OOCYTE MATURATION--VITAL STAIN-VIDEO MICROSCOPY ANALYSIS

卵母细胞成熟--活体染色-视频显微镜分析

• 批准号: 3317884
• 负责人: DAVID F ALBERTINI
• 金额: $ 15万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-04-01 至 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3317884
• 关键词: actins; animal tissue; biological signal transduction; cell cell interaction; cell cycle proteins; centrosome; confocal scanning microscopy; digital imaging; egg /ovum; electron microscopy; fluorescence microscopy; fluorescent dye /probe; follicle stimulating hormone; genetic translation; hormone regulation /control mechanism; immunocytochemistry; immunofluorescence technique; in situ hybridization; laboratory mouse; meiosis; microinjections; mitotic spindle apparatus; oogenesis; phosphorylation; posttranscriptional RNA processing; posttranslational modifications; protein kinase; protein metabolism; tissue /cell culture; video recording system; western blottings

Oogenesis culminates in the production of viable and developmentally competent ova as a result of an intricate series of metabolic changes in the cumulus-oocyte complex induced by hormones at ovulation. Disturbances in oocyte maturation lead to reproductive failure, manifest as infertility, spontaneous abortion, or birth defects, when errors in chromosome segregation occur during meiosis or when the hormonal requirements for the maturation process are not met. This proposal seeks to define important physiological aspects of oocyte maturation in mammals with respect to protein metabolism, meiotic cell cycle control and somatic cell-gamete interactions using a multidisciplinary approach. The major aims are: (1) To establish how changes in protein metabolism are regulated during meiotic maturation in the mouse oocyte. Translational, and post- translational metabolic processes will be evaluated with respect to meiotic competence acquisition and meiotic resumption/completion. The role of the cytoskeleton in mRNA processing will be analyzed by a combination of biochemical, morphological and micromanipulation procedures. (2) To determine how components of Maturation-Promoting Factor (MPF) mediate meiotic cell cycle progression. The role of centrosomes in activation of the M-phase kinase, localization of p34cdc2/cyclins and morphogenesis of the meiotic spindle will be analyzed using fluorescence digital imaging microscopy and biochemical techniques. (3) To define the physiological basis for information transfer between follicle cells and oocytes. Compositional and functional parameters of junctional contact sites will be analyzed in mouse and cow oocytes during meiotic maturation using vital stain video, fluorescence ratio and confocal imaging light microscopic methods. Information obtained from these studies will help to understand the causes of infertility in women, provide new avenues for improving assisted reproductive technologies, and open new directions for the study of environmentally-based determinants of reproductive failure in mammals.

卵子发生最终产生可存活且可发育的卵子 有能力的卵子是一系列复杂的代谢变化的结果 排卵时激素诱导的卵丘-卵母细胞复合体。 卵母细胞成熟障碍导致明显的生殖失败 如不孕症、自然流产或出生缺陷，当错误 染色体分离发生在减数分裂期间或当激素 不满足成熟过程的要求。 该提案寻求 定义哺乳动物卵母细胞成熟的重要生理方面 关于蛋白质代谢、减数分裂细胞周期控制和 使用多学科方法进行体细胞-配子相互作用。 这 主要目标是： (1) 确定蛋白质代谢的变化是如何调节的 小鼠卵母细胞减数分裂成熟。 翻译和后期 翻译代谢过程将根据以下方面进行评估 减数分裂能力的获得和减数分裂的恢复/完成。 细胞骨架在 mRNA 加工中的作用将通过以下方式进行分析 生物化学、形态学和显微操作的结合 程序。 (2) 确定成熟促进因子(MPF)的成分 介导减数分裂细胞周期进程。 中心体的作用 M 期激酶的激活，p34cdc2/细胞周期蛋白的定位 减数分裂纺锤体的形态发生将使用 荧光数字成像显微镜和生化技术。 (3) 定义人与人之间信息传递的生理基础 卵泡细胞和卵母细胞。 组成和功能 将在小鼠中分析连接接触位点的参数 和减数分裂成熟过程中的牛卵母细胞使用活体染色视频， 荧光比率和共焦成像光显微方法。 从这些研究中获得的信息将有助于理解 为女性不孕不育的原因提供新的改善途径 辅助生殖技术，为研究开辟新方向 哺乳动物生殖失败的环境决定因素。