Sexual dimorphism in embryo responses to maternal regulatory factors

描述(申请人提供)：植入前胚胎的发育程序很容易因微环境的改变而改变。由此产生的发育轨迹的变化可能会产生长期的影响，并延伸到出生后的生活中。在植入前阶段，男性胚胎和女性胚胎对环境变化的反应可能不同。我们的牛胚胎实验室最近的数据表明，对微环境变化的性别二型性反应可能是通过对母体调节信号的性别特异性反应来调节的。在胚胎发育的第5-7天用集落刺激因子2(CSF2)处理胚胎，在移植给雌性胚胎和在怀孕第15天恢复后，胚胎的特征会发生性别特异性的变化。如果这种性二态现象是一种普遍现象，就需要进行针对性别的分析，以了解控制发育的基本机制、环境对胚胎发育的影响，以及提高女性生育力的治疗方法。目前的提案试图确定对胚胎发育的监管在多大程度上取决于性别。目前有两个直接的目标：建立一个模型，以了解CSF2如何在胚胎发育中发挥性别特异性影响的分子基础，以及评估性二态现象在着床前发育调控中是否普遍存在(即，存在于不止一个调控分子中)。为了实现这些目标，我们将利用我们实验室的大量初步数据来描述CSF2和另外两个子宫调节因子-胰岛素样生长因子1(IGF1)和Dickkopf相关蛋白1(Dkk1)-如何改变牛囊胚的发育。特别是，我们将测试当使用X或Y分类的精子产生胚胎时，CSF2、IGF1和Dkk1对胚胎的作用是否以性别特有的方式发生。对于目标1，将确定牛桑椹胚和囊胚对CSF2的反应的性别差异。将进行四个实验，以评价CSF2对胚胎的性别二型性调节，包括桑椹胚和囊胚中的基因表达，抑制细胞凋亡，以及增加囊胚内细胞团中的细胞数量。目的2将通过测试IGF1和Dkk1是否以性别特异性的方式对牛胚胎发挥作用，来确定其他调控因子是否存在性别二型性。实验将确定男性和女性胚胎在IGF1对胚泡发育、基因表达和细胞凋亡的作用方面的差异，以及Dkk1对胚泡中细胞分配给上胚层、原始内胚层和滋养外胚层细胞的作用。这些实验对于发展研究模型以描述建立性别二型性的机制(目标1)和确定性别二型性作为胚胎发育调节的重要决定因素的重要性(目标2)是重要的。

DESCRIPTION (provided by applicant): The developmental program of the preimplantation embryo is readily modified by alterations in the microenvironment. The resultant change in trajectory of development can have long-acting effects that extend into post-natal life. Male and female embryos can respond differently to environmental modification during the preimplantation period. Recent data from our laboratory with bovine embryos indicate that sexual dimorphism in response to changes in the microenvironment could be mediated by gender-specific responses to maternal regulatory signals. Treatment of embryos with colony-stimulating factor 2 (CSF2) from Day 5-7 of development caused gender-specific alterations in characteristics of embryos after transfer to females and recovery at Day 15 of gestation. If this phenomenon of sexual dimorphism is a general one, gender-specific analysis will be required to understand basic mechanisms controlling development, environmental effects on embryonic development, and therapeutic approaches to improving fertility in women. The current proposal seeks to determine the extent to which regulation of embryonic development depends on gender. There are two immediate goals: to develop a model for understanding the molecular basis for how CSF2 exerts gender-specific effects on embryonic development and to evaluate whether the phenomenon of sexual dimorphism in regulation of preimplantation development is widespread (i.e., exists for more than one regulatory molecule). To meet these objectives, we will take advantage of a large amount of preliminary data from our laboratory that describes how CSF2 and two other uterine regulatory factors, insulin-like growth factor 1 (IGF1) and dickkopf-related protein 1 (DKK1), modify development of the bovine blastocyst. In particular, we will test whether actions of CSF2, IGF1 and DKK1 on embryos seen previously occur in a gender-specific manner when embryos are produced using X- or Y-sorted spermatozoa. For Aim 1, gender differences in response of bovine morulae and blastocysts to CSF2 will be determined. Four experiments will be conducted to evaluate sexual dimorphism in regulation of the embryo by CSF2 with respect to gene expression in morulae and blastocysts, inhibition of apoptosis, and increase in numbers of cells in the inner cell mass of the blastocyst. Aim 2 will determine whether sexual dimorphism exists for other regulatory factors by testing whether IGF1 and DKK1 exert actions on bovine embryos in a gender-specific manner. Experiments will be performed to determine differences between male and female embryos with respect to IGF1 actions on blastocyst development, gene expression and apoptosis and with respect to DKK1 actions on allocation of cells in the blastocyst to cells of the epiblast, primitive endoderm, and trophectoderm lineages. These experiments are important to develop research models for delineating mechanisms by which sexual dimorphism is established (Aim 1) and to determine the importance of sexual dimorphism as an important determinant of regulation of embryonic development (Aim 2).