Cellular and Molecular Aspects of Implantation

描述（由申请人提供）：着床后卵膜的形成被认为是胚胎免受母体损伤的保护屏障。因此，植入缺陷和子宫脱卵可能是早期妊娠丢失的原因。鉴定胚胎细胞类型和胚胎因子用于着床和脱卵将是提高生育能力的一项重大成就。产生限制细胞旁通透性屏障的紧密连接（TJ）仅存在于上皮细胞中，而不存在于间质中。我们首次注意到，妊娠第6天，围绕着床囊胚的初级蜕膜区（PDZ）的基质细胞表达TJ及其相关蛋白occludin、claudin-1和zoonula occluden -1 （ZO-1）和ZO-2，而次级蜕膜区（SDZ）的基质细胞则不表达TJ。我们的初步结果表明，囊胚的滋养细胞（Tr）而不是内细胞团（ICM）诱导着床和脱体细胞反应，并在PDZ中正确表达ZO-1。我们还提供证据表明，囊胚产生的肝素结合-表皮生长因子样生长因子（HB-EGF）对着床、脱体细胞化和ZO-1在PDZ中的表达很重要。这些观察结果，再加上静脉注射后正常囊胚诱导的大分子染料通过PDZ的限制，而不是人工诱导的脱个体化，表明胚胎HB-EGF影响着床、PDZ的形成和PDZ中TJ的发生，从而形成一个无血管渗透性屏障来保护胚胎。因此，我们的具体目的是在小鼠模型中确定：1)Tr在着床过程中对PDZ形成的影响，2)胚胎HB-EGF在着床过程中对PDZ形成的影响，3)HB-EGF在基质细胞向具有上皮特征的蜕膜细胞转化过程中的影响，4)PDZ作为母胎界面屏障的重要性。我们将使用多种实验方法，包括RT-PCR， Northern和原位杂交，免疫组织化学/免疫荧光，Western blotting，磷酸化，电子显微镜，胚胎培养和转移，细胞培养，体内/体外腺病毒载体驱动的基因传递等来实现我们的目标。这些结果将进一步加深我们对子宫保护机制对妊娠早期着床胚胎保护的认识。

DESCRIPTION (provided by applicant): The formation of deciduum following implantation is considered to function as a protective barrier to the embryo from the maternal insults. Thus, defective implantation and uterine decidualization may account for early pregnancy losses. Identification of embryonic cell-type(s) and embryonic factor(s) for implantation and decidualization will be a major achievement to improve fecundity. The tight junction (TJ) that creates a barrier restricting the paracellular permeability is present only in cells of the epithelium, but not in the stroma. We noticed for the first time that the stromal cells of the primary decidual zone (PDZ) surrounding the implanting blastocyst, but not the secondary decidual zone (SDZ), on day 6 of pregnancy express the TJ and its associated proteins, occludin, claudin-1, and zonula occludens-1 (ZO-1) and ZO-2. Our preliminary results show that the trophoblast (Tr), but not the inner cell mass (ICM), of the blastocyst induces implantation and decidualization reactions with correct expression of ZO-1 in the PDZ. We also provide evidence here that heparin binding-epidermal growth factor-like growth factor (HB-EGF) produced by the blastocyst is important for implantation, decidualization, and ZO-1 expression in the PDZ. These observations together with the restriction of passage of macromolecular dye to the PDZ induced by normal blastocysts after intravenous administration, but not by artificially induced decidualization, suggest that the embryonic HB-EGF influences implantation, the PDZ formation, and genesis of the TJ in the PDZ which creates an avascular permeability barrier to protect the embryo. Therefore, our specific aims are to determine in a murine model: 1) Influence of the Tr in implantation and the PDZ formation, 2) Influence of embryonic HB-EGF in implantation and the PDZ formation, 3) Influence of HB-EGF in the transformation of stromal cells to decidual cells with epithelial characteristics, and 4) The importance of the PDZ as a barrier at the maternal-fetal interface. We will use multiple experimental approaches including RT-PCR, Northern and in situ hybridizations, immunohistochemistry/ immunofluorescence, Western blotting, phosphorylation, electron microscopy, embryo culture and transfer, cell culture, in vivo/in vitro adenoviral vector-driven gene delivery, and others to accomplish our goals. These results will further our understanding regarding uterine protective mechanism(s) to the safeguard of the implanting embryo during early pregnancy.