The mineralocorticoid receptor may compensate for the loss of the glucocorticoid receptor at specific stages of mammary gland development

To study the role of glucocorticoid receptor (GR) at different stages of mammary gland development, mammary anlage were rescued from GR-/- mice by transplantation into the cleared fat pad of wild-type mice. In virgin mice, GR-/- outgrowths displayed abnormal ductal morphogenesis characterized by distended lumena, multiple layers of luminal epithelial cells in some regions along the ducts, and increased periductal stroma. In contrast, the loss of GR did not result in overt phenotypic changes in mammary gland development during pregnancy, lactation, and involution. Surprisingly, despite the known synergism between glucocorticoids and prolactin in the regulation of milk protein gene expression, whey acidic protein and beta-casein mRNA levels were unaffected in GR-/- transplants as compared with wild-type transplants. That mineralocorticoid receptor (MR) might compensate for the loss of GR was suggested by the detection of MR in the mammary gland at d 1 of lactation. This hypothesis was tested using explant cultures derived from the GR-/- transplants in which the mineralocorticoid fludrocortisone was able to synergistically induce beta-casein gene expression in the presence of prolactin and insulin. These studies suggest that MR may compensate for the absence of GR at some, but not at all stages of mammary gland development.     

Partial characterization and ontogeny of renal cytosolic glucocorticoid receptors in mouse kidney

The glucocorticoid receptor (GR) was partially characterized in mouse renal cytosol. A sensitive and reproducible [3H]dexamethasone binding assay suitable for use with small quantities of cytosolic protein, was developed. Studies defined the optimal equilibrium binding conditions, metabolism of [3H]dexamethasone in adult renal cytosol, specificity of binding of the GR, and molecular weight of the GR-[3H]dexamethasone complex by gel filtration chromatography. The assay was subsequently used to measure the renal GR during different stages of foetal and postnatal development, as well as in glomerular and renal tubular preparations from adult mice. An almost linear increase in GR occurred from day 13 to day 18 of gestation with levels rising from 100 to 201 fmol/mg cytosol protein; this was followed by a sharp rise in receptor concentration just after birth to 343 fmol/mg cytosol protein. Adult levels, 410-433 fmol/mg cytosol protein, were reached by 2 weeks after birth. The equilibrium dissociation constants (Kd) of the [3H]dexamethasone-receptor complex were similar in adult and in embryonic cytosols (range, 2.8-11.8 nM; mean +/- SD = 6.5 +/- 2.9 nM). Specific binding was assessed to be 3- to 5-fold greater in tubular than in glomerular preparations. These data on the localization and ontogeny of GR during murine metanephric development provide a basis for study of glucocorticoid-mediated effects on various models of congenital and acquired renal disease.     

Glucocorticoid activity during lung maturation is essential in mesenchymal and less in alveolar epithelial cells

Corticosteroid treatment is an established therapy for preterm infants, and germline inactivation of the glucocorticoid receptor (GR) gene in the mouse leads to respiratory failure and postnatal lethality. Although glucocorticoids have been thought to critically act in epithelial cells inducing the functional maturation of the lung, inactivation of the GR gene exclusively in the epithelium of the developing murine lung did not impair survival. In contrast, mice lacking GR specifically in mesenchyme-derived cells displayed a phenotype strongly reminiscent of GR knockout animals and died immediately after birth. Detailed analysis of gene expression allows the conclusion that GR acts in cells of the fibroblast lineage controlling their proliferation rate and the composition of the extracellular matrix.     

The glucocorticoid receptor is increased in Atm-/- thymocytes and in Atm-/- thymic lymphoma cells, and its nuclear translocation counteracts c-myc expression

