Inhibition of the glucocorticoid receptor attenuates proteinuric kidney diseases in multiple species.

BACKGROUND AND HYPOTHESIS: Glucocorticoids are the treatment of choice for proteinuric patients with minimal-change disease (MCD) and primary focal and segmental glomerulosclerosis (FSGS). Immunosuppressive as well as direct effects on podocytes are believed to mediate their actions. In this study, we analyzed the anti-proteinuric effects of inhibition of the glucocorticoid receptor (GR) in glomerular epithelial cells, including podocytes. METHODS: We employed genetic and pharmacological approaches to inhibit the GR. Genetically, we used Pax8-Cre/GRfl/fl mice to specifically inactivate the GR in kidney epithelial cells. Pharmacologically, we utilized a glucocorticoid antagonist called mifepristone. RESULTS: Genetic inactivation of GR, specifically in kidney epithelial cells, using Pax8-Cre/GRfl/fl mice, ameliorated proteinuria following protein overload. We further tested the effects of pharmacological GR inhibition in three models and species: the puromycin-aminonucleoside-induced nephrosis model in rats, the protein overload model in mice and the inducible transgenic NTR/MTZ zebrafish larvae with specific and reversible podocyte injury. In all three models, both pharmacological GR activation and inhibition consistently and significantly ameliorated proteinuria. Additionally, we translated our findings to humans, where three nephrotic adult patients with MCD or primary FSGS with contraindications or insufficient responses to corticosteroids, were treated with mifepristone. This treatment resulted in a clinically relevant reduction of proteinuria. CONCLUSIONS: Thus, across multiple species and proteinuria models, both genetic and pharmacological GR inhibition was at least as effective as pronounced GR activation. While, the mechanism remains perplexing, GR inhibition may be a novel and targeted therapeutic approach to treat glomerular proteinuria potentially bypassing adverse actions of steroids.

Genomic determinants implicated in the glucocorticoid-mediated induction of KLF9 in pulmonary epithelial cells.

Ligand-activated glucocorticoid receptor (GR) elicits variable glucocorticoid-modulated transcriptomes in different cell types. However, some genes, including Krüppel-like factor 9 (KLF9), a putative transcriptional repressor, demonstrate conserved responses. We show that glucocorticoids induce KLF9 expression in the human airways in vivo and in differentiated human bronchial epithelial (HBE) cells grown at air-liquid interface (ALI). In A549 and BEAS-2B pulmonary epithelial cells, glucocorticoids induce KLF9 expression with similar kinetics to primary HBE cells in submersion culture. A549 and BEAS-2B ChIP-seq data reveal four common glucocorticoid-induced GR binding sites (GBSs). Two GBSs mapped to the 5'-proximal region relative to KLF9 transcription start site (TSS) and two occurred at distal sites. These were all confirmed in primary HBE cells. Global run-on (GRO) sequencing indicated robust enhancer RNA (eRNA) production from three of these GBSs in BEAS-2B cells. This was confirmed in A549 cells, plus submersion, and ALI culture of HBE cells. Cloning each GBS into luciferase reporters revealed glucocorticoid-induced activity requiring a glucocorticoid response element (GRE) within each distal GBS. While the proximal GBSs drove modest reporter induction by glucocorticoids, this region exhibited basal eRNA production, RNA polymerase II enrichment, and looping to the TSS, plausibly underlying constitutive KLF9 expression. Post glucocorticoid treatment, interactions between distal and proximal GBSs and the TSS correlated with KLF9 induction. CBP/P300 silencing reduced proximal GBS activity, but negligibly affected KLF9 expression. Overall, a model for glucocorticoid-mediated regulation of KLF9 involving multiple GBSs is depicted. This work unequivocally demonstrates that mechanistic insights gained from cell lines can translate to physiologically relevant systems.

New insights into the roles of glucocorticoid signaling dysregulation in pathological cardiac hypertrophy.

