Human primordial germ cell-like cells specified from resetting precursors develop in human hindgut organoids.


Type
Article
Change log
Authors
Alves-Lopes, João Pedro  ORCID logo  https://orcid.org/0000-0003-3042-6988
Wong, Frederick CK 
Surani, M Azim 
Abstract

Human primordial germ cells (hPGCs), the precursors of eggs and sperm, start their complex development shortly after specification and during their migration to the primitive gonads. Here, we describe protocols for specifying hPGC-like cells (hPGCLCs) from resetting precursors and progressing them with the support of human hindgut organoids. Resetting hPGCLCs (rhPGCLCs) are specified from human embryonic stem cells (hESCs) transitioning from the primed into the naive state of pluripotency. Hindgut organoids are also derived from hESCs after a sequential differentiation into a posterior endoderm/hindgut fate. Both rhPGCLCs and hindgut organoids are combined and co-cultured for 25 d. The entire procedure takes ~1.5 months and can be successfully implemented by a doctoral or graduate student with basic skills and experience in hESC cultures. The co-culture system supports the progression of rhPGCLCs at a developmental timing analogous to that observed in vivo. Compared with previously developed hPGCLC progression protocols, which depend on co-cultures with mouse embryonic gonadal tissue, our co-culture system represents a developmentally relevant model closer to the environment that hPGCs first encounter after specification. Together with the potential for investigations of events during hPGC specification and early development, these protocols provide a practical approach to designing efficient models for in vitro gametogenesis. Notably, the rhPGCLC-hindgut co-culture system can also be adapted to study failings in hPGC migration, which are associated with the etiology of some forms of infertility and germ cell tumors.

Description
Keywords
3206 Medical Biotechnology, 32 Biomedical and Clinical Sciences, Stem Cell Research - Embryonic - Human, Regenerative Medicine, Stem Cell Research, 1 Underpinning research, 1.1 Normal biological development and functioning
Journal Title
Nat Protoc
Conference Name
Journal ISSN
1754-2189
1750-2799
Volume Title
Publisher
Springer Science and Business Media LLC
Rights
Publisher's own licence
Sponsorship
Wellcome Trust (209475/Z/17/Z)
Wellcome Trust (096738/Z/11/Z)
Medical Research Council (MR/P009948/1)
BBSRC (BB/T01346X/1)
European Commission Horizon 2020 (H2020) Marie Sk?odowska-Curie actions (836291)
Swedish Research Council (2019-06316) Swedish Research Council (2022-01467)