Possible Role of 5′AMP-Activated Protein Kinase in the Metformin-Mediated Arrest of Bovine Oocytes at the Germinal Vesicle Stage During In Vitro Maturation1
Possible Role of 5′AMP-Activated Protein Kinase in the Metformin-Mediated Arrest of Bovine Oocytes at the Germinal Vesicle Stage During In Vitro Maturation1
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DOI：
10.1095/biolreprod.107.060848
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发表时间：
2007-09
期刊：
影响因子：
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通讯作者：
L. Tosca;S. Uzbekova;C. Chabrolle;J. Dupont
中科院分区：
文献类型：
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作者：
L. Tosca;S. Uzbekova;C. Chabrolle;J. Dupont
摘要5′-腺苷酸活化蛋白激酶（AMPK）的激活参与了小鼠和猪卵巢卵母细胞减数分裂成熟过程。然而，其对卵母细胞的影响似乎是物种特异性的。我们研究了AMPK和丝裂原活化蛋白激酶（MAPK 3/1）在牛体外成熟（IVM）过程中的磷酸化模式，以及AMPK激活剂二甲双胍对卵丘-卵母细胞复合体（COCs）和裸露牛卵母细胞（DO）中卵母细胞成熟的影响。在牛COCs中，PRKAA Thr 172磷酸化降低，而MAPK 3/1磷酸化在卵母细胞和卵丘细胞中增加。二甲双胍（5和10 mM）可将COC中的卵母细胞阻滞在GV期，但在DO中则不然。在COCs中，这种停滞与卵丘细胞扩增的抑制、PRKAA Thr 172磷酸化的增加以及卵母细胞和卵丘细胞中MAPK 3/1磷酸化的减少有关。然而，添加AMPK抑制剂化合物C（10 μM）加速了牛COCs卵母细胞GV分解（GVBD）过程的启动，而没有任何MAPK 3/1磷酸化的改变。二甲双胍降低卵母细胞中AURKA和CCNB 1蛋白水平。此外，IVM 1小时后，二甲双胍降低RPS 6磷酸化，增加EEF 2磷酸化，表明二甲双胍处理COCs的卵母细胞蛋白质合成速率较低。用MEK抑制剂U 0126（100微摩尔）处理COCs后，大多数卵母细胞在GVBD阶段后被阻滞。因此，在牛COC中，二甲双胍在GV期阻断减数分裂进程，激活PRKAA，并在IVM期间抑制卵母细胞和卵丘细胞中的MAPK 3/1磷酸化。此外，卵丘细胞对于二甲双胍对牛卵母细胞成熟的影响至关重要，而MAPK 3/1磷酸化则不然。
Abstract The 5′AMP-activated protein kinase (AMPK) activation is involved in the meiotic maturation of oocytes in the ovaries of mice and pigs. However, its effects on the oocyte appear to be species-specific. We investigated the patterns of AMPK and mitogen-activated protein kinases (MAPK3/1) phosphorylation during bovine in vitro maturation (IVM) and the effects of metformin, an AMPK activator, on oocyte maturation in cumulus-oocyte complexes (COCs) and denuded bovine oocytes (DOs). In bovine COCs, PRKAA Thr172 phosphorylation decreased, whereas MAPK3/1 phosphorylation increased in both oocytes and cumulus cells during IVM. Metformin (5 and 10 mM) arrested oocytes at the GV stage in COCs but not in DOs. In COCs, this arrest was associated with the inhibition of cumulus cell expansion, an increase in PRKAA Thr172 phosphorylation, and a decrease in MAPK3/1 phosphorylation in both oocytes and cumulus cells. However, the addition of compound C (10 μM), an inhibitor of AMPK, accelerated the initiation of the GV breakdown (GVBD) process without any alteration of MAPK3/1 phosphorylation in oocytes from bovine COCs. Metformin decreased AURKA and CCNB1 protein levels in oocytes. Moreover, after 1 h of IVM, metformin decreased RPS6 phosphorylation and increased EEF2 phosphorylation, suggesting that protein synthesis rates were lower in oocytes from metformin-treated COCs. Most oocytes were arrested after the GVBD stage following the treatment of COCs with the MEK inhibitor, U0126 (100 micromoles). Thus, in bovine COCs, metformin blocks meiotic progression at the GV stage, activates PRKAA, and inhibits MAPK3/1 phosphorylation in both the oocytes and cumulus cells during IVM. Moreover, cumulus cells were essential for the effects of metformin on bovine oocyte maturation, whereas MAPK3/1 phosphorylation was not.