Pathological cardiac hypertrophy is a process of abnormal remodeling of the myocardium in response to stress overload or ischemia that results in myocardial injury, which is an independent risk factor for the increased morbidity and mortality of heart failure. Elevated circulating glucocorticoids (GCs) levels are associated with an increased risk of pathological cardiac hypertrophy, but the exact role remains unclear. In the heart, GCs exerts physiological and pharmacological effects by binding the glucocorticoid receptor (GR, NR3C1). However, under the state of tissue damage or oxidative stress, GCs can also bind the closely related mineralocorticoid receptor (MR, NR3C2) to exert a detrimental effect on cardiac function. In addition, the bioavailability of GCs at the cellular level is mainly regulated by tissue-specific metabolic enzymes 11ß-hydroxysteroid dehydrogenases (11ß-HSDs), including 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) and type 2 (11ß-HSD2), which catalyze the interconversion of active GCs. In this paper, we provide an overview of GC signaling and its physiological roles in the heart and highlight the dynamic and diverse roles of GC signaling dysregulation, mediated by excessive ligand GCs levels, GR/MR deficiency or overexpression, and local GCs metabolic disorder by 11ß-HSDs, in the pathology of cardiac hypertrophy. Our findings will provide new ideas and insights for the search for appropriate intervention targets for pathological cardiac hypertrophy.

Physiological Convergence and Antagonism Between GR and PPARγ in Inflammation and Metabolism.

Nuclear receptors (NRs) are transcription factors that modulate gene expression in a ligand-dependent manner. The ubiquitously expressed glucocorticoid receptor (GR) and peroxisome proliferator-activated receptor gamma (PPARγ) represent steroid (type I) and non-steroid (type II) classes of NRs, respectively. The diverse transcriptional and physiological outcomes of their activation are highly tissue-specific. For example, in subsets of immune cells, such as macrophages, the signaling of GR and PPARγ converges to elicit an anti-inflammatory phenotype; in contrast, in the adipose tissue, their signaling can lead to reciprocal metabolic outcomes. This review explores the cooperative and divergent outcomes of GR and PPARγ functions in different cell types and tissues, including immune cells, adipose tissue and the liver. Understanding the coordinated control of these NR pathways should advance studies in the field and potentially pave the way for developing new therapeutic approaches to exploit the GR:PPARγ crosstalk.

Ascorbic acid along with ciprofloxacin regulates S. aureus induced microglial inflammatory responses and oxidative stress through TLR-2 and glucocorticoid receptor modulation.

Microglial inflammatory responses play a central role in the pathogenesis of S. aureus induced brain infections. Upon activation, microglia produces free radicals (ROS/RNS) and disrupts the cellular antioxidant defense to combat invading microorganisms. Despite conventional antibiotic or steroid therapy, microglial over-activation could not be controlled. So, an attempt had been taken by using a natural antioxidant ascorbic acid along with ciprofloxacin to regulate microglial over-activation by involving TLR-2 and glucocorticoid receptor (GR) in an in-vitro cell culture-based study. Combinatorial treatment during TLR-2 neutralization effectively reduced the bacterial burden at 60 min compared to the GR blocking condition (p < 0.05). Moreover, the infection-induced H2O2, O2.-, and NO release in microglial cell culture was diminished possibly by enhancing SOD and catalase activities in the same condition (p < 0.05). The arginase activity was markedly increased after TLR-2 blocking in the combinatorial group compared to single treatments (p < 0.05). Experimental results indicated that combinatorial treatment may act through up-regulating GR expression by augmenting endogenous corticosterone levels. However, better bacterial clearance could further suppress the TLR-2 mediated pro-inflammatory NF-κB signaling. From Western blot analysis, it was concluded that ciprofloxacin-ascorbic acid combination in presence of anti-TLR-2 antibody exhibited 81.25% inhibition of TLR-2 expression while the inhibition for GR was 3.57% with respect to the infected group. Therefore, during TLR-2 blockade ascorbic acid combination might be responsible for the restoration of redox balance in microglia via modulating TLR-2/GR interaction. The combination treatment could play a major role in the neuroendocrine-immune regulation of S. aureus induced microglial activation.

Expression and clinicopathological significance of glucocorticoid receptor, SGK1, and NDRG1 in hormone-naïve prostate carcinoma.

