Recently, mice have been produced from Freeze-drying (FD) somatic cells by nuclear transfer, but the success rate was very low compared to FD spermatozoa. Since spermatozoa, unlike somatic cells, are haploid cell and their nuclei were hardened by protamine, it is unclear which is responsible for tolerance to FD treatment. Here, we attempt to produce offspring from FD spermatid, a haploid sperm progenitor cell, but nuclei were not yet replaced by protamine as same as somatic cells. We developed the method to collect FD spermatids from testicular suspension. When FD spermatids were injected into oocytes, healthy offspring were obtained, despite the significantly reduced success rate compared to FD spermatozoa. Offspring were also obtained from FD spermatids derived from immature male mice, which have not yet produced spermatozoa. These results indicate that nuclear protaminization, rather than haploid nuclei, is the key process responsible for tolerance to FD treatment.

Whether mammalian embryos develop normally under microgravity remains to be determined. However, embryos are too small to be handled by inexperienced astronauts who orbit Earth on the International Space Station (ISS). Here we describe the development of a new device that allows astronauts to thaw and culture frozen mouse 2-cell embryos on the ISS without directly contacting the embryos. First, we developed several new devices using a hollow fiber tube that allows thawing embryo without practice and observations of embryonic development. However, the general vitrification method requires liquid nitrogen, which is not available on the ISS. Therefore, we developed another new device, Embryo Thawing and Culturing unit (ETC) employing a high osmolarity vitrification method, which preserves frozen embryos at −80°C for several months. Embryos flushed out of the ETC during thawing and washing were protected using a mesh sheet. Although embryonic development could not be observed in the ETC, thawed embryos formed blastocysts after 4 days of culture without direct contact. This ETC will enable untrained astronauts to thaw and culture frozen embryos on the ISS, as well as to serve as an important advance in fields such as clinical infertility and animal biotechnology.