

Specialties Biochemistry
Biophysics
Cell Biology
Molecular Biology

Date 2021 Jun 2

PMID 34065661

Citations 6

Authors

Swati Mishra

Jasin Taelman

Yolanda W Chang

Annekatrien Boel

Petra De Sutter

Bjorn Heindryckx

Susana M Chuva de Sousa Lopes

Affiliations

Soon will be listed here.

Abstract

The second trimester of human development is marked by asynchronous gonadal development hampering the isolation of homogenous populations of early and late fetal germ cells (FGCs). We evaluated the feasibility of using surface markers TNAP, PDPN, EPCAM and ITGA6 to isolate FGCs as well as human primordial germ cell-like cells (hPGCLCs) derived from embryonic stem cells (hESCs) from both sexes by fluorescence-activated cell sorting (FACS). Our results suggest that a combination of TNAP and PDPN was sufficient to separate populations of premeiotic FGCs and hPGCLCs in both sexes. This combination of antibodies also proved efficient in separating 'mitotic' from 'retinoic-acid responsive' female FGCs. Furthermore, we report that the differentiation efficiency of TNAP+PDPN+ hPGCLCs from hESCs was sex-independent, but the ability to propagate differed considerably between the sexes. In contrast to male, female hPGCLCs retained their characteristics and exhibited robust colony-forming ability when cultured for five days in medium containing LIF, forskolin and FGF2. We conclude that marked sex differences exist in the isolation and propagation of human FGCs and hPGCLCs. Our study provides novel insights relevant for the optimization of in vitro gametogenesis in humans.

Citing Articles

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References

Gell J, Liu W, Sosa E, Chialastri A, Hancock G, Tao Y . An Extended Culture System that Supports Human Primordial Germ Cell-like Cell Survival and Initiation of DNA Methylation Erasure. Stem Cell Reports. 2020; 14(3):433-446. PMC: 7066331. DOI: 10.1016/j.stemcr.2020.01.009. View

Li L, Yang R, Yin C, Kee K . Studying human reproductive biology through single-cell analysis and in vitro differentiation of stem cells into germ cell-like cells. Hum Reprod Update. 2020; 26(5):670-688. DOI: 10.1093/humupd/dmaa021. View

Anderson R, Fulton N, Cowan G, Coutts S, Saunders P . Conserved and divergent patterns of expression of DAZL, VASA and OCT4 in the germ cells of the human fetal ovary and testis. BMC Dev Biol. 2007; 7:136. PMC: 2211489. DOI: 10.1186/1471-213X-7-136. View

Sybirna A, Tang W, Pierson Smela M, Dietmann S, Gruhn W, Brosh R . A critical role of PRDM14 in human primordial germ cell fate revealed by inducible degrons. Nat Commun. 2020; 11(1):1282. PMC: 7062732. DOI: 10.1038/s41467-020-15042-0. View

Li L, Dong J, Yan L, Yong J, Liu X, Hu Y . Single-Cell RNA-Seq Analysis Maps Development of Human Germline Cells and Gonadal Niche Interactions. Cell Stem Cell. 2017; 20(6):858-873.e4. DOI: 10.1016/j.stem.2017.03.007. View

Gomes Fernandes M, Bialecka M, Salvatori D, Chuva de Sousa Lopes S . Characterization of migratory primordial germ cells in the aorta-gonad-mesonephros of a 4.5-week-old human embryo: a toolbox to evaluate in vitro early gametogenesis. Mol Hum Reprod. 2018; 24(5):233-243. PMC: 6018722. DOI: 10.1093/molehr/gay011. View

Mishra S, Taelman J, Popovic M, Tilleman L, Duthoo E, van der Jeught M . Activin A-derived human embryonic stem cells show increased competence to differentiate into primordial germ cell-like cells. Stem Cells. 2021; 39(5):551-563. PMC: 8248136. DOI: 10.1002/stem.3335. View