Glucocorticoid receptor (GR) has been implicated in prostate carcinoma growth and progression. Glucocorticoid receptor beta (GRß) acts as an inhibitor of GR; however, its function is not well understood. Serum- and glucocorticoid-regulated kinase 1 (SGK1) is a GR-responsive gene that phosphorylates N-myc downstream-regulated gene 1 (NDRG1) and is involved in cancer growth and invasion. However, the expression of GR, GRß, SGK1, and NDRG1 in prostate cancer and their relationship with clinicopathological and functional significance remain unknown. The association between the status of GR, GRß, SGK1, and NDRG1 immunoreactivity and clinicopathological variables was analyzed in patients with prostate carcinoma to explore their clinical significance. In prostate carcinoma cases, the relative abundance of GR and NDRG1 immunoreactivity was inversely and significantly associated with the primary tumor stage (pT), while GR immunoreactivity was inversely and significantly associated with the Ki-67 score. The relative expression status of NDRG1 was significantly associated with that of GR. However, no significant correlation was observed between any of the clinicopathological parameters and GRß and SGK1 expression. Our findings indicate that GR and NDRG1 expression status is correlated with clinicopathological features in patients with prostate cancer.

Glucocorticoid receptor dimerization in the cytoplasm might be essential for nuclear localization.

The glucocorticoid receptor (GR) plays an important role in steroid-dependent regulation of metabolism, development, and the immune response in humans. Although GR is known to be activated by the binding of glucocorticoid, the mechanism of action is poorly understood. We investigated dimerization of GR in the cytoplasm and nuclear trans-localization in response to treatment with the ligand dexamethasone. GFP-tagged GR and FLAG-tagged GR were co-expressed in COS-1 cells, and cell lysates were subjected to co-immunoprecipitation assay with anti-GFP antibody to determine their dimerization. FLAG-GR was co-precipitated with GFP-GR in the cytoplasmic fraction of COS-1 cells. Treatment with the GR agonist dexamethasone significantly decreased the cytoplasmic interaction between FLAG- and GFP-GR, and significantly increased interaction of the GRs in the nuclear fraction. The two amino acids, Pro625 and Ile628 known to be located in GR-GR dimer interface, were mutated to alanine and the influence of the mutation on dimerization, ligand-dependent nuclear localization, and transcriptional activities were determined. Mutant GR showed a dramatic decrease in interaction in the cytoplasmic fraction and no detectable nuclear translocation in the presence or absence of dexamethasone. Furthermore, luciferase assays showed that mutant GR showed no detectable transcriptional activation via the GR-responsive DNA element (GRE) compared to the wild-type. Our results suggest that GR exists as a dimer in the cytoplasm and this dimerization may be essential for GRE-mediated transcriptional activation following ligand binding.

Involvement of BMP-15 in glucocorticoid actions on ovarian steroidogenesis by rat granulosa cells.

To elucidate the impact of glucocorticoids on ovarian steroidogenesis and its molecular mechanism by focusing on bone morphogenetic proteins (BMPs), we examined the effect of dexamethasone (Dex) on estradiol and progesterone synthesis by using primary culture of rat granulosa cells. It was revealed that Dex treatment dose-dependently decreased estradiol production but increased progesterone production induced by follicle-stimulating hormone (FSH) by granulosa cells. In accordance with the effects of Dex on estradiol synthesis, Dex suppressed P450arom mRNA expression and cAMP synthesis induced by FSH. Dex treatment in turn enhanced basal as well as FSH-induced levels of mRNAs encoding the enzymes for progesterone synthesis including P450scc and 3ßHSD but not StAR and 20αHSD. Of note, Dex treatment significantly upregulated transcription of the BMP target gene Id-1 and Smad1/5/9 phosphorylation in the presence of BMP-15 among the key ovarian BMP ligands. It was also found that Dex treatment increased the expression level of BMP type-I receptor ALK-6 among the type-I and -II receptors for BMP-15. Inhibitory Smad6/7 expression was not affected by Dex treatment. On the other hand, BMP-15 treatment upregulated glucocorticoid receptor (GR) expression in granulosa cells. Collectively, it was revealed that glucocorticoids elicit differential effects on ovarian steroidogenesis, in which GR and BMP-15 actions are mutually enhanced in granulosa cells.

