Lee Yongjin, Shim Joohyun, Ko Nayoung, Kim Hyoung-Joo, Park Jae-Kyung, Kwak Kyungmin, Kim Hyunil, Choi Kimyung
Optipharm Inc, 63, Osongsaengmyeong 6-ro, Osong-eup, Heungdeok-gu, Cheongju-si, Chungcheongbuk-do, 28158, Republic of Korea.
Optipharm Inc, 63, Osongsaengmyeong 6-ro, Osong-eup, Heungdeok-gu, Cheongju-si, Chungcheongbuk-do, 28158, Republic of Korea.
Theriogenology. 2019 Mar 15;127:80-87. doi: 10.1016/j.theriogenology.2019.01.001. Epub 2019 Jan 4.
The objective of this study was to examine the effect of alanine treatment during in vitro maturation (IVM) on oocyte maturation and embryonic development in pigs. To this end, we investigated the nuclear maturation, intraoocyte glutathione (GSH) content of IVM oocytes, and embryonic development after parthenogenesis (PA) and somatic cell nuclear transfer (SCNT). In addition, we analyzed the expression of genes associated with apoptosis and embryonic development in IVM oocytes, 4-cell stage embryos, and blastocysts produced via PA and SCNT. To determine the optimal concentration of alanine to promote the maturation and development of PA and SCNT embryos, various concentrations (0, 0.363, 1, 5, and 10 mM) of alanine were added to IVM medium during oocyte maturation. The proportion of metaphase II (MII) oocytes after IVM did not differ according to the concentration of alanine. However, significantly higher intraoocyte GSH content was observed in oocytes treated with 0.363 mM alanine compared with that in untreated oocytes. However, treatment of recipient oocytes with 5 or 10 mM alanine during IVM decreased the GSH content in mature oocytes compared to that in control oocytes. Oocytes matured in the presence of 0.363 mM alanine showed significantly increased rates of cleavage and blastocyst formation after PA and SCNT compared to untreated oocytes. PA and SCNT embryos from the 0.363 mM alanine-treated group of MII oocytes showed significantly higher transcript levels of POU5F1 and FGFR2, which are associated with oocyte quality and embryonic development, than the untreated group. Our results suggest that treatment of pig oocytes with 0.363 mM alanine during IVM improves embryonic developmental competence after PA and SCNT by increasing intraoocyte GSH content and increasing the mRNA expression of POU5F1 and FGFR2.
本研究的目的是检测体外成熟（IVM）过程中丙氨酸处理对猪卵母细胞成熟和胚胎发育的影响。为此，我们研究了IVM卵母细胞的核成熟、卵母细胞内谷胱甘肽（GSH）含量，以及孤雌生殖（PA）和体细胞核移植（SCNT）后的胚胎发育情况。此外，我们分析了通过PA和SCNT产生的IVM卵母细胞、4细胞期胚胎和囊胚中与细胞凋亡和胚胎发育相关基因的表达。为了确定促进PA和SCNT胚胎成熟和发育的丙氨酸最佳浓度，在卵母细胞成熟过程中向IVM培养基中添加了不同浓度（0、0.363、1、5和10 mM）的丙氨酸。IVM后中期II（MII）卵母细胞的比例根据丙氨酸浓度的不同没有差异。然而，与未处理的卵母细胞相比，用0.363 mM丙氨酸处理的卵母细胞中观察到卵母细胞内GSH含量显著更高。然而，与对照卵母细胞相比，在IVM期间用5或10 mM丙氨酸处理受体卵母细胞会降低成熟卵母细胞中的GSH含量。与未处理的卵母细胞相比，在0.363 mM丙氨酸存在下成熟的卵母细胞在PA和SCNT后显示出显著提高的分裂率和囊胚形成率。来自0.363 mM丙氨酸处理组的MII卵母细胞的PA和SCNT胚胎显示出与未处理组相比显著更高的POU5F1和FGFR2转录水平，这与卵母细胞质量和胚胎发育相关。我们的结果表明，在IVM期间用0.363 mM丙氨酸处理猪卵母细胞可通过增加卵母细胞内GSH含量以及增加POU5F1和FGFR2的mRNA表达来提高PA和SCNT后的胚胎发育能力。
