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Regulation of Ca2+ influx in mouse oocytes and eggs during maturation and fertilization to improve assisted reproductive technologies and modulate fertility

调节小鼠卵母细胞和卵在成熟和受精过程中的 Ca2 流入，以改进辅助生殖技术并调节生育力

基本信息

批准号：
10401470
负责人：
Rafael Antonio Fissore
金额：
$ 25.33万
依托单位：
UNIVERSITY OF MASSACHUSETTS AMHERST
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-17 至 2024-05-31
项目状态：
已结题

项目摘要

There is a persistent gap in our knowledge of the molecules and mechanisms that mediate Ca2+ influx into oocytes and eggs. Ca2+ influx is required for egg activation and embryo development in all mammals, including human beings. This gap in our knowledge therefore represents a serious impediment, as until it is filled we cannot physiologically modulate Ca2+ influx or objectively diagnose and treat infertility associated with disturbances in Ca2+ homeostasis. The long-term goal is to understand how Ca2+ homeostasis is regulated in oocytes and eggs and identify its molecular effectors. The objective here is to identify the Ca2+ channel(s) that mediate Ca2+ influx during maturation and fertilization and characterize their regulatory mechanisms. Mouse oocytes/eggs are a great model because they display distinctive Ca2+ entry during maturation and fertilization, and Ca2+ release is required for egg activation. The central hypothesis is that expression and/or distinct regulation of underdetermined Ca2+ channels on the plasma membrane underlies Ca2+ influx in oocytes, its inactivation during maturation and its recovery after fertilization. This hypothesis was conceived based on extensive preliminary data. The rationale for this research is that once the channels are identified, a better understanding of the molecular determinants of oocyte maturation and fertilization will be gained. The findings here also have the potential to translate into therapeutic methods to assist infertile couples in this country. We plan to test our central hypothesis by pursuing the following specific aims: 1) Identify the Ca2+ channel(s) that mediate Ca2+ influx in oocytes prior to and during maturation; and 2) Identify the Ca2+ influx channel(s) that support oscillations after fertilization. Under Aim 1, Ca2+ imaging, pharmacology, conditional knockout mice, and electrophysiology will be used to identify the active channel(s) and to assess the impact of Ca2+ influx on maturation; the role of a TRPM7-like current recently discovered by the applicant and collaborators will be closely examined. Under Aim 2, the signaling mechanism(s) whereby fertilization stimulates Ca2+ influx and the contributing channel(s) will be determined. A novel approach that overcomes the inactivation of Ca2+ influx in eggs will facilitate these studies. Genetic models and electrophysiology will confirm the function of these channel(s) during fertilization. The research in this application is innovative because it combines electrophysiology, pharmacology and new KO lines, approach that has served to identify two new channels in oocytes, including TRPM7, whose global deletion is embryonic lethal at E7.5. The contribution of the proposed project is significant because it is expected to allow physiological modulation of Ca2+ entry in oocytes and eggs that will produce new conditions and activation protocols for use in the clinic. It will also expand our understanding of the impact of Ca2+ homeostasis on oocyte competence and infertility.

我们对调节钙离子内流的分子和机制的了解一直存在空白。变成卵母细胞和卵子。在所有哺乳动物中，卵子激活和胚胎发育都需要钙离子内流，包括人类在内。因此，我们知识上的这种差距是一个严重的障碍，直到它充满了我们无法从生理上调节钙离子内流或客观诊断和治疗不孕不育与钙离子动态平衡失调有关。长期目标是了解钙离子是如何动态平衡在卵母细胞和卵子中受到调节，并确定其分子效应器。这里的目标是成熟受精过程中介导钙离子内流的钙通道(S)的鉴定及特性他们的监管机制。小鼠卵母细胞/卵子是一个很好的模型，因为它们表现出独特的卵子在成熟和受精过程中需要钙离子进入，而激活卵子需要钙离子的释放。中环假说是血浆上钙离子通道的表达和/或明显的调节卵母细胞膜是钙离子内流的基础，成熟过程中钙离子失活，成熟后恢复。受精。这一假设是基于广泛的初步数据而提出的。这样做的理由是研究表明，一旦确定了通道，就会更好地理解分子决定因素将获得卵母细胞成熟和受精的可能性。这里的发现也有可能转化为以治疗方法来帮助这个国家的不孕夫妇。我们计划测试我们的中心假设通过追求以下具体目标：1)确定介导卵母细胞内钙内流的钙通道(S) 成熟前和成熟中；2)确定支持成熟后振荡的钙离子内流通道(S) 受精。在目标1下，钙离子成像、药理学、条件性基因敲除小鼠和电生理将被用来识别活跃的通道(S)，并评估钙离子内流对成熟的影响；申请人和合作者最近发现的TRPM7类洋流的作用将密切相关检查过了。在目标2下，受精刺激钙内流的信号机制(S)和供稿渠道(S)将确定。一种克服钙离子失活的新方法卵子的大量涌入将促进这些研究。遗传模型和电生理学将证实这一功能这些通道(S)在受精过程中。这一应用的研究具有创新性，因为它结合了电生理学、药理学和新的KO系，这一方法已用于识别两个新的卵母细胞中的通道，包括TRPM7，其全局缺失在e7.5时是胚胎致死的。这个拟议项目的贡献很大，因为预计它将允许生理调节在卵母细胞和卵子中的钙离子进入将产生新的条件和激活方案用于诊所。这也将扩大我们对钙离子稳态对卵母细胞能力的影响的理解。和不孕不育。

项目成果

期刊论文数量（11）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Multidrug resistance transporter-1 dysfunction perturbs meiosis and Ca2+ homeostasis in oocytes.

DOI：
10.1530/rep-22-0192
发表时间：
2023-01-01
期刊：
REPRODUCTION
影响因子：
3.8
作者：
Nabi, Dalileh;Bosi, Davide;Gupta, Neha;Thaker, Nidhi;Fissore, Rafael;Brayboy, Lynae M.
通讯作者：
Brayboy, Lynae M.

SPERM FACTORS AND EGG ACTIVATION: ICSI and the discovery of the sperm factor and PLCZ1.