Sato E, Miyamoto H, Koide S S
Department of Animal Science, Faculty of Agriculture, Kyoto University, Japan.
Mol Reprod Dev. 1990 Aug;26(4):391-7. doi: 10.1002/mrd.1080260415.
The viability of oocytes cultured in vitro was determined by the trypan blue exclusion test. Isolated porcine oocytes with or without cumulus cells cultured in modified Krebs-Ringer medium undergo cell death after 48 h. The addition of glycosaminoglycans (GAGs) prepared from porcine follicular fluid (pFF) to the medium delayed or prevented the onset of cell death in vitro. GAGs at concentrations of 0.25 mg/ml or greater prevented cell death in a dose-dependent manner. To identify the active factor, GAGs were purified from pFF by ethanol precipitation, chromatography on Dowex 1-x2, and high performance liquid chromatography (HPLC) on TSK gel DEAE-2 SW column. The fraction with a retention time nearly coincident with that of hyaluronic acid possessed high oocyte viability promoting activity. The present results suggest that the viability of oocytes in vitro is influenced by the presence of specific GAGs separated from follicular fluid.
通过台盼蓝排斥试验来确定体外培养的卵母细胞的活力。在改良的Krebs-Ringer培养基中培养的带有或不带有卵丘细胞的分离猪卵母细胞在48小时后会发生细胞死亡。向培养基中添加从猪卵泡液（pFF）中制备的糖胺聚糖（GAGs）可延迟或防止体外细胞死亡的发生。浓度为0.25mg/ml或更高的GAGs以剂量依赖的方式防止细胞死亡。为了鉴定活性因子，通过乙醇沉淀、在Dowex 1-x2上进行色谱分离以及在TSK凝胶DEAE-2 SW柱上进行高效液相色谱（HPLC）从pFF中纯化GAGs。保留时间与透明质酸几乎一致的馏分具有高的促进卵母细胞活力的活性。目前的结果表明，体外卵母细胞的活力受从卵泡液中分离出的特定GAGs的存在影响。
