FOLLICULAR MORPHOGENESIS DURING PERINATAL DEVELOPMENT

围产期发育期间的卵泡形态发生

• 批准号: 6740919
• 负责人: SHYAMAL K. ROY
• 金额: $ 25.73万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6740919
• 关键词: epidermal growth factor; follicle stimulating hormone; growth factor receptors; hamsters; hormone regulation /control mechanism; immunofluorescence technique; newborn animals; oogenesis; polymerase chain reaction; receptor expression; transforming growth factors

The mechanisms regulating the onset of follicular morphogenesis remain unresolved, but the differentiation of granulosa cells from pluripotential somatic cells and their interaction with oocytes appears to be an essential early step. In contrast to rats and mice, ovaries of newborn Syrian hamsters contain oocytes in the 1st meiotic prophase, and undifferentiated somatic cells. Therefore, we have an ideal animal model for studying the mechanism involved in producing the first cohort of follicles. Preliminary studies have shown that: [1] immunoinactivation of endogenous FSH results in nearly complete inhibition of primordial follicle formation and this can be reversed by equine chorionic gonadotropin (eCG), a hormone with FSH activity unaffected by the highly specific anti-FSH serum; [2] the failure of follicular formation is correlated with an appreciable reduction in the transcription and translation of the epidermal growth factor (EGF) receptor gene consistent with our earlier findings that FSH functions via EGF and its receptor; [3] exposure of neonatal ovaries in vivo or in vitro to eCG upregulates the expression of EGF, EGF-receptor and TGF-beta receptor associated with accelerated granulosa cell and follicle differentiation and [4] increased expression of FSH-receptor mRNA correlates with increased FSH function. We hypothesize that FSH controls the differentiation of pluripotential somatic cells into granulosa cells during the first phases of follicular morphogenesis by mechanisms that involve the spatio-temporal interaction of EGF and TGF-beta receptors with their appropriate ligands. This will be tested in vivo using anti-FSH antiserum and in vitro using fetal and early postnatal hamster ovaries exposed to FSH and/or growth factors. Measured parameters include [1] morphometric quantitation of follicle formation; [2] quantitative evaluation by Western immunoblotting, of the significance of EGF and TGF-beta ligand and receptor expression during granulosa cell differentiation; their cell- specific subtle expression will be identified by immunofluorescence; [3] correlation of changes in EGF and TGF-beta receptor mRNA, determined by RT-PCR quantitation, with gonadotropin-regulated differentiation and [4] measurement of ovarian steroidogenic activity to determine the functional significance of follicular development following in vitro hormone and/or growth factor manipulation. Results will contribute substantially to our understanding of the mechanisms involved in the initial steps of folliculogenesis, i.e., the formation of primordial follicles, which will prove valuable for improving the treatment and management of human infertility.

调节卵泡形态发生开始的机制仍未解决，但颗粒细胞从多能体细胞的分化及其与卵母细胞的相互作用似乎是重要的早期步骤。与大鼠和小鼠不同，新生叙利亚仓鼠的卵巢含有处于第一次减数分裂前期的卵母细胞和未分化的体细胞。因此，我们有一个理想的动物模型来研究第一批卵泡产生的机制。初步研究表明： [1] 内源性 FSH 的免疫失活导致原始卵泡形成几乎完全受到抑制，并且这种情况可以被马绒毛膜促性腺激素 (eCG) 逆转，eCG 是一种具有 FSH 活性的激素，不受高度特异性的抗 FSH 血清的影响； [2] 卵泡形成的失败与表皮生长因子 (EGF) 受体基因转录和翻译的明显减少相关，这与我们早期的发现一致，即 FSH 通过 EGF 及其受体发挥作用； [3] 新生儿卵巢在体内或体外暴露于 eCG 会上调与加速颗粒细胞和卵泡分化相关的 EGF、EGF 受体和 TGF-β 受体的表达，并且 [4] FSH 受体 mRNA 表达的增加与 FSH 功能的增强相关。我们假设 FSH 在卵泡形态发生的第一阶段控制多能体细胞分化为颗粒细胞，其机制涉及 EGF 和 TGF-β 受体与其适当配体的时空相互作用。这将使用抗 FSH 抗血清进行体内测试，并使用暴露于 FSH 和/或生长因子的胎儿和出生后早期仓鼠卵巢进行体外测试。测量参数包括 [1] 卵泡形成的形态定量； [2]通过Western免疫印迹定量评估EGF和TGF-β配体和受体表达在颗粒细胞分化过程中的重要性；它们的细胞特异性微妙表达将通过免疫荧光来识别； [3] 通过 RT-PCR 定量测定 EGF 和 TGF-β 受体 mRNA 变化与促性腺激素调节的分化的相关性，以及 [4] 测量卵巢类固醇生成活性，以确定体外激素和/或生长因子操作后卵泡发育的功能意义。结果将大大有助于我们了解卵泡发生的初始步骤（即原始卵泡的形成）所涉及的机制，这对于改善人类不孕症的治疗和管理具有重要意义。