Biochemical & Proteomic Analysis of Cumulus Cell Secreted Zinc Inhibitory Factor

生化

• 批准号: 8583461
• 负责人: FRANCISCO J DIAZ
• 金额: $ 7.48万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-21 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8583461
• 关键词: Acute; Area; Biochemical; Cell Communication; Cells; Complex; Conditioned Culture Media; Data; Defect; Development; Diagnosis; Diet; EGF gene; Embryo; Embryonic Development; Excision; Failure; Female; Fertility; Fertilization; Functional disorder; Health; Heating; Homeostasis; Hour; In Vitro; Infertility; Knowledge; Label; Mediating; Meiosis; Metabolism; Metaphase; Methods; Molecular Weight; Mus; Nuclear; Oocytes; Ovarian; Ovulation; Paracrine Communication; Pathway interactions; Pentetic Acid; Preparation; Procedures; Process; Proteins; Proteomics; Pump; Reagent; Regulation; Reproductive Health; Reproductive Medicine; Research Activity; Role; Sampling; Somatic Cell; Technology; Testing; Transition Elements; Trypsin; Zinc; Zinc deficiency; egg; granulosa cell; improved; in vivo; liquid chromatography mass spectrometry; mass spectrometer; metal chelator; nano; novel; oocyte maturation; poor egg quality; premature; prevent; public health relevance; reproductive function; success; treatment duration; zygote

DESCRIPTION (provided by applicant): Compromised fertility is often caused by problems during the periovulatory transition. Meiotic defects, ovulation failure and poor egg quality all contribute to decreased fertility. Oocyte-granulosa cell communication is very important for fertilization and embryo development. However, the specific paracrine signals and the mechanisms controlled by secreted factors in the follicle are not completely know or understood. The LH surge causes dramatic changes in the cumulus-oocyte complex (COC), including expansion of the cumulus cells and oocyte cytoplasmic and nuclear maturation. Recent evidence strongly supports a role for zinc-mediated processes in the regulation of oocyte maturation and fertilization. Cumulus cells secrete a zinc inhibitory factor (ZIF) that potently suppresses zinc accumulation in the oocyte before ovulation. This product is a novel paracrine signal regulating zinc metabolism in the follicle. The objective of this proposal is the biochemica and proteomic analysis of ZIF produced by cumulus cells. In specific aim 1, we will fractionate cumulus cell secreted products according to apparent molecular weight. Next, all secreted proteins in the fraction containing the highest ZIF activity will be analyzed by a sensitive nano-liquid chromatography mass spectrometry (nano LC MS) method. In aim 2, the effect on oocyte fertility of a preparation of conditioned medium containing high ZIF activity will be tested. Together results from these aims will develop a valuable reagent able to precisely control zinc homeostasis during in vitro maturation/fertilization procedures.

描述（由申请人提供）：造成的生育能力通常是由卵巢周围过渡期间的问题引起的。减数分裂缺陷，排卵衰竭和卵质量差，都导致生育能力降低。卵母细胞颗粒细胞通信对于受精和胚胎发育非常重要。但是，卵泡中分泌因子控制的特定旁分泌信号和机制尚未完全了解或理解。 LH激增会导致积云 - 核细胞络合物（COC）的急剧变化，包括卵丘细胞的扩展和卵母细胞的细胞质和核成熟。最近的证据强烈支持锌介导的过程在调节卵母细胞成熟和施肥中的作用。积云细胞分泌一个锌抑制因子（ZIF），该因子在排卵前有效抑制锌在卵母细胞中的积累。该产品是一种调节卵泡中锌代谢的新型旁分泌信号。该提案的目的是对积云细胞产生的ZIF的生物化学和蛋白质组学分析。在特定的目标1中，我们将根据明显的分子量分割积云细胞分泌的产物。接下来，将通过敏感的纳米液色谱质谱法（Nano LC MS）方法分析包含最高ZIF活性的馏分中的所有分泌蛋白。在AIM 2中，将测试对含有高ZIF活性的条件培养基制备的卵母细胞生育能力的影响。这些目标的结果将共同开发出一种有价值的试剂，能够在体外成熟/受精程序中精确控制锌稳态。