We have previously demonstrated that spontaneous DNA synthesis in immature thymocytes of Atm-/- mice is elevated, and that treatment with the glucocorticoid dexamethasone (Dex) attenuates this increased DNA synthesis and prevents the development of thymic lymphomas. Deregulation of c-myc may drive the uncontrolled proliferation of Atm-/- thymocytes, since upregulation of c-myc parallels the elevated DNA synthesis in the cells. In this study, we show that the glucocorticoid receptor (GR) is expressed at high levels in Atm-/- thymocytes and in Atm-/- thymic lymphoma cells, although serum glucocorticoid (GC) levels in Atm-/- mice are similar to those in Atm+/+ mice. In cultured Atm-/- thymic lymphoma cells treated with Dex, GR nuclear translocation occurs, resulting in suppression of DNA synthesis and c-myc expression at both the mRNA and protein levels. Interestingly, the GR antagonist RU486 also causes GR nuclear translocation, but does not affect DNA synthesis and c-myc expression in Atm-/- thymic lymphoma cells. As expected, RU486 reverses the suppressive effects of Dex on DNA synthesis and c-myc expression. Administration of Dex to Atm-/- mice decreases the elevated c-Myc protein levels in their thymocytes. These findings suggest that GC/GR signaling plays an important role in regulating c-myc expression in Atm-/- thymocytes and thymic lymphoma cells.     

Glucocorticoid receptor antagonizes EGFR function to regulate eyelid development

The glucocorticoid receptor (GR) plays a crucial role in epidermal morphogenesis during embryonic development, as demonstrated by analyzing genetically modified mouse models of GR gain- and loss-of-function. Eyelid formation constitutes a useful model to study epithelial development, as it requires coordinated regulation of keratinocyte proliferation, apoptosis and migration. We have analyzed this biological process in GR(-/-) embryos during ontogeny. Our data demonstrate that GR deficiency results in delayed and impaired eyelid closure, as illustrated by increased keratinocyte proliferation and apoptosis along with impaired differentiation in GR(-/-) eyelid epithelial cells. These defects are due, at least in part, to the lack of antagonism between GR and epidermal growth factor receptor (EGFR) signaling, causing sustained activation of the MAPK/AP-1 pathway and the upregulation of keratin K6 at embryonic stage E18.5. Additionally, we demonstrate that GR regulates epithelial cell migration in vitro by interfering with EGFR-mediated signaling. Overall, GR/EGFR antagonism appears as a major mechanism regulating ocular epithelial development.     

糖皮质激素受体及其选择性调节剂研究进展

糖皮质激素（glucocorticoids,GCs）是临床上广泛使用的一类抗炎药物,在体内主要通过糖皮质激素受体（glucocorticoid receptor,GR）发挥生理和药理作用。GR是核受体超家族的成员之一,为配体激活的转录因子,在机体的多种生理和病理活动中扮演重要的角色。随着对GR信号通路的深入研究,寻找针对糖皮质激素受体的新型调节剂,以期将抗炎作用和现有糖皮质激素的副作用相分离,已经成为新药发现的研究热点。本文对近年来GR的分子结构、生物学作用及其选择性调节剂的研究进展作一简要的介绍。     

Maternal postpartum learned helplessness (LH) affects maternal care by dams and responses to the LH test in adolescent offspring

It is known that the early environment affects the mental development of rodent and human offspring. However, it is not known specifically whether a postpartum depressive state influences the depressive state in offspring. Using learned helplessness (LH) in rats as an animal model of depression, we examined the influence of maternal postpartum LH on responses to the LH test of offspring. Dam rats were judged as LH or non-helpless (nLH) on postnatal days (PN) 2-3, and maternal behavior was recorded during PN2-14. On PN 45-46, offspring were subjected to the LH test. Plasma corticosterone (CORT) levels, hippocampal levels of glucocorticoid receptor (GR) and brain-derived neurotrophic factor (BDNF) mRNA were measured before and after the LH test in offspring. Active nursing in LH dams was significantly lower than that in nLH dams. Susceptibility to LH in the offspring of LH dams was significantly higher than in those of nLH dams, and was negatively correlated with active nursing by LH dams. The GR mRNA levels before and after the LH test were lower in the offspring of LH dams than in those of nLH dams, and the reduced basal GR mRNA and protein might have resulted in the higher CORT response after the LH test. There was no significant difference in BDNF mRNA in the offspring of LH and nLH dams. These findings suggest that early postpartum LH decreased active nursing and increased depression-like behavior in the adolescent offspring via dysfunction of the hypothalamic-pituitary-adrenal axis.     

A chimeric Cre recombinase inducible by synthetic,but not by natural ligands of the glucocorticoid receptor.