Ginsenoside compound K- a potential drug for rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory and autoimmune disease, if prescription of effective delayed, the articular disturbances may lead to disability. Ginsenoside compound K (GCK) is the main degradation product of oral ginsenosides in the human intestine. Numerous researches in vitro and in vivo have recorded the anti-arthritic effect of GCK, we discuss the mechanisms from the following three aspects, including anti-inflammatory, immune-regulatory, and bone-protective, respectively, in this review, and the anti-arthritic mechanism of GCK may be related to the effect on TNF-α-TNFR2, glucocorticoid receptor (GR) and ß-arrestin1/2. We also describe the anti-anemia effect of GCK to open the possibility that GCK can be used as an effective disease-modifying anti-rheumatic drug (DMARD).

Dexamethasone for Severe COVID-19: How Does It Work at Cellular and Molecular Levels?

The coronavirus disease 2019 (COVID-19) caused by infection of the severe respiratory syndrome coronavirus-2 (SARS-CoV-2) significantly impacted human society. Recently, the synthetic pure glucocorticoid dexamethasone was identified as an effective compound for treatment of severe COVID-19. However, glucocorticoids are generally harmful for infectious diseases, such as bacterial sepsis and severe influenza pneumonia, which can develop respiratory failure and systemic inflammation similar to COVID-19. This apparent inconsistency suggests the presence of pathologic mechanism(s) unique to COVID-19 that renders this steroid effective. We review plausible mechanisms and advance the hypothesis that SARS-CoV-2 infection is accompanied by infected cell-specific glucocorticoid insensitivity as reported for some other viruses. This alteration in local glucocorticoid actions interferes with undesired glucocorticoid to facilitate viral replication but does not affect desired anti-inflammatory properties in non-infected organs/tissues. We postulate that the virus coincidentally causes glucocorticoid insensitivity in the process of modulating host cell activities for promoting its replication in infected cells. We explore this tenet focusing on SARS-CoV-2-encoding proteins and potential molecular mechanisms supporting this hypothetical glucocorticoid insensitivity unique to COVID-19 but not characteristic of other life-threatening viral diseases, probably due to a difference in specific virally-encoded molecules and host cell activities modulated by them.

Rapid Screening of Glucocorticoid Receptor (GR) Effectors Using Cortisol-Detecting Sensor Cells.

Cortisol, a stress hormone, plays key roles in mediating stress and anti-inflammatory responses. As abnormal cortisol levels can induce various adverse effects, screening cortisol and cortisol analogues is important for monitoring stress levels and for identifying drug candidates. A novel cell-based sensing system was adopted for rapid screening of cortisol and its functional analogues under complex cellular regulation. We used glucocorticoid receptor (GR) fused to a split intein which reconstituted with the counterpart to trigger conditional protein splicing (CPS) in the presence of targets. CPS generates functional signal peptides which promptly translocate the fluorescent cargo. The sensor cells exhibited exceptional performance in discriminating between the functional and structural analogues of cortisol with improved sensitivity. Essential oil extracts with stress relief activity were screened using the sensor cells to identify GR effectors. The sensor cells responded to peppermint oil, and L-limonene and L-menthol were identified as potential GR effectors from the major components of peppermint oil. Further analysis indicated L-limonene as a selective GR agonist (SEGRA) which is a potential anti-inflammatory agent as it attenuates proinflammatory responses without causing notable adverse effects of GR agonists.

Inhibition of expression of glucocorticoids receptors may contribute to postpartum depression.

Although postpartum depression (PPD) is the leading cause of disability worldwide, its molecular mechanisms are poorly understood. Recent evidence has suggested that impaired glucocorticoid receptor (GR), the signaling of key molecules of the HPA axis, plays a key role in the behavioral and neuroendorcrine alterations of major depression. However, the role of GR in postpartum period, which following with the abrupt withdrawal of placental corticotropin releasing hormone (CRH) and resulting in a re-equilibration of the maternal HPA axis in the days of post-delivery, is still not entirely clear. Previously, a hormone-simulated pregnancy (HSP), and the subsequent 'postpartum' withdrawal in estrogen has been employed to mimic the fluctuations in estradiol associated with pregnancy and postpartum. Using the HSP model, we investigated here the effect of 'postpartum' withdrawal in estrogen as well as depression- and anxiety-like behavior by intra-hippocampal infusion with GR inhibitor-RU486. Following the successful acquisition of PPD model by withdrawal in estrogen, reduced GR expression was observed in hippocampus. Further, HSP-rats suffered intra-hippocampal RU486 infusion presented depression- and anxiety-like behavior as postpartum depression. Together, these results suggest an important, though complex, role for GR in the behavioral regulation of postpartum depression.

