Studies on the genes and molecules involved in human endometrial cell proliferation and differentiation

对于人类的成功着床，子宫内膜的精确计时变化，包括蜕膜基质细胞的转化，被认为是非常重要的。虽然蜕膜化主要受黄体分泌孕酮的控制，但最近的研究表明，各种其他因素在蜕膜化的调节中起着局部中介作用。孕酮对子宫内膜间质细胞的作用可能存在一些第二信使分子。在本研究中，我们建立了一种人蜕膜的体外模型。在孕酮存在下培养的人子宫内膜间质细胞进行形态分化，并产生催乳素等蜕膜蛋白。通过这个体外蜕膜化模型，我们试图确定与蜕膜化有关的基因。从增生期子宫内膜组织中分离人子宫内膜间质细胞，用Poly(A)RNAs构建cDNA文库。C DNA消减，表达…用更多的Rray和差异显示的方法寻找孕酮诱导子宫内膜间质细胞分化的相关基因。结果鉴定出组织转谷氨酰胺酶(Tgase)、TIMP-3、CD63、IL-15等基因，以及未见报道的UK10基因。孕酮作用6h可诱导组织Tgase基因表达，1h内可诱导未见报道的UK10基因表达，孕酮作用7~10d后CD63表达下调，IL-15表达不受调控。酶组织Tgase是Tgase家族中的一员，催化多肽中谷氨酰胺残基的g-羧基与多肽中赖氨酸残基的伯胺或e-氨基之间的钙依赖的酰基转移反应。组织Tgase在各种器官和组织中都有报道。组织Tgase的生物学功能尚不完全清楚，但已被认为在调节细胞生长和分化、细胞膜钙依赖的交联、细胞程序性死亡以及细胞因子的激活和二聚化等方面发挥作用。提示组织Tgase在蜕膜化中起重要作用，因为在组织Tgase基因中加入特定底物竞争物或反义寡核苷酸可抑制孕酮诱导的蜕膜化。虽然子宫内膜组织Tgase的底物(S)尚不清楚，但该分子(S)应该是孕酮诱导基质细胞分化所必需的。至于UK10基因，我们发现该基因在分泌中期的子宫内膜中有表达，在植入失败的患者中表达降低。孕酮可明显提高子宫内膜间质细胞IL-15的表达水平和IL-15的产生。提示IL-15的产生可刺激子宫NK细胞的产生，是妊娠成功的关键事件。此外，我们还首次阐明了孕酮调节基因--20a-羟基类固醇脱氢酶。较少

For successful implantation in humans, precisely timed changes in the endometrium including the decidual transformation of stromal cells are believed to be very important. Although decidualization is primarily under the control of progesterone secreted by corpus luteum, recent studies have demonstrated that various other factors act as local mediators in the regulation of decidualization. And there should exist some second messenger molecules in progesterone action on endometrial stromal cells. In this study, we have developed an in vitro model of human decidualizaiton. Human endometrial stromal cells cultured in the presence of progesterone undergo morphological differentiation and produce decidual proteins such as PRL. With this model of in vitro decidualization, we tried to identify the genes involved in decidualization. Human endometrial stromal cells from proliferative endometrial tissues were isolated and cDNA libraries were made from poly (A)+RNAs. cDNA subtraction, expression a … More rray, and differential display methods were used to identify genes involved in endometrial stromal cell differentiation induced by progesterone. As a results, several genes including tissue transglutaminase (Tgase), TIMP-3, CD63, IL-15, and unreported gene of UK10 were identified. Tissue Tgase gene was induce after 6 hours culture with progesterone and the unreported gene, UK10, was induced within 1 hour culture, CD63 was down regulated and IL-15 was unregulated by progesterone after 7-10 days culture with progesterone. The enzyme tissue Tgase is a member of the Tgase family that catalyzes calcium-dependent acyl transfer reaction between g-carboxamide group of the glutamine residues in peptides and either primary amines or e-amino groups of the lysine residues in peptides. Tissue Tgase is reported in various organs and tissues. The biological function of tissue Tgase is not completely understood, but it is suggested that the enzyme has roles in regulation of cell growth and differentiation, in calcium-dependent cross linking of cell membrane, in the programmed cell death, and in the activation and dimerization of cytokines. The important role of tissue Tgase in decidualization was suggested, since the addition of a specific substrate competitor, or the antisense oligodeoxynucleotide to tissue Tgase gene inhibited progesterone-induced decidualizaiton. Although the substrate(s) of tissue Tgase in endometrium for is unknown, the molecule(s) should be indispensable for stromal cell differentiation induced by progesterone. As to UK10 gene, we found the gene was expressed in mid-secretary phase endometrium, and its expression was decreased in patients of implantation failure. IL-15 mRNA levels as well as IL-15 production by endometrial stromal cells was apparently increased by progesterone. And it was suggested that IL-15 production stimulates the induction of uterine NK cells and is a key event for the successful pregnancy establishment. In addition, we have clarified a gene for progesterone regulation, 20a-hydroxysteroid dehydrogenase for the first time. Less