Glucocorticoid Receptor Transcriptional Activity Research Articles

Characterization of a new selective glucocorticoid receptor modulator with anorexigenic activity

Obesity, a worldwide epidemic, leads to various metabolic disorders threatening human health. In response to stress or fasting, glucocorticoid (GC) levels are elevated to promote food intake. This involves GC-induced expression of the orexigenic neuropeptides in agouti-related protein (AgRP) neurons of the hypothalamic arcuate nucleus (ARC) via the GC receptor (GR). Here, we report a selective GR modulator (SGRM) that suppresses GR-induced transcription of genes with non-classical glucocorticoid response elements (GREs) such as Agrp-GRE, but not with classical GREs, and via this way may serve as a novel anti-obesity agent. We have identified a novel SGRM, 2-O-trans-p-coumaroylalphitolic acid (Zj7), a triterpenoid extracted from the Ziziphus jujube plant, that selectively suppresses GR transcriptional activity in Agrp-GRE without affecting classical GREs. Zj7 reduces the expression of orexigenic genes in the ARC and exerts a significant anorexigenic effect with weight loss in both high fat diet-induced obese and genetically obese db/db mouse models. Transcriptome analysis showed that Zj7 represses the expression of a group of orexigenic genes including Agrp and Npy induced by the synthetic GR ligand dexamethasone (Dex) in the hypothalamus. Taken together, Zj7, as a selective GR modulator, showed beneficial metabolic activities, in part by suppressing GR activity in non-classical GREs in orexigenic genes. This study demonstrates that a potential anorexigenic molecule may allow GRE-specific inhibition of GR transcriptional activity, which is a promising approach for the treatment of metabolic disorders.

Selective Modulation of the Human Glucocorticoid Receptor Compromises GR Chromatin Occupancy and Recruitment of p300/CBP and the Mediator Complex

Exogenous glucocorticoids are frequently used to treat inflammatory disorders and as adjuncts for the treatment of solid cancers. However, their use is associated with severe side effects and therapy resistance. Novel glucocorticoid receptor (GR) ligands with a patient-validated reduced side effect profile have not yet reached the clinic. GR is a member of the nuclear receptor family of transcription factors and heavily relies on interactions with coregulator proteins for its transcriptional activity. To elucidate the role of the GR interactome in the differential transcriptional activity of GR following treatment with the selective GR agonist and modulator dagrocorat compared to classic (ant)agonists, we generated comprehensive interactome maps by high-confidence proximity proteomics in lung epithelial carcinoma cells. We found that dagrocorat and the antagonist RU486 both reduced GR interaction with CREB-binding protein/p300 and the mediator complex compared to the full GR agonist dexamethasone. Chromatin immunoprecipitation assays revealed that these changes in GR interactome were accompanied by reduced GR chromatin occupancy with dagrocorat and RU486. Our data offer new insights into the role of differential coregulator recruitment in shaping ligand-specific GR-mediated transcriptional responses.

Glucocorticoid Receptor (GR) Activation Is Associated with Increased cAMP/PKA Signaling in Castration-Resistant Prostate Cancer.

In castration-resistant prostate cancer (CRPC), increased glucocorticoid receptor (GR) expression and ensuing transcriptional activity have been proposed as an oncogenic "bypass" mechanism in response to androgen receptor (AR) signaling inhibition (ARSi). Here, we report that GR transcriptional activity acquired following ARSi is associated with the upregulation of cyclic adenosine monophosphate (cAMP)-associated gene expression pathways in both model systems and metastatic prostate cancer patient samples. In the context of ARSi, the expression of GR-mediated genes encoding cAMP signaling pathway-associated proteins can be inhibited by treatment with selective GR modulators (SGRMs). For example, in the context of ARSi, we found that GR activation resulted in upregulation of protein kinase inhibitor beta (PKIB) mRNA and protein levels, leading to nuclear accumulation of the cAMP-dependent protein kinase A catalytic subunit (PKA-c). Increased PKA-c, in turn, is associated with increased cAMP response element-binding protein phosphorylation and activity. Furthermore, enzalutamide and SGRM combination therapy in mice bearing CRPC xenografts delayed CRPC progression compared with enzalutamide therapy alone, and reduced tumor PKIB mRNA expression. Supporting the clinical importance of GR/PKA signaling activation in CRPC, we found a significant enrichment of both cAMP pathway signaling-associated gene expression and high NR3C1 (GR) activity in patient-derived xenograft models and metastatic human CRPC samples. These findings suggest a novel mechanism linking CRPC-induced GR transcriptional activity with increased cAMP signaling in AR-antagonized CRPC. Furthermore, our findings suggest that GR-specific modulation in addition to AR antagonism may delay GR+ CRPC time to recurrence, at least in part, by inhibiting tumor cAMP/PKA pathways.

