Sasseville Maxime, Albuz Firas K, Côté Nancy, Guillemette Christine, Gilchrist Robert B, Richard François J
Département des Sciences Animales, Centre de Recherche en Biologie de la Reproduction, Université Laval, Sainte-Foy, Québec, Canada.
Biol Reprod. 2009 Aug;81(2):415-25. doi: 10.1095/biolreprod.108.074450. Epub 2009 Apr 8.
The phosphodiesterase (PDE) family is a group of enzymes that catalyzes the transformation of cyclic nucleotides into 5' nucleotides. Based on rodents, the current mammalian model of PDE distribution in the ovarian follicle predicts Pde3a in the oocyte and Pde4d in the somatic cells. Using bovine as an experimental model, the present results showed that PDE3 was the predominant PDE activity in oocytes. However, cumulus cell cAMP-PDE activity was predominantly resistant to inhibition by 3-isobutyl-methylxantine, indicating PDE8 activity (60% of total PDE activity) and a minor role for PDE4 (<5%). A total of 20% of total oocyte PDE activity was also attributed to PDE8. The PDE activity measurements in mural granulosa cells from 2 to 6 mm in diameter suggest the presence of PDE4 and PDE8. In granulosa cells from follicles >10 mm, total PDE and PDE8 activities along with PDE8A protein level were increased compared with smaller follicles. The RT-PCR experiments showed that cumulus cells expressed PDE8A, PDE8B, and PDE10A. Western blot experiments showed PDE8A, PDE8B, and PDE4D proteins in mural granulosa cells and cumulus-oocyte complexes. PDE8 inhibition using dipyridamole in a dose-dependent manner increased cAMP levels in the cumulus-oocyte complexes and delayed oocyte nuclear maturation. These results are the first to demonstrate the functional presence of PDE8 in the mammalian ovarian follicle. This challenges the recently described cell-specific expression of cAMP-PDEs in the ovarian follicle and the notion that PDE4 is the predominant granulosa/cumulus cell PDE. These findings have implications for our understanding of hormonal regulation of folliculogenesis and the potential application of PDE inhibitors as novel contraceptives.
磷酸二酯酶（PDE）家族是一类催化环核苷酸转化为5'核苷酸的酶。基于啮齿动物，目前关于PDE在卵巢卵泡中分布的哺乳动物模型预测卵母细胞中有Pde3a，体细胞中有Pde4d。以牛作为实验模型，目前的结果表明PDE3是卵母细胞中主要的PDE活性。然而，卵丘细胞的cAMP-PDE活性主要对3-异丁基-甲基黄嘌呤的抑制具有抗性，表明存在PDE8活性（占总PDE活性的60%），而PDE4的作用较小（<5%）。卵母细胞总PDE活性的20%也归因于PDE8。对直径2至6毫米的壁颗粒细胞进行的PDE活性测量表明存在PDE4和PDE8。在直径大于10毫米的卵泡的颗粒细胞中，与较小卵泡相比，总PDE和PDE8活性以及PDE8A蛋白水平均有所增加。RT-PCR实验表明卵丘细胞表达PDE8A、PDE8B和PDE10A。蛋白质印迹实验表明壁颗粒细胞和卵丘-卵母细胞复合体中存在PDE8A、PDE8B和PDE4D蛋白。使用双嘧达莫以剂量依赖性方式抑制PDE8可提高卵丘-卵母细胞复合体中的cAMP水平，并延迟卵母细胞核成熟。这些结果首次证明了PDE8在哺乳动物卵巢卵泡中的功能性存在。这对最近描述的卵巢卵泡中cAMP-PDE的细胞特异性表达以及PDE4是颗粒/卵丘细胞主要PDE的观点提出了挑战。这些发现对于我们理解卵泡发生的激素调节以及PDE抑制剂作为新型避孕药的潜在应用具有重要意义。