MLL2 regulates glucocorticoid receptor-mediated transcription of ENACα in human retinal pigment epithelial cells.

Glucocorticoids require the glucocorticoid receptor (GR), a type of ligand-dependent nuclear receptor to transmit their downstream effects. Upon glucocorticoid binding, GR associates with glucocorticoid response elements (GREs) and recruits other transcriptional coregulators to activate or repress target gene transcription. Many SET-domain family proteins have been demonstrated to contribute to GR-mediated transcriptional activity. However, whether histone H3K4-specific methyltransferase plays a cell-type-specific role in GR transcriptional regulation remains poorly understood. In this report, we examined MLL2 (KMT2D), a histone-lysine methyltransferase that catalyzes histone H3 lysine 4 methylation (H3K4me). Furthermore, we demonstrated that MLL2 specifically regulates the transcription of some GR target genes (e.g., ENACα and FLJ20371) in ARPE-19 cells, but has no effect in A549 cells. Mechanistically, co-immunoprecipitation assays revealed that MLL2 is associated with GR in a ligand-independent manner in APRE-19 cells. Moreover, chromatin immunoprecipitation analyses demonstrated that MLL2 could co-occupy glucocorticoid response elements (GREs) of GR target genes along with GR following Dex stimulation. Finally, the FAIRE-qPCR results illustrated that MLL2 is pivotal in establishing chromatin structure accessibility at the GREs of ARPE-19 specific genes in the presence of Dex. Taken together, our study determined that MLL2 regulates GR-mediated transcription in a cell-type-specific manner, and we provide a molecular mechanism to explain the specific role of MLL2 in regulating GR target gene expression in ARPE-19 cells.

Dexamethasone along with ciprofloxacin modulates S. aureus induced microglial inflammation via glucocorticoid (GC)-GC receptor-mediated pathway.

Microglial inflammation is the hallmark of S. aureus induced brain abscesses. Conventional antibiotic therapy could not regulate inflammation and the use of steroids in CNS infection remained controversial. To address this issue the effect of dexamethasone along with ciprofloxacin on microglial inflammation has been attempted both in glucocorticoid receptor (GR) opened and blocked condition. We have investigated the effects of ciprofloxacin (0.24 µg/ml, pre-treatment) and dexamethasone (150 nM, pre-treatment) in combination with murine microglia infected with S. aureus for 30, 60 and 90 min by either keeping GR opened or blocked with GR antagonist RU486. Alterations in cellular motility, intracellular killing assay, free radical production, antioxidant enzyme activities, corticosterone, and cytokine levels were determined. The expressions of TLR-2, GR, and other inflammatory markers were determined in terms of this combinatorial treatment. Combination treatment significantly (p < 0.05) reduced the bacterial burden of microglia only when GR remained open and effectively suppressed S. aureus induced oxidative stress by augmenting SOD and catalase enzyme activity and suppressing other pro-inflammatory markers at 90 min. Arginase activity, a critical determinant of microglial polarization was found to be higher after treatment at 60 and 90 min. This situation was reversed when this combination treatment was applied by keeping GR blocked using GR antagonist RU486. Therefore, it can be concluded that combination treatment of ciprofloxacin and dexamethasone could regulate S. aureus induced microglial activation, in the presence of functional GR via utilizing glucocorticoid (GC)-GR pathway and ultimately confers protection to the host from brain inflammation.

[Effect of Puyu Capsules on behavior,HPA axis and CREB-BDNF pathway expression in hippocampus of depressed mice].

