Exclusive generation of rat spermatozoa in sterile mice utilizing blastocyst complementation with pluripotent stem cells

Blastocyst complementation denotes a technique that aims to generate organs, tissues, or cell types in animal chimeras via injection of pluripotent stem cells (PSCs) into genetically compromised blastocyst-stage embryos. Here, we report on successful complementation of the male germline in adult chimeras following injection of mouse or rat PSCs into mouse blastocysts carrying a mutation in Tsc22d3, an essential gene for spermatozoa production. Injection of mouse PSCs into Tsc22d3-Knockout (KO) blastocysts gave rise to intraspecies chimeras exclusively embodying PSC-derived functional spermatozoa. In addition, injection of rat embryonic stem cells (rESCs) into Tsc22d3-KO embryos produced interspecies mouse-rat chimeras solely harboring rat spermatids and spermatozoa capable of fertilizing oocytes. Furthermore, using single-cell RNA sequencing, we deconstructed rat spermatogenesis occurring in a mouse-rat chimera testis. Collectively, this study details a method for exclusive xenogeneic germ cell production in vivo, with implications that may extend to rat transgenesis, or endangered animal species conservation efforts.

Automated summary

This study successfully achieved complementation of functional spermatozoa in adult chimeras through the technique of blastocyst complementation. Using mouse and rat pluripotent stem cells, the researchers were able to generate chimeras with either species-specific sperm or interspecies sperm capable of fertilization. Single-cell RNA sequencing was used to study spermatogenesis in the chimeric testis. The findings of this study have important implications for germ cell research and conservation efforts for endangered animal species.