Hormonal regulation of estrogen and progestin receptors in decidual cells.
Hormonal regulation of estrogen and progestin receptors in decidual cells.
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蜕膜细胞中雌激素和孕激素受体的激素调节。
DOI：
10.1095/biolreprod35.2.475
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发表时间：
1986
影响因子：
3.6
通讯作者：
Takeda,A
中科院分区：
文献类型：
--
作者：
Leavitt,WW;Takeda,A
总雌激素受体（Re）和总孕酮受体（Rp）测定仓鼠蜕膜组织和蜕膜细胞培养的胞浆和核组分。血清类固醇（雌二醇和孕酮）和蜕膜受体的相关性显示，在第一个四天的蜕膜化，这是不可归因于血清类固醇水平的变化显着的损失。采用蜕膜细胞-组织培养系统研究孕酮戒断后受体的恢复反应。将蜕膜细胞铺板并在含有5%马血清的Ham’s F12/Dulbecco’s改良Eagle’s培养基中生长，所述马血清补充有胰岛素、转铁蛋白、硒和孕酮（10 ng/ml）。在培养48小时内，通过相位显微镜观察到大的多核蜕膜细胞。在含孕酮的培养基中培养72 h时，蜕膜细胞中仅检测到Rp。在孕酮存在下维持的细胞的胞质溶胶或细胞核中未检测到Re（< 200 fmol/mg DNA）。然而，当孕激素从培养基中删除，胞质液Re恢复逐步从8小时至16小时的培养。孕激素戒断也引起细胞质和细胞核Rp的平行增加，雌二醇治疗（2 ng/ml）结合孕激素戒断进一步增强了蜕膜细胞培养物中的Rp水平。这些结果与培养的蜕膜细胞表明，孕酮下调Re和Rp，Re恢复迅速孕酮撤出后，和Re系统是有能力响应雌激素的作用，在RP induction.We使用的密度漂移方法，以确定resistin增加细胞核Re在仓鼠蜕膜细胞在3小时内的营业额。从子宫内膜中分离蜕膜细胞，并在含有正常氨基酸（1H，12 C，14 N）的无胆甾酮培养基中培养2天。将汇合的单层细胞暴露于1 nM雌二醇（E2）1小时，以最大限度地提高细胞核部分中被占用的Re量。然后，在时间0，将细胞转移到补充有致密（2 H、13 C、15 N）氨基酸和1 nM E2或E2加100 nM黄体酮的培养基中。在Re用密集氨基酸标记1、3、6和9 h后，用10 mM吡哆醛−5′磷酸提取核Re，并用125 I-碘雌二醇（5 nM）标记。代表预先存在的和新合成的Re的两个放射性峰通过蔗糖密度梯度离心分离。E2单独作用时，蜕膜细胞核Re的半衰期为3.7h。用E2+孕酮处理使Re的半衰期缩短至1.9 h，甾体激素诱导的Re降解早在类固醇处理后3 h就很明显。E2刺激Re合成，孕酮阻断E2诱导的Re合成。这些结果明确表明，孕酮的作用在下调Re的主要网站是增强Re营业额与随后的抑制作用E2诱导的补充Re。因此，蜕膜细胞培养物可以作为一个模型系统，以研究Re和Rp系统的上调和下调。
Total estrogen receptor (Re) and total progestin receptor (Rp) were measured in the cytosol and nuclear fractions from hamster deciduomal tissue and decidual cell cultures. Correlation of serum steroid (estradiol and progesterone) and deciduomal receptor profiles revealed a significant loss of Re during the first four days of decidualization that was not attributable to changes in serum steroid levels. A decidual cell-tissue culture system was used to study the receptor’s recovery response to progesterone withdrawal. Decidual cells were plated and grown in Ham’s F12/Dulbecco’s modified Eagle’s medium with 5% horse serum supplemented with insulin, transferrin, selenium and progesterone (10 ng/ml). Within 48 h of culture large, multinucleate decidual cells were observed by phase microscopy. At 72 h of culture in medium containing progesterone, only Rp was detectable in decidual cells. Re was not detectable (< 200 fmol/mg DNA) in either cytosol or nuclei from cells maintained in the presence of progesterone. However, when progesterone was deleted from the medium, cytosol Re recovered progressively from 8 h to 16 h of culture. Progesterone withdrawal also caused parallel increases in cytosol and nuclear Rp, and estradiol treatment (2 ng/ml) in combination with progesterone withdrawal further enhanced Rp levels in decidual cell cultures. These results with cultured decidual cells demonstrate that progesterone down-regulates Re and Rp, Re recovers rapidly upon progesterone withdrawal, and the Re system is competent to respond to estrogen action in terms of Rp induction.We used the density-shift method to determine that progestin increases the turnover of nuclear Re in hamster decidual cells within 3 h. Hamster decidual cells were isolated from the endometrium and cultured in progesterone-free medium containing normal amino acids (1H,12C,14N) for 2 days. Confluent monolayers of cells were exposed to 1 nM estradiol (E2) for 1 h to maximize the amount of occupied Re in the nuclear fraction. Then, at time 0, cells were transferred to medium supplemented with dense (2H,13C,15N) amino acids and either 1 nM E2or E2plus 100 nM progesterone. After Re was labeled with dense amino acids for 1, 3, 6 and 9 h, nuclear Re was extracted with 10 mM pyridoxal −5′ phosphate and labeled with125I-iodoestradiol (5 nM). Two radioactive peaks representing preexisting and newly synthesized Re were separated by sucrose density-gradient centrifugation. The halflife of nuclear Re in decidual cells was 3.7 h when cells were treated with E2alone. Treatment with E2+ progesterone shortened the half life of Re to 1.9 h, and progesterone-induced Re degradation was evident as early as 3 h after steroid treatment. E2stimulated the synthesis of Re, and progesterone blocked E2-induced Re synthesis. These results demonstrate unequivocally that the primary site of progesterone action in down-regulating Re is the enhancement of Re turnover with a subsequent inhibitory effect on E2-induced replenishment of Re. Thus, decidual cell cultures can serve as a model system to study the up- and down-regulation of Re and Rp systems.
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