Molecular characterization of the follicle defects in the growth differentiation factor 9-deficient ovary.
Molecular characterization of the follicle defects in the growth differentiation factor 9-deficient ovary.
复制标题
DOI：
10.1210/mend.13.6.0309
复制
发表时间：
1999-06
影响因子：
--
通讯作者：
J. Elvin;Changning Yan;Pei Wang;K. Nishimori;M. Matzuk
中科院分区：
文献类型：
--
作者：
J. Elvin;Changning Yan;Pei Wang;K. Nishimori;M. Matzuk
生长分化因子-9（GDF-9）是转化生长因子-β超家族的分泌型成员，在哺乳动物卵母细胞中从3a型初级卵泡阶段开始以高水平表达。我们以前已经证明，GDF-9缺陷的雌性小鼠是不育的，因为在3b型初级卵泡阶段的卵泡发生的早期阻滞。为了解决由于缺乏GDF-9而导致的分子缺陷，我们分析了几个重要的卵巢标志物基因的表达。我们的研究的主要发现如下：1）对于几种卵泡膜细胞层标记物[即17 α-羟化酶、LH受体（LHR）和c-kit（kit配体的受体）]，在GDF-9缺陷型卵泡周围没有可检测的信号。这表明在不存在GDF-9的情况下，卵泡不能发出募集卵泡膜细胞前体以包围卵泡的信号; 2）GDF-9缺陷型小鼠的初级卵泡显示kit配体和白细胞介素-α的上调。这表明，这两个重要的分泌的生长因子，表达在颗粒细胞，可能直接调节GDF-9的旁分泌方式。通过卵母细胞上的c-kit信号传导，kit配体的上调可能直接涉及GDF-9缺陷型卵母细胞的大小增加和卵母细胞的最终死亡; 3）在卵母细胞损失后，GDF-9缺陷型卵泡的细胞保持在类固醇生成簇中，其在组织学上类似于小黄体。然而，在分子水平上，这些细胞对黄体标志物（例如LHR和P-450侧链裂解）和非黄体标志物（例如白介素α和P-450芳香酶）均呈阳性。这表明最初卵母细胞的存在阻止了黄体化标记物的表达，但是在早期时间点GDF-9的缺乏改变了颗粒细胞的分化程序;增殖细胞核抗原（PCNA）或Ki-67染色及TUNEL法证实通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling）标记，GDF-9缺陷型3b初级卵泡的颗粒细胞不能增殖，但也不能经历细胞死亡。这表明，3b型卵泡的颗粒细胞需要GDF-9的持续生长，也成为有能力进行凋亡，可能通过分化事件。因此，这些研究启发了我们的旁分泌作用的GDF-9，以及正常的步骤，颗粒细胞和卵泡膜细胞的生长和分化的卵泡内。
Growth differentiation factor-9 (GDF-9), a secreted member of the transforming growth factor-beta superfamily, is expressed at high levels in the mammalian oocyte beginning at the type 3a primary follicle stage. We have previously demonstrated that GDF-9-deficient female mice are infertile because of an early block in folliculogenesis at the type 3b primary follicle stage. To address the molecular defects that result from the absence of GDF-9, we have analyzed the expression of several important ovarian marker genes. The major findings of our studies are as follows: 1) There are no detectable signals around GDF-9-deficient follicles for several theca cell layer markers [i.e. 17alpha-hydroxylase, LH receptor (LHR), and c-kit, the receptor for kit ligand]. This demonstrates that in the absence of GDF-9, the follicles are incompetent to emit a signal that recruits theca cell precursors to surround the follicle; 2) The primary follicles of GDF-9-deficient mice demonstrate an up-regulation of kit ligand and inhibin-alpha. This suggests that these two important secreted growth factors, expressed in the granulosa cells, may be directly regulated in a paracrine fashion by GDF-9. Up-regulation of kit ligand, via signaling through c-kit on the oocyte, may be directly involved in the increased size of GDF-9-deficient oocytes and the eventual demise of the oocyte; 3) After loss of the oocyte, the cells of the GDF-9-deficient follicles remain in a steroidogenic cluster that histologically resembles small corpora lutea. However, at the molecular level, these cells are positive for both luteal markers (e.g. LHR and P-450 side chain cleavage) and nonluteal markers (e.g. inhibin alpha and P-450 aromatase). This demonstrates that initially the presence of the oocyte prevents the expression of luteinized markers, but that the absence of GDF-9 at an early timepoint alters the differentiation program of the granulosa cells; and 4) As demonstrated by staining with either proliferating cell nuclear antigen (PCNA) or Ki-67 and TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling) labeling, the granulosa cells of GDF-9-deficient type 3b primary follicles fail to proliferate but also fail to undergo cell death. This suggests that granulosa cells of type 3b follicles require GDF-9 for continued growth and also to become competent to undergo apoptosis, possibly through a differentiation event Thus, these studies have enlightened us as to the paracrine roles of GDF-9 as well as the normal steps of granulosa cell and theca cell growth and differentiation within ovarian follicles.