This study aimed to investigate the antidepressant effects of Puyu Capsules and its potential mechanism. The antidepressant activity of Puyu Capsules was evaluated by forced swimming test(FST) and tail suspension test(TST) after subchronic administration in mice. Next, the mice were subjected to a chronic unpredictable stress(CUS) protocol for a period of 28 d to induce depressive-like behaviors. Then, a sucrose preference test, open-field test and novelty-suppressed feeding test were performed to evaluate the antidepressant effect of Puyu Capsules. After the behavioral test, the adrenal index was calculated; the levels of serum corticosterone(CORT) and adrenocorticotropic hormone(ACTH) were detected by enzyme-linked immunosorbent assay(ELISA); the levels of glucocorticoid receptor(GR), protein expression of brain-derived neurotrophic factor(BDNF), and the ratio of phosphorylated cAMP response element binding protein(CREB) to total CREB were detected by Western blot to explore the antidepressant function and mechanism of Puyu Capsules. The results suggested that Puyu Capsules had significant antidepressant effects on both the depression model and CUS model. At the same time, the drug could prevent the change of adrenal index induced by CUS and reverse the abnormal activation of CORT and ACTH in the serum of depressed mice. Finally, Puyu Capsules could also reverse the lower expression of pCREB, BDNF and GR in the hippocampus of CUS mice. In conclusion, Puyu Capsules produced significant antidepressant effects, and the mechanism was closely related to hypothalamic pituitary adrenal(HPA) axis activity, GR and CREB-BDNF pathway expression.

Placental glucocorticoid receptor and 11ß-hydroxysteroid dehydrogenase-2 recruitment indicates impact of prenatal adversity upon postnatal development in mice.

Prenatal stress may increase concentrations of maternal glucocorticoids, which restrict fetal growth, with variable impact upon postnatal development. Among key regulators of stress hormone effects are the glucocorticoid receptor (GR) and 11ß-hydroxysteroid dehydrogenase-2 (11ßHSD2), the enzyme that inactivates glucocorticoid. This study utilized mice selectively bred for social dominance (Dom) or submissiveness (Sub), respectively exhibiting resilience or sensitivity to stress, to test whether stress-induced alterations in placental GR and 11ßHSD2 protein expression may mediate divergent effects of prenatal adversity upon postnatal development. Pregnant Dom and Sub dams underwent prenatal restraint stress (PRS) for 45 min on gestational days (GD) 15-17. PRS induced a similar spike in serum corticosterone concentrations of dams from each strain on GD15 (p < .001, n = 8), and impaired fetal growth (p < .01, n = 5 litters), although Dom placentae were larger than Sub placentae (p < .01). Among placentae from Dom dams, PRS elevated protein contents of both GR (p < .05, n = 5 litters) and 11ßHSD2 (p < .01) on GD19. In contrast, GR contents were reduced among placentae from PRS-exposed Sub mice (p < .01), without changes in 11ßHSD2 content. Correspondingly, Dom PRS pup growth recovered by PND14, yet Sub PRS pups remained underweight into adolescence (p < .0001, n = 40 pups). Thus, prenatal stress more strongly increased placental GR and 11ßHSD2 levels among Dom mice than in Subs. Increased GR may improve placental function and up-regulate 11ßHSD2 expression, protecting fetuses from effects of prenatal stress upon postnatal development. Placental recruitment of GR and 11ßHSD2 are potential markers of stress-induced developmental disorders, in accordance with maternal resilience or sensitivity to stress.

False responses of Renilla luciferase reporter control to nuclear receptor TR4.

Renilla luciferase reporter is a widely used internal control in dual luciferase reporter assay system, where its transcription is driven by a constitutively active promoter. However, the authenticity of the Renilla luciferase response in some experimental settings has recently been questioned. Testicular receptor 4 (TR4, also known as NR2C2) belongs to the subfamily 2 of nuclear receptors. TR4 binds to a direct repeat regulatory element in the promoter of a variety of target genes and plays a key role in tumorigenesis, lipoprotein regulation, and central nervous system development. In our experimental system using murine pituitary corticotroph tumor AtT20 cells to investigate TR4 actions on POMC transcription, we found that overexpression of TR4 resulted in reduced Renilla luciferase expression whereas knockdown TR4 increased Renilla luciferase expression. The TR4 inhibitory effect was mediated by the TR4 DNA-binding domain and behaved similarly to the GR and its agonist, Dexamethasone. We further demonstrated that the chimeric intron, commonly present in various Renilla plasmid backbones such as pRL-Null, pRL-SV40, and pRL-TK, was responsible for TR4's inhibitory effect. The results suggest that an intron-free Renilla luciferase reporter may provide a satisfactory internal control for TR4 at certain dose range. Our findings advocate caution on the use of Renilla luciferase as an internal control in TR4-directed studies to avoid misleading data interpretation.

