Effects of different concentrations of amino acids in the culture medium on preimplantation mouse embryo development in vitro

Abstract

Abstract: Objective To evaluate the effects of amino acids (AA) on the development of in vitro cultured preimplantation embryos of Kunming mice, and define the optimal AA concentration for embryo culture.Methods Totally 630 zygotes were collected from the oviducts of superovulated female Kunming mice, which were cultured in protein-free potassium simplex optimized medium (mKSOM) supplemented with Eagle’s essential amino acids and Eagle’s non-essential amino acids of different concentrations (mKSOM, mKSOM+1/16AA, mKSOM+1/8AA, mKSOM+1/4AA, mKSOM+1/2AA, mKSOM+AA, and mKSOM+2AA).Results The embryos cultured with the amino acids showed higher development rate to both 8-cell embryo stage and blastocyst stage than those cultured without amino acids. The correlation of amino acid concentration with 8-cell and blastocyst development rates conformed to the cubic model, with the highest development rate to both of the two stages observed at half of the amino acid concentration.Conclusion Amino acids can promote the development of preimplantation Kunming mouse embryos, but excessively high concentration of amino acids impair embryo development possibly because of metabolic and osmotic pressure changes of the embryos as well as toxicity of ammonium resulting from the metabolism of amino acids.

CLC Number:

Q95-3

DENG Xing¹, WANG Shuang¹, HUANG Wu-jian², LIU Qiu-zhen¹, XIE Wei-bing¹, ZHANG Qing-ling¹, FANG Wei-yi³, LIU Teng-fei³, HAN Chun¹, DU Sha-sha³, WU Li-sha¹, DING Yan-qing¹. Effects of different concentrations of amino acids in the culture medium on preimplantation mouse embryo development in vitro[J]. Journal of Southern Medical University, 2005, 25(03): 241-245.

References

[1] Biggers JD. Reflections on the culture of the preimplantation embryo [J]. Int J Dev Biol,1998;42 (7):879-84.
[2] 钟瑜,潘善培,张春雪,等.人输卵管上皮细胞共培养系统对小鼠胚胎体外发育的影响[J].生殖医学杂志,1996,5(3):162-5.Zhong Y,Pan SP,Zhang CX,et al. Improvement of mouse early embryo development in vitro by co-cultured with human oviductal epithelial cells[J]. J Reprod Med,1996,5(3):162-5.
[3] Taniguchi F,Harada T,Nara M,et al. Coculture with a human granulosa cell line enhanced the development of murine preimplantation embryos via SCF/c-kit system [J]. Assist Reprod Genet,2004,21 (6):223-8.
[4] Chatot CL,Ziomek CA,Bavister BD,et al. An improved culture medium supports development of random-bred 1-cell mouse embryos in vitro[J]. J Reprod Fertil,1989,86(2):679-88.
[5] Lawitts JA,Biggers JD. Culture of preimplantation embryos [J].Methods Enzymol,1993,225:153-64.
[6] Chatot CL,Lewis JL,Torres I,et al. Development of 1-cell embryos from different strains of mice in CZB media[J]. Biol Reprod,1990,42(3):432-40.
[7] Kamata K,Todoriki H,Kusumoto M,et al. Toxicological evaluation of an in vitro culture of mouse embryos:effect of non-carcinogenic mutagens for the development of embryos [J]. Nippon Yakurigaku Zasshi,1986,88(2):167-72.
[8] Lane M,Gardner DK. EDTA stimulates development of cleavage stage mouse embryos by inhibiting the glycolytic enzyme phosophoglycerate kinase[J]. Biol Reprod,1997,57 (Suppl 1):193.
[9] Cholewa JA,Whitten WK. Development of two-cell mouse embryos in the absence of a fixed-nitrogen source [J]. J Reprod Fertil,1970,22(3):553-5.
[10] Biggers JD,Summers MC,McGinnis LK. Polyvinyl alcohol and amino acids as substitutes for bovine serum albumin in mouse preimplantation embryo culture media [J]. Hum Reprod Update,1997,3(2):125-35.
[11] 黄敏珍,陈士岭,邢福祺,单精子注入卵胞浆过程对胚胎发育的影响[J].第一军医大学学报,2001,21(5):363-4Huang MZ,Chen SL,Xing FQ. Impact of intracytoplasmic sperm injection procedure on embryonic development[J]. J First Mil Med Univ/Di Yi Jun Yi Da Xue Xue Bao,2001,21(5):363-4
[12] Schini SA,Bavister BD. Development of golden hamster embryos through the two-cell block in chemically-defined media [J]. J Exp Zool,1988,245(1):111-45.
[13] Miller,JGO,Schultz GA. Amino acid content of preimplantation rabbit embryos and fluids of the reproductive tract [J]. Biol Reprod,1987,36(1):125-9.
[14] Gardner,DK,Leese HJ. Concentrations of nutrients in mouse oviduct fluid and their effects on embryo development and metabolism in vitro[J]. Reprod Fertil,1990,88(1):361-8.
[15] Seshagiri PB,Bavister BD. Glucose and phosphate inhibit respiration and oxidative metabolism in cultured hamster eight-cell embryos:evidence for the ’Crabtree effect’[J]. Mol Reprod Dev,1991,30(2):105-11.
[16] Gardner DK,Lane M. The 2-cell block in CF1 mouse embryos is associated with an increase in glycolysis and a decrease in tricarboxylic acid glycolysis and a decrease in tricarboxylic acid (TCA) cycle activity:alleviation of the 2-cell block is associated with the restoration of in vivo metabolic pathway activities [J]. Biol Reprod,1993,49 (Suppl. 1):52.
[17] Barnett DK,Bavister BD. Inhibitory effect of glucose and phosphate on the second cleavage division of hamster embryos:is it linked to metabolism[J] ? Hum Reprod,1996,11(1):177-83.
[18] Borland RM,Biggers JD,Lechene CP,et al. Elemental composition of fluid in the human Fallopian tube [J]. J Reprod Fertil,1980,58(2):479-82.
[19] van Winkle LJ,Campione AL. Amino acid transport regulation in preimplantation mouse embryos:effects of amino acid content and pre- and peri-implantation development [J]. Theriogenology,1996,45:69-80.
[20] Dumoulin,JCM,van Wissen LCP,Menheere PPCA,et al. Taurine acts as an osmolyte in human and mouse oocytes and embryos [J].Biol Reprod,1997,56(3):739-44.
[21] Edwards LJ,Williams DA,Gardner DK. Intracellular pH of the preimplantation mouse embryo:effects of extracellular pH and weak acids[J]. Mol Reprod Dev. 1998,50(4):434-42.
[22] Edwards LJ,Williams DA,Gardner DK. Intracellular pH of of the mouse preimplantation embryo:amino acids act as buffers of intracellular pH[J]. Hum Reprod,1998,13(12):3441-8
[23] Gardner D.K. and Lane M.,Amino acids and ammonium regulate the development of mouse embryos in culture [J]. Biol Reprod,1993,48(2):377-85.
[24] Lane,M,Gardner DK. Increase in postimplantation development of cultured mouse embryos by amino acids and induction of fetal retardation and exencephaly by ammonium ions [J]. Reprod Fertil,1994,102(2):305-12.
[25] Lane M,Gardner DK. Differential regulation of mouse embryo developments and viability by amino acids[J]. Reprod Fertil,1997,109:153-64.