Lin Qingyi, Le Quynh Anh, Takebayashi Koki, Hirata Maki, Tanihara Fuminori, Thongkittidilok Chommanart, Sawamoto Osamu, Kikuchi Takeshi, Otoi Takeshige
Bio-Innovation Research Center, Tokushima University, Tokushima, Japan.
Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima, Japan.
Reprod Domest Anim. 2022 May;57(5):556-563. doi: 10.1111/rda.14095. Epub 2022 Feb 13.
This study developed an efficient method for liquid storage of in vitro-derived porcine blastocysts at ambient temperature for 24 hr. We evaluated the effects of new chemically defined media (cell wash and preservation solution, Cellstor -W [Cell-W] and cell suspension and preservation solution, Cellstor -S [Cell-S]) for short-term storage. In the first experiment, in vitro-derived blastocyst were stored at 25ºC for 24 hr in Cell-W solution, Cell-S solution and pig embryo culture (PBM) medium. There were no differences in the rates of survival and development of stored blastocysts between the Cell-S and Cell-W solutions, but the total cell number of embryos that survived after storage in Cell-S solution was significantly higher (p < .05) than that in the Cell-W solution. In the second experiment, Cell-S solution was used to store the in vitro-derived blastocysts at 20°C, 25°C and 30°C. Storage at 20°C resulted in a significant decrease in the rates of survival and development of stored blastocysts compared to storage at 25°C or 30°C. No differences in survival and development rates were observed between storage at 25°C and 30°C, but the damage to the embryo quality after storage and culture was significantly lower at 25°C than at 30°C. In the third experiment, Cell-S solution was supplemented with β-mercaptoethanol and curcumin, either alone or in combination, as antioxidant agents. Although the supplementation with curcumin did not improve survival, it significantly increased the development rate of stored blastocysts compared with the control blastocysts stored without antioxidants. In conclusion, when porcine blastocysts were stored at 25°C for 24 hr, a Cell-S solution may be effective for maintaining the survival and development of in vitro embryos.
本研究开发了一种在常温下将体外培养的猪囊胚液体保存24小时的有效方法。我们评估了新型化学成分明确的培养基(细胞冲洗和保存液,Cellstor -W [Cell-W]以及细胞悬浮和保存液,Cellstor -S [Cell-S])用于短期保存的效果。在第一个实验中,将体外培养的囊胚在Cell-W溶液、Cell-S溶液和猪胚胎培养基(PBM)中于25℃保存24小时。Cell-S溶液和Cell-W溶液保存的囊胚在存活率和发育率上没有差异,但在Cell-S溶液中保存后存活胚胎的总细胞数显著高于(p < 0.05)Cell-W溶液中的胚胎。在第二个实验中,使用Cell-S溶液在20℃、25℃和30℃下保存体外培养的囊胚。与在25℃或30℃下保存相比,在20℃下保存导致保存的囊胚存活率和发育率显著降低。在25℃和30℃下保存的存活率和发育率没有差异,但在25℃下保存和培养后对胚胎质量的损害明显低于30℃。在第三个实验中,向Cell-S溶液中单独或联合添加β-巯基乙醇和姜黄素作为抗氧化剂。虽然添加姜黄素没有提高存活率,但与未添加抗氧化剂保存的对照囊胚相比,它显著提高了保存囊胚的发育率。总之,当猪囊胚在25℃保存24小时时,Cell-S溶液可能对维持体外胚胎的存活和发育有效。