Cyclic nucleotide phosphodiesterase 3A-deficient mice as a model of female infertility
Cyclic nucleotide phosphodiesterase 3A-deficient mice as a model of female infertility
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DOI：
10.1172/jci2004210804
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发表时间：
2004-07-01
影响因子：
15.9
通讯作者：
Manganiello, V
中科院分区：
文献类型：
--
作者：
Masciarelli, S;Horner, K;Manganiello, V
由于cAMP在体外阻碍哺乳动物和两栖动物卵母细胞的减数分裂成熟，并且环核苷酸磷酸二酯酶3A（PDE3A）主要负责卵母细胞cAMP水解，因此我们通过同源重组产生了PDE3A缺陷小鼠。 Pde3a(-/-)雌性存活并排卵正常数量的卵母细胞，但完全不育，因为排卵的卵母细胞在生发囊泡阶段停滞，因此无法受精。 Pde3a(-/-) 卵母细胞缺乏 cAMP 特异性 PDE 活性，cAMP 水平升高，并且无法在体外自发成熟（长达 48 小时）。通过用腺苷-3',5'-环硫代磷酸酯、Rp-异构体 (Rp-cAMPS) 抑制蛋白激酶 A (PKA) 或注射蛋白激酶抑制肽 (PKI) 或编码磷酸酶 CDC25 的 mRNA，可恢复 Pde3a(-/-) 卵母细胞减数分裂的成熟，这证实 cAMP-PKA 信号传导的增加是减数分裂阻断的原因。经历生泡破裂的Pde3a(-/-)卵母细胞表现出MPF和MAPK的激活，完成第一次减数分裂，挤出极体，并具有精子受精能力。我们相信这些发现提供了第一个遗传证据，表明体内和体外减数分裂的恢复需要 PDE3A 活性。 Pde3a(-/-) 小鼠代表减数分裂成熟和排卵分离的体内模型，这强调了抑制卵母细胞成熟作为避孕的潜在策略。
Since cAMP blocks meiotic maturation of mammalian and amphibian oocytes in vitro and cyclic nucleotide phosphodiesterase 3A (PDE3A) is primarily responsible for oocyte cAMP hydrolysis, we generated PDE3A-deficient mice by homologous recombination. The Pde3a(-/-) females were viable and ovulated a normal number of oocytes but were completely infertile, because ovulated oocytes were arrested at the germinal vesicle stage and, therefore, could not be fertilized. Pde3a(-/-) oocytes lacked cAMP-specific PDE activity, contained increased cAMP levels, and failed to undergo spontaneous maturation in vitro (up to 48 hours). Meiotic maturation in Pde3a(-/-) oocytes was restored by inhibiting protein kinase A (PKA) with adenosine-3',5'-cyclic monophosphorothioate, Rp-isomer (Rp-cAMPS) or by injection of protein kinase inhibitor peptide (PKI) or mRNA coding for phosphatase CDC25, which confirms that increased cAMP-PKA signaling is responsible for the meiotic blockade. Pde3a(-/-) oocytes that underwent germinal vesicle breakdown showed activation of MPF and MAPK, completed the first meiotic division extruding a polar body, and became competent for fertilization by spermatozoa. We believe that these findings provide the first genetic evidence indicating that resumption of meiosis in vivo and in vitro requires PDE3A activity. Pde3a(-/-) mice represent an in vivo model where meiotic maturation and ovulation are dissociated, which underscores inhibition of oocyte maturation as a potential strategy for contraception.