Reactive oxygen species in bovine oocyte maturation in vitro
Reactive oxygen species in bovine oocyte maturation in vitro
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DOI：
10.1071/rd08198
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发表时间：
2009-01-01
影响因子：
1.9
通讯作者：
Dalvit, Gabriel C.
中科院分区：
文献类型：
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作者：
Morado, Sergio A.;Cetica, Pablo D.;Dalvit, Gabriel C.
活性氧在卵母细胞体外成熟过程中的作用一直存在争议。本研究的目的是确定在牛卵母细胞体外成熟过程中，在不同的ROS产生调节剂的存在下，ROS产生可能的波动。卵丘-卵母细胞复合体在培养基199（对照）中于39 ℃、5%CO_2、潮湿空气中培养22小时，培养基中有或无0.6mm半胱氨酸、1 mm 1-氯-2，4-二硝基苯（CDNB）、2 μ m二苯基碘、0.5mm N-硝基-L-精氨酸甲酯或10 μ m硝普钠（SNP）。此外，还使用了呼吸链效应物氰化钾（KCN; 1 mm）和羰基氰间氯苯腙（0.42 μ m）。减数分裂成熟由MII的存在决定。以2 '，7'-二氯二氢荧光素二醋酸酯（DCHF-DA）与荧光素二醋酸酯（FDA）的比例在不同时间点评价裸露卵母细胞中的活性氧产生。ROS水平，表示为DCHF-DA：FDA，波动整个22小时的成熟取决于所施加的治疗。在KCN和SNP存在下孵育12 h后，ROS水平升高，而在半胱氨酸存在下孵育12 h后，ROS水平降低（P < 0.05）。CDNB和SNP均损害减数分裂进程。牛卵母细胞成熟过程中较高的代谢活性需求与ROS产生的伴随减少相一致。这些结果表明，12 h是牛卵母细胞体外成熟的一个临界点，因为它与此时ROS的产生密切相关。
The role of reactive oxygen species (ROS) in the in vitro maturation (IVM) of oocytes remains controversial. The aim of the present study was to determine possible fluctuations in ROS production during bovine oocyte IVM in the presence of different modulators of ROS generation. Cumulus-oocyte complexes were cultured in medium 199 (control) in the absence or presence of 0.6mm cysteine, 1mm 1-choro-2,4-dinitro benzene (CDNB), 2 mu m diphenyliodonium, 0.5mm N-nitro-L-arginine methyl ester or 10 mu m sodium nitroprusside (SNP) at 39 degrees C, in 5% CO2 in humidified air for 22 h. In addition, the respiratory chain effectors potassium cyanide (KCN; 1mm) and carbonyl cyanide m-chlorophenylhydrazone (0.42 mu m) were used. Meiotic maturation was determined by the presence of MII. ROS production was evaluated in denuded oocytes at different time points as the ratio of 2',7'-dichlorodihydrofluorescein diacetate (DCHF-DA) to fluorescein diacetate (FDA). ROS levels, expressed as DCHF-DA: FDA, fluctuated throughout the 22 h of maturation depending on the treatment applied. At 12 h incubation in the presence of KCN and SNP, ROS levels were increased, whereas ROS levels after 12 h in the presence of cysteine were reduced (P < 0.05). Both CDNB and SNP impaired meiotic progression. The higher metabolic activity demand during bovine oocyte maturation coincides with a concomitant reduction in ROS generation. These results suggest that 12 h would be a critical point for bovine oocyte IVM because it is closely related to the production of ROS at this time.