Paracrine actions of growth differentiation factor-9 in the mammalian ovary
Paracrine actions of growth differentiation factor-9 in the mammalian ovary
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DOI：
10.1210/me.13.6.1035
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发表时间：
1999-06-01
影响因子：
--
通讯作者：
Matzuk, MM
中科院分区：
文献类型：
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作者：
Elvin, JA;Clark, AT;Matzuk, MM
尽管转化生长因子-β (TGF-β) 超家族是最大的分泌性生长因子家族，但令人惊讶的是，其信号通路中的下游靶基因却很少被发现。同样，调节重要的卵母细胞-体细胞相互作用的卵母细胞衍生的分泌因子的身份仍然很大程度上未知。例如，已知卵母细胞分泌旁分泌生长因子，其对于卵丘扩张、诱导透明质酸合成和抑制LH受体(LHR) mRNA合成是必需的。我们之前的研究表明，TGF-β 家族成员生长分化因子-9 (GDF-9) 的缺失会阻碍初级卵泡阶段的卵巢卵泡发生，从而导致不孕。在本研究中，我们证明小鼠 GDF-9 蛋白在从 3a 型卵泡阶段（包括窦卵泡）开始的所有卵母细胞中表达。为了探索 GDF-9 在卵泡发生和卵丘扩张后期的生物学功能，我们使用中国仓鼠卵巢细胞表达系统生产了成熟的糖基化重组小鼠 GDF-9。建立颗粒细胞培养系统，利用半定量 RT-PCR 确定 GDF-9 在调节几个关键卵巢基因产物中的作用。我们发现重组 GDF-9 诱导透明质酸合酶 2 (HAS2)、环氧合酶 2 (COX-2) 和类固醇生成急性调节蛋白 (StAR) mRNA 合成，但抑制尿激酶纤溶酶原激活剂 (uPA) 和 LHR mRNA 合成。与 GDF-9 对 StAR mRNA 的诱导一致，重组 GDF-9 在缺乏 FSH 的情况下增加了颗粒细胞孕酮合成。由于卵丘细胞中 HASP 的诱导和蛋白酶 uPA 的抑制是卵丘扩张过程中产生的富含透明质酸的细胞外基质的关键事件，因此我们确定 GDF-9 是否可以模拟该过程。使用卵母细胞切除的卵丘细胞-卵母细胞复合物，我们证明重组 GDF-9 在体外诱导卵丘扩张。这些研究表明，GDF-9 可以与颗粒细胞上的受体结合，调节多种基因产物的表达。因此，除了在早期卵泡发生过程中发挥生长和分化因子的关键作用外，GDF-9还作为卵母细胞分泌的旁分泌因子，调节参与卵丘扩张和维持最佳卵母细胞微环境的几种关键颗粒细胞酶，这些过程对于正常排卵、受精和女性生殖至关重要。
Although the transforming growth factor-beta (TGF-beta) superfamily is the largest family of secreted growth factors, surprisingly few downstream target genes in their signaling pathways have been identified. Likewise, the identities of oocyte-derived secreted factors, which regulate important oocyte-somatic cell interactions, remain largely unknown. For example, oocytes are known to secrete paracrine growth factor(s) which are necessary for cumulus expansion, induction of hyaluronic acid synthesis, and suppression of LH receptor (LHR) mRNA synthesis. Our previous studies demonstrated that absence of the TGF-beta family member, growth differentiation factor-9 (GDF-9), blocks ovarian folliculogenesis at the primary follicle stage leading to infertility. In the present study, we demonstrate that mouse GDF-9 protein is expressed in all oocytes beginning at the type 3a follicle stage including antral follicles. To explore the biological functions of GDF-9 in the later stages of folliculogenesis and cumulus expansion, we produced mature, glycosylated, recombinant mouse GDF-9 using a Chinese hamster ovary cell expression system. A granulosa cell culture system was established to determine the role of GDF-9 in the regulation of several key ovarian gene products using semiquantitative RT-PCR. We find that recombinant GDF-9 induces hyaluronan synthase 2 (HAS2), cyclooxygenase 2 (COX-2), and steroidogenic acute regulator protein (StAR) mRNA synthesis but suppresses urokinase plasminogen activator (uPA) and LHR mRNA synthesis. Consistent with the induction of StAR mRNA by GDF-9, recombinant GDF-9 increases granulosa cell progesterone synthesis in the absence of FSH. Since induction of HASP and suppression of the protease uPA in cumulus cells are key events in the production of the hyaluronic acid-rich extracellular matrix which is produced during cumulus expansion, we determined whether GDF-9 could mimic this process. Using oocytectomized cumulus cell-oocyte complexes, we show that recombinant GDF-9 induces cumulus expansion in vitro. These studies demonstrate that GDF-9 can bind to receptors on granulosa cells to regulate the expression of a number of gene products. Thus, in addition to playing a critical function as a growth and differentiation factor during early folliculogenesis, GDF-9 functions as an oocyte-secreted paracrine factor to regulate several key granulosa cell enzymes involved in cumulus expansion and maintenance of an optimal oocyte microenvironment, processes which are essential for normal ovulation, fertilization, and female reproduction.