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.

调节卵泡形态发生的机制尚未解决，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一个必不可少的早期步骤。与大鼠和小鼠相反，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，用于研究参与产生第一批卵泡的机制。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抑制，这可以被马绒毛膜促性腺激素（eCG）逆转，这是一种具有FSH活性的激素，不受高度特异性抗FSH血清的影响;[2]毛囊形成的失败与表皮生长因子转录和翻译的明显减少有关（EGF）受体基因与我们早期发现FSH通过EGF及其受体发挥功能一致; [3]体内或体外新生儿卵巢暴露于eCG上调EGF、EGF受体和TGF-β受体的表达，与加速颗粒细胞和卵泡分化相关[4] FSH受体mRNA表达增加与FSH功能增加相关。我们假设FSH通过EGF和TGF-β受体与其相应配体的时空相互作用机制，在卵泡形态发生的第一阶段控制多能体细胞向颗粒细胞的分化。将使用抗FSH抗血清进行体内试验，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试验。测量的参数包括[1]卵泡形成的形态定量; [2]通过蛋白质免疫印迹定量评价颗粒细胞分化期间EGF和TGF-β配体和受体表达的意义;通过免疫荧光鉴定它们的细胞特异性细微表达;[3]通过RT-PCR定量测定EGF和TGF-β受体mRNA变化的相关性，促性腺激素调节的分化和[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理解卵泡发生的初始步骤所涉及的机制，即，原始卵泡的形成，这将证明对改善人类不育症的治疗和管理有价值。