Human in vitro spermatogenesis as a regenerative therapy — where do we stand?

Spermatogenesis involves precise temporal and spatial gene expression and cell signalling to reach a coordinated balance between self-renewal and differentiation of spermatogonial stem cells through various germ cell states including mitosis, and meiosis I and II, which result in the generation of haploid cells with a unique genetic identity. Subsequently, these round spermatids undergo a series of morphological changes to shed excess cytoplast, develop a midpiece and tail, and undergo DNA repackaging to eventually form millions of spermatozoa. The goal of recreating this process in vitro has been pursued since the 1920s as a tool to treat male factor infertility in patients with azoospermia. Continued advances in reproductive bioengineering led to successful generation of mature, functional sperm in mice and, in the past 3 years, in humans. Multiple approaches to study human in vitro spermatogenesis have been proposed, but technical and ethical obstacles have limited the ability to complete spermiogenesis, and further work is needed to establish a robust culture system for clinical application. Several approaches have been used to date to reproduce human spermatogenesis in vitro, which, however, mostly result in incomplete differentiation. In this Review, the authors describe advantages and limitations of current technologies for in vitro spermatogenesis. New approaches that could lead to the clinical translation of this technology to treat patients with male factor infertility are also discussed. Key points Spermatogenesis includes a complex and highly regulated series of steps required for the production of mature, motile sperm. Efforts to recreate this process in vitro are ongoing to support the treatment of male factor infertility, particularly non-obstructive azoospermia (NOA). Advances in the ex vivo production of mature spermatids in animal models have provided new insights for the successful achievement of these results in humans. Several approaches to achieve human in vitro spermatogenesis have been described, such as 2D and 3D cultures including scaffold, organoid-based and bioprinted systems, which are supported by a variety of media and biomaterials. Complete spermatid differentiation in vitro has been shown in some studies, although this process frequently occurred at an accelerated rate compared with the expected course in vivo; moreover, these results still need to be independently replicated by other research groups. The future perspective