Stoop H, Honecker F, Cools M, de Krijger R, Bokemeyer C, Looijenga L . Differentiation and development of human female germ cells during prenatal gonadogenesis: an immunohistochemical study. Hum Reprod. 2005; 20(6):1466-76. DOI: 10.1093/humrep/deh800. View

Popovic M, Bialecka M, Gomes Fernandes M, Taelman J, van der Jeught M, De Sutter P . Human blastocyst outgrowths recapitulate primordial germ cell specification events. Mol Hum Reprod. 2019; 25(9):519-526. PMC: 6802404. DOI: 10.1093/molehr/gaz035. View

10.

Bialecka M, Young T, Chuva de Sousa Lopes S, Ten Berge D, Sanders A, Beck F . Cdx2 contributes to the expansion of the early primordial germ cell population in the mouse. Dev Biol. 2012; 371(2):227-34. DOI: 10.1016/j.ydbio.2012.08.018. View

11.

Mitsunaga S, Shioda T . Evolutionarily diverse mechanisms of germline specification among mammals: what about us?. Stem Cell Investig. 2018; 5:12. PMC: 5945857. DOI: 10.21037/sci.2018.04.03. View

12.

Sasaki K, Yokobayashi S, Nakamura T, Okamoto I, Yabuta Y, Kurimoto K . Robust In Vitro Induction of Human Germ Cell Fate from Pluripotent Stem Cells. Cell Stem Cell. 2015; 17(2):178-94. DOI: 10.1016/j.stem.2015.06.014. View

13.

Jorgensen A, Rajpert-De Meyts E . Regulation of meiotic entry and gonadal sex differentiation in the human: normal and disrupted signaling. Biomol Concepts. 2014; 5(4):331-41. DOI: 10.1515/bmc-2014-0014. View

14.

Greber B, Lehrach H, Adjaye J . Fibroblast growth factor 2 modulates transforming growth factor beta signaling in mouse embryonic fibroblasts and human ESCs (hESCs) to support hESC self-renewal. Stem Cells. 2006; 25(2):455-64. DOI: 10.1634/stemcells.2006-0476. View

15.

Gkountela S, Li Z, Vincent J, Zhang K, Chen A, Pellegrini M . The ontogeny of cKIT+ human primordial germ cells proves to be a resource for human germ line reprogramming, imprint erasure and in vitro differentiation. Nat Cell Biol. 2012; 15(1):113-22. PMC: 3786872. DOI: 10.1038/ncb2638. View

16.

Padgett D, Cathro H, Wick M, Mills S . Podoplanin is a better immunohistochemical marker for sarcomatoid mesothelioma than calretinin. Am J Surg Pathol. 2007; 32(1):123-7. DOI: 10.1097/PAS.0b013e31814faacf. View

17.

Irie N, Weinberger L, Tang W, Kobayashi T, Viukov S, Manor Y . SOX17 is a critical specifier of human primordial germ cell fate. Cell. 2014; 160(1-2):253-68. PMC: 4310934. DOI: 10.1016/j.cell.2014.12.013. View

18.

Kojima Y, Sasaki K, Yokobayashi S, Sakai Y, Nakamura T, Yabuta Y . Evolutionarily Distinctive Transcriptional and Signaling Programs Drive Human Germ Cell Lineage Specification from Pluripotent Stem Cells. Cell Stem Cell. 2017; 21(4):517-532.e5. DOI: 10.1016/j.stem.2017.09.005. View

19.

Shamblott M, Axelman J, Wang S, Bugg E, LITTLEFIELD J, Donovan P . Derivation of pluripotent stem cells from cultured human primordial germ cells. Proc Natl Acad Sci U S A. 1998; 95(23):13726-31. PMC: 24887. DOI: 10.1073/pnas.95.23.13726. View

20.

Kurilo L . Oogenesis in antenatal development in man. Hum Genet. 1981; 57(1):86-92. DOI: 10.1007/BF00271175. View