Chromatin accessibility and pioneer factor FOXA1 restrict glucocorticoid receptor action in prostate cancer.

Treatment of prostate cancer relies predominantly on the inhibition of androgen receptor (AR) signaling. Despite the initial effectiveness of the antiandrogen therapies, the cancer often develops resistance to the AR blockade. One mechanism of the resistance is glucocorticoid receptor (GR)-mediated replacement of AR function. Nevertheless, the mechanistic ways and means how the GR-mediated antiandrogen resistance occurs have remained elusive. Here, we have discovered several crucial features of GR action in prostate cancer cells through genome-wide techniques. We detected that the replacement of AR by GR in enzalutamide-exposed prostate cancer cells occurs almost exclusively at pre-accessible chromatin sites displaying FOXA1 occupancy. Counterintuitively to the classical pioneer factor model, silencing of FOXA1 potentiated the chromatin binding and transcriptional activity of GR. This was attributed to FOXA1-mediated repression of the NR3C1 (gene encoding GR) expression via the corepressor TLE3. Moreover, the small-molecule inhibition of coactivator p300's enzymatic activity efficiently restricted GR-mediated gene regulation and cell proliferation. Overall, we identified chromatin pre-accessibility and FOXA1-mediated repression as important regulators of GR action in prostate cancer, pointing out new avenues to oppose steroid receptor-mediated antiandrogen resistance.

The combination of Ephedrae herba and coixol from Coicis semen attenuate adiposity via glucocorticoid receptor regulation

The enhanced therapeutic effects and mechanisms of certain herbal combination in various herbal prescriptions are mostly unclear. A combination of two herbs, namely Ephedrae herba (EH) and Coicis semen (CS), has been commonly prescribed for obesity. In our previous work, the combination of EH and CS was studied using network pharmacological approach to predict its pharmacological targets and in vitro experiments to evaluate its efficacy on obesity. Although we demonstrated enhanced anti-adiposity effects of the combination on matured adipocytes, the molecular mechanisms and contributing compounds underlying the effects of EH-CS combination on adiposity or adipogenesis were not fully elucidated. The current study adopted integrated bioinformatics analysis to precisely validate potential targets of EH-CS by screening differentially expressed genes (DEGs) of morbid obesity patients from NCBI gene expression omnibus (GEO). Based on the functional cluster analysis of down-regulated DEGs, the anti-adipogenesis mechanism of EH-CS combination was speculated with KEGG enrichment analysis. Furthermore, we investigated the combinational effects of EH and coixol, or stigmasterol, the two compounds in CS which were expected to have main beneficial effects in metabolic diseases. Moreover, distinct effect of the combination on transcriptional activity of glucocorticoid receptor (GR) was investigated using electrophoretic mobility shift assay (EMSA). The EH-CS combination was predicted to modulate down-regulated genes which are involved in KEGG pathways crucial to metabolic disease in morbidly obese individuals. The combination of EH with CS compounds significantly increased the phosphorylation of acetyl-coA carboxylase (ACC), AMP-activated protein kinase (AMPK), and protein kinase B (AKT) in 3T3-L1 cells and decreased intracellular lipid accumulation. The two CS compounds significantly increased the anti-adipogenesis/lipogenesis effects of EH by inhibiting the gene expression levels. Finally, the combination of EH and coixol inhibited dexamethasone-induced GR translocation to the nucleus and transcriptional binding activity in adipocytes. The combination of EH and CS could be considered a therapeutic strategy for treating metabolic diseases, including obesity.

