Jang Goo, Lee Byeong Chun, Kang Sung Keun, Hwang Woo Suk
Department of Theriogenology and Biotechnology, College of Veterinary Medicine, Seoul National University, Seoul, Korea.
Reprod Fertil Dev. 2003;15(3):179-85. doi: 10.1071/rd02054.
The purpose of this study was to evaluate the effect of glycosaminoglycans (GAGs) added to the culture medium on the developmental competence of bovine embryos derived from in vitro fertilization (IVF) and from somatic cell nuclear transfer (SCNT). In vitro-matured oocytes were either inseminated with 1 x 10(6) spermatozoa mL(-1) or enucleated and reconstructed with bovine adult ear fibroblasts by SCNT. The embryos were then cultured in modified synthetic oviduct fluid (mSOF) containing 8 mg mL(-1) bovine serum albumin (BSA) (control mSOF) or control mSOF supplemented with various GAGs (hyaluronic acid, heparin or chondroitin sulfate) in a dose-dependent manner (0.1, 0.5 or 1.0 mg mL(-1)). Developmental competence was evaluated by monitoring the numbers of 2-cell embryos, 8-16-cell embryos and blastocysts. The mean cell number of flattened blastocysts stained with 5 microM bisbenzimide on Day 8 was counted. The percentage of blastocyst formation (IVF and SCNT embryos) from cleaved embryos was significantly higher (P < 0.05) in control mSOF supplemented with 0.5 mg mL(-1) hyaluronic acid (45% and 47%), heparin (40% and 47%) or chondroitin sulfate (38% and 44%) compared with control mSOF (30-31% and 30-33%). When compared with the efficacy of 0.5 mg mL(-1) GAGs, no significant differences were observed in the developmental competence of both IVF and SCNT embryos. Supplementing control mSOF with 0.5 mg mL(-1) GAGs had no effect on the cell number of IVF embryos. In contrast, supplementing 0.5 mg mL(-1) of hyaluronic acid, heparin or chondroitin sulfate to control mSOF significantly (P < 0.05) increased the numbers of total cells (93-98 v. 88 cells) and trophectoderm (TE) cells (64-66 v. 55 cells), and decreased the inner cell mass (ICM) to TE cell ratio (48.2-49.8 v. 61.3) in SCNT blastocysts compared with embryos in control mSOF. In conclusion, supplementation of culture media with GAGs may improve the development of bovine IVM-IVF and SCNT embryos to the blastocyst stage. The GAGs increased the quality of blastocysts by increasing total cell numbers in the SCNT embryos.
本研究的目的是评估添加到培养基中的糖胺聚糖（GAGs）对体外受精（IVF）和体细胞核移植（SCNT）获得的牛胚胎发育能力的影响。体外成熟的卵母细胞要么用1×10⁶个精子/mL进行授精，要么通过SCNT去核并与牛成年耳成纤维细胞进行重构。然后将胚胎培养在含有8mg/mL牛血清白蛋白（BSA）的改良合成输卵管液（mSOF）中（对照mSOF），或者是添加了不同剂量（0.1、0.5或1.0mg/mL）GAGs（透明质酸、肝素或硫酸软骨素）的对照mSOF中。通过监测2细胞胚胎、8 - 16细胞胚胎和囊胚的数量来评估发育能力。对第8天用5μM双苯甲酰胺染色的扁平囊胚的平均细胞数进行计数。与对照mSOF（30 - 31%和30 - 33%）相比，添加0.5mg/mL透明质酸（45%和47%）、肝素（40%和47%）或硫酸软骨素（38%和44%）的对照mSOF中，来自分裂胚胎的囊胚形成率（IVF和SCNT胚胎）显著更高（P < 0.05）。与0.5mg/mL GAGs的效果相比，IVF和SCNT胚胎的发育能力未观察到显著差异。向对照mSOF中添加0.5mg/mL GAGs对IVF胚胎的细胞数没有影响。相反，与对照mSOF中的胚胎相比，向对照mSOF中添加0.5mg/mL的透明质酸、肝素或硫酸软骨素显著（P < 0.05）增加了SCNT囊胚的总细胞数（93 - 98对88个细胞）和滋养外胚层（TE）细胞数（64 - 66对55个细胞），并降低了内细胞团（ICM）与TE细胞的比例（48.2 - 49.8对61.3）。总之，向培养基中添加GAGs可能会改善牛体外成熟 - IVF和SCNT胚胎发育到囊胚阶段的情况。GAGs通过增加SCNT胚胎中的总细胞数提高了囊胚的质量。
Zotero 插件