Hamster: a unique model for studying implantation

描述(申请人提供)：卵巢孕酮是所有研究物种着床所必需的。然而，在这一过程中对胚胎雌激素的需求仍然是一个悬而未决的问题，除了在小鼠和大鼠中，胚胎不具备合成雌激素的机制。植入可以发生在几个物种，包括仓鼠，也许在没有卵巢雌激素的情况下，如果提供黄体酮，人类也可能这样做。然而，我们最近的研究表明：(1)雌激素而不是孕酮调节去卵巢仓鼠子宫中肝素结合的EGF-样生长因子基因(Hegf1)的表达；(2)在黄体酮诱导的去垂体怀孕仓鼠中，Hegf1的表达发生在胚泡周围的子宫腔上皮细胞中，而在没有循环雌激素的情况下。这些结果表明，仓鼠胚泡可能是雌激素的替代来源，局部作用于子宫，诱导Hegf1基因表达。事实上，我们的初步实验表明，细胞色素P450芳香酶蛋白存在于仓鼠桑椹和囊胚中。此外，在培养中用类固醇细胞色素P450芳香酶抑制剂处理金黄地鼠桑椹胚会显著抑制胚泡的形成。这些观察，加上我们对植入前仓鼠胚胎和子宫中雌激素受体-α表达的初步结果，使我们假设该物种不依赖卵巢雌激素，但需要胚胎雌激素来启动胚胎和着床所需的子宫变化。然而，胚胎雌激素的合成及其在金黄地鼠胚泡形成、子宫容受性和着床中的作用尚未得到解决。因此，我们的具体目标是在仓鼠身上研究：(1)仓鼠着床前胚胎是否产生雌激素？(2)桑胚-囊胚转化是否需要胚胎雌激素？(3)建立子宫着床接受性是否需要胚胎雌激素？(4)在没有胚胎雌激素的情况下，系统雌激素是否诱导子宫接受性和着床？我们将利用RT-PCR、原位杂交、免疫组织化学、免疫荧光、Western blotting、胚胎培养和移植等多种方法来实现我们的目标。随着对胚胎发育和子宫着床容受性机制的更好理解，有关妇女生育和不孕的问题将得到更有效的管理。

DESCRIPTION (provided by applicant): Ovarian progesterone is essential for implantation in all species studied. However, the requirement of embryonic estrogen in this process is still an unsettled issue, except in mice and rats in which embryos do not possess the machinery for estrogen synthesis. Implantation can occur in several species including hamsters and perhaps in humans in the absence of ovarian estrogen if progesterone is provided. However, our recent study shows that: (1) estrogen, but not progesterone, regulates the uterine expression of heparin binding EGF-like growth factor gene (Hegfl) in ovariectomized hamsters, and (2) Hegfl expression occurs in the uterine luminal epithelium surrounding the blastocyst at the time of implantation in the absence of circulating estrogen in progesterone-primed hypophysectomized pregnant hamsters. These results suggest that hamster blastocysts could be an alternative source of estrogen that acts locally to induce Hegfl gene expression in the uterus. Indeed, our preliminary experiments suggest the presence of cytochrome P450 aromatase protein in hamster morulae and blastocysts. Furthermore, the exposure of hamster morulae to a steroidal cytochrome P450 aromatase inhibitor in culture significantly inhibits the formation of blastocysts. These observations together with our preliminary results of estrogen receptor-alpha expression in preimplantation hamster embryos and uterus at the time of implantation led us to postulate that this species does not depend on ovarian estrogen, but requires embryonic estrogen to initiate embryonic and uterine changes required for implantation. However, neither the synthesis nor the role of embryonic estrogen in blastocyst formation, uterine receptivity and implantation in hamsters has been resolved. Thus, our specific aims to study in hamsters: (1) Do hamster preimplantation embryos produce estrogen? (2) Is embryonic estrogen required for morula-blastocyst transformation? (3) Is embryonic estrogen required for the establishment of uterine receptivity for implantation? (4) Does systemic estrogen induce uterine receptivity and implantation in the absence of embryonic estrogen? We will use multiple approaches including RT-PCR, in situ hybridization, immunohistochemistry, immunofluorescence, Western blotting, embryo culture and transfer and others to accomplish our goals. With better understanding of the mechanisms of embryo development and uterine receptivity for implantation, issues concerning fertility and infertility in women will be more effectively managed.