Ginsenoside compound K exerts anti-inflammatory effects through transcriptional activation and transcriptional inhibition of glucocorticoid receptor in rheumatoid arthritis fibroblast-like synoviocytes

Ginsenoside compound K (GCK) has anti-inflammatory and immunoregulatory effects, and glucocorticoid receptor (GR) has been considered as its potential target. But the mechanism by which GCK exerts its anti-inflammatory effects after GR activation remains unclear. In this study, molecular docking, isothermal titration calorimetry, siRNA of GR and GRA458T mutation were used to confirm the anti-inflammatory mechanism of GCK targeting GR in fibroblast-like synoviocytes (FLS). The results showed that the key binding sites of GR and GCK were identified as ASN564, MET560 and ASN638, with binding levels at the μm level. In addition, the inhibitory effect of GCK on the proliferation of FLS and the secretion of inflammatory cytokines (IL-6, IL-8, and IL-1β) were mediated by transcriptional activation of GR, but on the migration, invasion, and TNF-α secretion of FLS were mediated by transcriptional inhibition of GR. These actions exert anti-inflammatory effects through indirect and direct inhibition of NF-κB transcriptional activity, respectively. In conclusion, this study elucidates that GCK can directly bind to and activate GR. Furthermore, after activation, GR mediates the anti-inflammatory effects of GCK through two mechanisms: transcriptional activation and transcriptional inhibition.

Impaired glucocorticoid receptor function attenuates herpes simplex virus 1 production during explant-induced reactivation from latency in female mice.

A correlation exists between stress and increased episodes of human alpha-herpes virus 1 reactivation from latency. Stress increases corticosteroid levels; consequently, the glucocorticoid receptor (GR) is activated. Recent studies concluded that a GR agonist, but not an antagonist, accelerates productive infection and reactivation from latency. Furthermore, GR and certain stress-induced transcription factors cooperatively transactivate promoters that drive the expression of infected cell protein 0 (ICP0), ICP4, and VP16. This study revealed female mice expressing a GR containing a serine to alanine mutation at position 229 (GRS229A) shed significantly lower levels of infectious virus during explant-induced reactivation compared to male GRS229A or wild-type parental C57BL/6 mice. Furthermore, female GRS229A mice contained fewer VP16 + TG neurons compared to male GRS229A mice or wild-type mice during the early stages of explant-induced reactivation from latency. Collectively, these studies revealed that GR transcriptional activity has female-specific effects, whereas male mice can compensate for the loss of GR transcriptional activation.

OR22-04 The Oncogenic Action Of Glucocorticoid Receptor In Prostate Cancer

Abstract Disclosure: V. Paakinaho: None. Steroid receptors, such as androgen (AR), and glucocorticoid receptor (GR), are hormone-controlled transcription factors (TFs) that regulate transcription in multiple central cellular systems. The action of steroid receptors has a major role in the development and progression diseases, especially endocrine-related cancers, such as prostate cancer. Since AR is the key oncogenic TF driving the development and progression of prostate cancer, the receptor is the major precision drug target in the disease. Therefore, antiandrogen treatment, i.e., the inhibition of AR activity, is a common form of cancer therapy. Despite the initial effectiveness of the therapy, antiandrogen resistance often occurs and there is no curable treatment in many cases. In contrast to AR, GR has been thought to possess tumor suppressive properties in therapy-naïve prostate cancer. Nonetheless, GR can hijack the oncogenic role of AR in prostate cancer by replacing the antiandrogen inactivated AR thus mediating antiandrogen-resistance and becoming the driver of the disease. Yet, the genome-wide effects of GR in prostate cancer have largely remained elusive. The tumor suppressive versus the oncogenic consequences of GR activation are important to acknowledge since prostate cancer patients regularly receive glucocorticoids to reduce inflammation and alleviate therapy-related side effects. To unveil these issues, we integrated ChIP-seq, RNA-seq, ATAC-seq data and uncover several pivotal features of GR action in prostate cancer cells. We detected that the replacement of AR by GR in antiandrogen-exposed prostate cancer cells occurs almost exclusively at pre-accessible chromatin sites displaying the occupancy of FOXA1. Counterintuitively to the classical pioneer factor model, the silencing of FOXA1 potentiated the chromatin binding and transcriptional activity of GR. This was attributed to FOXA1-mediated repression of the NR3C1 (gene encoding GR) expression via the corepressor TLE3. Additionally, we discovered that the chromatin binding of GR as well as transcriptional regulation by the receptor differs between different prostate cancer subtypes. Moreover, GR activity in normal prostate cells diverges drastically from that of the cancer cells. In sum, our data highlight the impact FOXA1 harbors in GR-mediated antiandrogen-resistant prostate cancer. Furthermore, the distinct GR transcriptional activity in prostate cancer subtypes could potentially be used to determine if glucocorticoid exposure possess potential harm for the prostate cancer patient. Presentation: Saturday, June 17, 2023

