Identification of uterine genes induced by embryo during mice implantation.

在这个项目中，我们试图通过从怀孕第5天的小鼠的cDNA中减去从未怀孕小鼠的子宫中构建的cDNA来鉴定参与植入过程的基因。其中一个cDNA编码钙结合蛋白D-9k （Calbindin）。Northern blot分析显示，转录本的大小与大鼠calbindin mRNA相同，并在妊娠第5天小鼠的子宫内表达。我们发现，在妊娠第3天，当血清孕酮浓度较高时，calbindin mRNA的表达增加。事实上，黄体酮诱导钙结合蛋白mRNA的表达显著增加。原位杂交结果显示，妊娠第5天子宫腺上皮calbindin mRNA表达消失。在假妊娠小鼠中，第5天在腺上皮中观察到calbindin mRNA的表达，提示该分子不仅受性类固醇激素的调控，还受胚胎信号的调控。为了检验这种可能性，我们进行了胚胎移植实验。在对照子宫角腺上皮中检测到calbindin mRNA，而在胚胎移植后36 h子宫角中未检测到calbindin mRNA。这些结果表明，在妊娠第5天，胚泡的存在抑制了腺上皮calbindin mRNA的表达。综上所述，我们可以发现胚胎信号在着床过程中调节子宫内膜。我们现在正在分析从减去的cDNA文库中克隆的其他cDNA克隆。这种方法将有助于澄清生殖生理学。

In this project, we sought to identify the genes involved in the implantation process by subtracting cDNA constructed from the uteri of non-pregnant mice from cDNA from those of pregnancy day 5 mice. One of the identified cDNA encoded the calcium binding protein D-9k (Calbindin). Northern blot analysis showed that the size of the transcript was the same as rat calbindin mRNA, and that it was expressed in the uteri of pregnancy day 5 mice. We found that the expression of calbindin mRNA increased on day 3 of pregnancy when the serum progesterone concentration is high. In fact, progesterone induced a marked increase in the expression of calbindin mRNA.In situ hybridization showed that the uterine expression of calbindin mRNA disappeared in the glandular epithelium on day 5 of pregnancy. In the pseudopregnant mice, the expression of calbindin mRNA was observed in the glandular epithelium on day 5, suggesting that this molecule is regulated not only by the sex steroid hormones, but also by the embryonal signal. To examine this possibility, we performed the embryo transfer experiments. While calbindin mRNA was detected in the glandular epithelium of the control uterine horn, it was not detected in the embryo-transferred horn 36 h after embryo transfer. These results indicated that the expression of calbindin mRNA in the glandular epithelium was suppressed by the presence of blastocysts on day 5 of pregnancy.In conclusion we could found that embryonic signals modulate the endometrium during implantation. We are now analyzing other cDNA clones cloned from subtracted cDNA library. This approach will contribute towards the clarification of reproductive physiology.