We have developed a new ligand-dependent chimeric recombinase (Cre-GRdex) by fusing the site-specific Cre recombinase to the ligand binding domain (LBD) of a mutant human glucocorticoid receptor (GRdex). The synthetic glucocorticoid receptor (GR) ligands dexamethasone, triamcinolone acetonide and RU38486efficiently induce recombinase activity in F9 murine embryonal carcinoma cells expressing constitutively Cre-GRdex. In contrast, no recombinase activity was detected in the absence of ligand or in the presence of the natural GR ligands corticosterone, cortisol or aldosterone. Moreover, physiological concentrations of these natural GR ligands do not affect Cre-GRdexrecombinase activity induced by dexamethasone. Thus, as previously shown using Cre-oestrogen receptor (ER) fusion proteins, Cre-GRdexmight be useful for achieving loxP site-directed mutagenesis in cultured cells and spatio-temporally controlled somatic cell mutagenesis in transgenic mice.     

Disruption of β-arrestins blocks glucocorticoid receptor and severely retards lung and liver development in mice

In this study, the role of β-arrestin 1 and β-arrestin 2 in fetal lung and liver development was examined using Arrb1(-/-)Arrb2(-/-) mouse embryos. β-Arrestin 1/2 dual-null mice died shortly after birth and morphological examination revealed an obvious pulmonary hypoplasia and severe hepatic impairment. Western blot analysis demonstrated that GR protein levels in Arrb1(-/-)Arrb2(-/-) lung and liver tissues were significantly decreased compared to wild type embryos. Expression of GR proteins was confirmed in the nuclei of type II pneumocytes of 18.5 day embryos (E18.5) by immunofluorescence. The production of hepatic glucose and mRNA level of gluconeogenic enzymes were dramatically reduced in E18.5 Arrb1(-/-)Arrb2(-/-) liver. These results suggest that GR is an important downstream effector of the β-arrestin signaling pathway involved in regulation of lung and liver development. However, no obvious changes in GR expression following in vitro modulation of β-arrestin 1/2 indicated the existence of an indirect regulatory relationship between GR and the β-arrestin signaling pathway.     

Spatial and temporal patterns of expression of 11beta-hydroxysteroid dehydrogenase types 1 and 2 messenger RNA and glucocorticoid receptor protein in the murine placenta and uterus during late pregnancy

To gain insight into the role of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) enzymes and actions of glucocorticoids in the murine placenta and uterus, the expression pattern of the mRNA for 11beta-HSD1 and 11beta-HSD2 and the glucocorticoid receptor (GR) protein were determined from Embryonic Day 12.5 (E12.5, term = E19) to E18.5 by in situ hybridization and immunohistochemistry, respectively. Consistent with its putative role in regulating the transplacental passage of maternal glucocorticoid to the fetus, 11beta-HSD2 mRNA was highly expressed in the labyrinthine zone (the major site of maternal/fetal exchange) at E12.5, and its level decreased dramatically at E16.5, when it became barely detectable. Remarkably, the silencing of 11beta-HSD2 gene expression coincided with the onset of 11beta-HSD1 gene expression in the labyrinth at E16.5 when moderate levels of 11beta-HSD1 mRNA were detected and maintained to E18.5. By contrast, neither 11beta-HSD1 mRNA nor 11beta-HSD2 mRNA were detected in any cell types within the basal zone from E12.5 to E18.5. Moreover, the expression of 11beta-HSD1 and 11beta-HSD2 in the decidua exhibited a high degree of cell specificity in that the mRNA for both 11beta-HSD1 and 11beta-HSD2 was detected in the decidua-stroma but not in the compact decidua. A distinct pattern was also observed within the endometrium where the mRNA for 11beta-HSD1 was expressed in the epithelium, whereas that for 11beta-HSD2 was confined strictly to the stroma. By comparison, the expression of GR in the placenta and uterus was ubiquitous and unremarkable throughout late pregnancy. In conclusion, the present study demonstrates for the first time remarkable spatial and temporal patterns of expression of 11beta-HSD1 and 11beta-HSD2 and GR in the murine placenta and uterus and highlights the intricate control of not only transplacental passage of maternal glucocorticoid to the fetus but also local glucocorticoid action during late pregnancy.