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Recently, complete in vitro generation of male gametes starting from pluripotent stem cells was obtained in a mouse model with live offspring as a result. This breakthrough was probably due to the use of a stepwise differentiation protocol taking the tightly regulated in vivo situation into account. As shown previously, factors of the TGFβ superfamily, metabolites of vitamin A, growth hormones, sex steroids and, most importantly, somatic cell support are major regulators of the development, survival, proliferation and differentiation of male gametes. However, up till now, all differentiation protocols starting from human pluripotent stem cells only focused on one or two of these substantive factors, not taking any timeframe into account, leading to promising but unsatisfying results with low efficiency. Therefore, progress might be achieved by including a stepwise differentiation protocol, including all proven contributing regulators, and therefore mimicking more closely human in vivo spermatogenesis and its temporo-spatial organization. In this review, the indispensable regulators of in vivo spermatogenesis and the outcomes of related human in vitro studies are discussed with the aim of unravelling the most successful combinations of medium factors to be used in future differentiation protocols.

Human in vitro spermatogenesis from pluripotent stem cells: in need of a stepwise differentiation protocol?