Role of spliceosome proteins in the regulation of glucocorticoid receptor isoforms by cortisol and dehydroepiandrosterone.

Dehydroepiandrosterone (DHEA) can counteract the activity of cortisol by modulating the glucocorticoid receptor ß (GRß) expression and antagonizing the binding of GRα to the glucocorticoid responsive element (GRE) in RACK1 (Receptor for Activated C Kinase 1) promoter. These observations are important in the context of immunosenescence and can be extended to recognize a complex hormonal balance in the control of GR isoform expression and consequently in the expression of GR responsive genes. To elucidate the mechanism of DHEA on GR alternative splicing, we investigated its possible involvement in the expression of proteins such as the Serine/arginine (SR)-Rich Splicing Factors (SRSF) regulating GR splicing, specifically SRSF9 and SRSF3 also known as SRp30c and SRp20 respectively. We demonstrated that DHEA can induce the up-regulation of GR mRNA which is preferentially directed toward the ß isoform. The effect is due to an increase in expression of the splicing factor SRSF9. On the other hand cortisol up-regulated SRSF3, the splicing factor promoting GRα isoform. We demonstrated that DHEA and cortisol modulate SRSF9 and SRSF3 in a different way and our data suggest that the anti-glucocorticoid effect of DHEA, among other mechanisms, is also exerted by modulating the expression of proteins involved in the splicing of the GR pre-mRNA.

Update on molecular mechanisms of corticosteroid resistance in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is an inflammatory and irreversible pulmonary disorder that is characterized by inflammation and airway destruction. In recent years, COPD has become a global epidemic due to increased air pollution and exposure to cigarette smoke. Current therapeutics using bronchiodialator and anti-inflammatory corticosteroids are most widely used for all patients with persistent COPD, but these approaches are disappointing due to limited improvement in symptom control and survival rate. More importantly, a certain number of COPD patients are resistant to the corticosteroid treatment and their symptoms worsen. Therefore, more effective anti-inflammatory drugs and combinational treatment are required. Understanding of the underlying molecular and immunological mechanisms is critical to developing new therapeutics. Lung inflammation and the released pro-inflammatory cytokines affect glucocorticoid receptor (GR), histone deacetylase 2 (HDAC2) and surfactant protein D (SP-D) activities in many cell types. Macrophages, neutrophils, airway epithelial cells and lymphocytes are involved in the induction of corticosteroid resistance. This review updated the recent advances in molecular and immunological mechanisms of steroid resistance among patients and animal models with COPD. Meanwhile we discussed novel therapeutic approaches in controlling lung inflammation and improving corticosteroid sensitivity among the steroid resistant patients with COPD.

Glucocorticoids facilitate the transcription from the human cytomegalovirus major immediate early promoter in glucocorticoid receptor- and nuclear factor-I-like protein-dependent manner.

Human cytomegalovirus (HCMV) is a common and usually asymptomatic virus agent in healthy individuals. Initiation of HCMV productive infection depends on expression of the major immediate early (MIE) genes. The transcription of HCMV MIE genes is regulated by a diverse set of transcription factors. It was previously reported that productive HCMV infection is triggered probably by elevation of the plasma hydroxycorticoid level. However, it is poorly understood whether the transcription of MIE genes is directly regulated by glucocorticoid. Here, we found that the dexamethasone (DEX), a synthetic glucocorticoid, facilitates the transcription of HCMV MIE genes through the MIE promoter and enhancer in a glucocorticoid receptor (GR)-dependent manner. By competitive EMSA and reporter assays, we revealed that an NF-I like protein is involved in DEX-mediated transcriptional activation of the MIE promoter. Thus, this study supports a notion that the increased level of hydroxycorticoid in the third trimester of pregnancy reactivates HCMV virus production from the latent state.