Kobe Research Laboratories, Eisai Co., Ltd. 6-8- 2 Minatozima Minamimachi, Chuo-ku, Kobe, 650-0047, Japan.
Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Chuo-ku, Kumamoto, 860- 0811, Japan.
Sci Rep. 2024 Oct 15;14(1):24198. doi: 10.1038/s41598-024-76073-x.
The cryopreservation of rat embryos is useful for efficiently archiving rat resources in bioresource repositories. The cryopreserved fertilized oocytes can be quickly reanimated to rats with homozygous mutations using embryo transfer. In addition, cryopreserved rat fertilized oocytes are easier to transport than live animals. Before cryopreservation, fertilized oocytes are obtained by mating or in vitro fertilization. However, it is not clear which fertilized oocytes are most suited to cryopreservation. In this study, we performed a systematic comparison of the fertilizing ability, cryotolerance, and developmental ability of cryopreserved fertilized oocytes at the pronuclear stage produced either by mating (in vivo) or in vitro fertilization (in vitro) in SD and F344 rats. In vivo-fertilized oocytes had higher cryotolerance and developmental ability than in vitro-fertilized oocytes in SD and F344 rats. Furthermore, the fertilization ability, cryotolerance, and developmental ability of vitrified-warmed fertilized oocytes differed between SD and F344 rats. In conclusion, our study suggests that in vivo-fertilized rat oocytes were more suitable for cryopreservation. Our protocol provides an optimized system for the management of rat colonies using fertilized oocytes cryopreservation and contributes to the 3Rs principle by reducing the number of animals used for research.
胚胎的冷冻保存对于在生物资源库中有效地保存大鼠资源非常有用。通过胚胎移植，可以将冷冻保存的同源突变受精卵迅速复苏为纯合突变大鼠。此外，与活体动物相比，冷冻保存的大鼠受精卵更容易运输。在冷冻保存之前，可以通过交配或体外受精获得受精卵。然而，目前尚不清楚哪种受精卵最适合冷冻保存。在这项研究中，我们在 SD 和 F344 大鼠中对通过交配（体内）或体外受精（体外）产生的原核期冷冻受精卵的受精能力、冷冻耐受性和发育能力进行了系统比较。体内受精的受精卵在 SD 和 F344 大鼠中的冷冻耐受性和发育能力均高于体外受精的受精卵。此外，冷冻解冻的受精卵在 SD 和 F344 大鼠之间的受精能力、冷冻耐受性和发育能力也存在差异。总之，我们的研究表明，体内受精的大鼠受精卵更适合冷冻保存。我们的方案提供了一个优化的系统，用于使用受精卵冷冻保存来管理大鼠群体，通过减少用于研究的动物数量，符合 3R 原则。
