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  • In vitro reconstitution of the whole male germ-cell development from mouse pluripotent stem cells.

In vitro reconstitution of the whole male germ-cell development from mouse pluripotent stem cells.

Cell stem cell (2021-09-09)
Yukiko Ishikura, Hiroshi Ohta, Takuya Sato, Yusuke Murase, Yukihiro Yabuta, Yoji Kojima, Chika Yamashiro, Tomonori Nakamura, Takuya Yamamoto, Takehiko Ogawa, Mitinori Saitou
ABSTRACT

Mammalian male germ-cell development consists of three distinct phases: primordial germ cell (PGC) development, male germ-cell specification for spermatogonium development, and ensuing spermatogenesis. Here, we show an in vitro reconstitution of whole male germ-cell development by pluripotent stem cells (PSCs). Mouse embryonic stem cells (mESCs) are induced into PGC-like cells (mPGCLCs), which are expanded for epigenetic reprogramming. In reconstituted testes under an optimized condition, such mPGCLCs differentiate into spermatogonium-like cells with proper developmental transitions, gene expression, and cell-cycle dynamics and are expanded robustly as germline stem cell-like cells (GSCLCs) with an appropriate androgenetic epigenome. Importantly, GSCLCs show vigorous spermatogenesis, not only upon transplantation into testes in vivo but also under an in vitro culture of testis transplants, and the resultant spermatids contribute to fertile offspring. By uniting faithful recapitulations of the three phases of male germ-cell development, our study creates a paradigm for the in vitro male gametogenesis by PSCs.

MATERIALS
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Bovine Serum Albumin, cold ethanol fraction, pH 5.2, ≥96%
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Cyclosporin A, 97.0-101.5% (on dried basis)
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Mitomycin C from Streptomyces caespitosus, powder, BioReagent, suitable for cell culture
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Deoxyribonuclease I from bovine pancreas, lyophilized powder, Protein ≥85 %, ≥400 Kunitz units/mg protein
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ESGRO® Recombinant Mouse LIF Protein, ESGRO Leukemia Inhibitory Factor (LIF) supplement for mouse ES cell culture. Each vial contains 10^7 units/ml.
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