SAT016 Small Compounds Modulate Glucocorticoid Receptor Function By Binding To A Novel Allosteric Pocket

Abstract Disclosure: A. Jimenez-Panizo: None. The glucocorticoid receptor (GR) is a ubiquitously expressed transcription factor that controls metabolic and homeostatic processes essential for life. This receptor exerts its functions by binding to corticosteroids through its ligand-binding domain (LBD), which harbors a hydrophobic pocket where ligands are encapsulated. Synthetic glucocorticoids, as Dexamethasone, are widely used in the clinic for the treatment of chronic inflammatory diseases. Unfortunately, their prolonged use is associated with critical side effects and most of the mechanisms underlying these collateral issues are still unknown. In this work, we present a deep biochemical and biophysical analysis showing how small molecules are able to bind a new allosteric pocket in solution and how these compounds can modulate GR transcriptional activity in cells. These findings can be essential to further understand GR signaling and how to better design synthetic corticosteroids to avoid side effects. Presentation: Saturday, June 17, 2023

SAT010 Role Of Glucocorticoids In Induction Of Autophagy During Fetal Lung Development

Abstract Disclosure: R. Mishra: None. Development of the fetal lung and production of pulmonary surfactant is a complex process that involves crosstalk between the mesenchyme and epithelium, branching morphogenesis, thinning of the bronchi alveolar ducts and alveolar septa, differentiation of alveolar type II cells, increased synthesis of surfactant lipids and proteins and their trafficking to lamellar bodies (LBs). The regulation of these processes is incompletely understood. Autophagy has recently been shown to be critical for lung morphogenesis and lamellar body biogenesis. In studies of global and lung epithelial cell-specific knockout of a number of autophagy-related genes in mice, there was found to be neonatal respiratory distress and lethality at birth. Interestingly, the lung phenotype of mice with an epithelial cell-specific knockout of the autophagy gene, Beclin 1 (Becn1), was very similar to that of mice with a targeted deletion of the glucocorticoid receptor (GR) gene (GRKO). Moreover, this phenotype was also observed in mice double-deficient in steroid receptor coactivators (Src)-1 and 2 (Src-1/-2dKO), which serve an essential role in GR transcriptional activity in fetal lung. Therefore we were interested to determine whether GCs acting through the GR regulate autophagy gene expression in fetal lung. In preliminary studies, we made the exciting discovery that nuclear levels of phospho (p)-GRS220 increased markedly in mouse fetal lung (MFL) at 16.5 dpc (compared to 15.5 dpc) and continued to increase toward term (18.5 dpc). Importantly, nuclear levels of total GR remained unchanged from 15.5 -18.5 dpc. Moreover, we observed that treatment of cultured MFL epithelial cells with the synthetic glucocorticoid, dexamethasone (Dex), upregulated expression of the autophagy genes, Becn1, Atg7 and Lc3b. Dex Induction of autophagy genes was prevented by co-incubation with the potent small-molecule inhibitor, SI-2, which selectively reduces cellular protein levels of the SRC family of GR coactivators. The importance of SRC-1 and SRC-2 in the regulation of autophagy gene expression is further emphasized by our finding that Becn1 and Atg7 expression was significantly reduced in lungs of Src-1/-2 dKO fetal mice at term. Previously, we found that glucocorticoids increase expression of transcription factor, C/EBPβ in an SRC-1/SRC-2-dependent manner which further upregulates C/EBPα in MFL epithelial cells. In ChIP-qPCR studies, we observed an increased binding of p- GRS220 and C/EBPα to the promoter regions of autophagy genes in MFL with advancing gestation. We also observed that siRNA-mediated knockdown of C/EBPα prevented Dex induction of autophagy gene expression. These exciting collective findings suggest that glucocorticoids together with SRC-1 and SRC-2 enhance autophagic flux in fetal lung via the C/EBP signaling pathway. Presentation: Saturday, June 17, 2023

Influence of dexamethasone on the interaction between glucocorticoid receptor and SOX9: A molecular dynamics study

The glucocorticoid receptor (GR) is a nuclear receptor that controls critical biological processes by regulating the transcription of specific genes. GR transcriptional activity is modulated by a series of ligands and coenzymes, where a ligand can act as an agonist or antagonist. GR agonists, such as the glucocorticoids dexamethasone (DEX) and prednisolone, are widely prescribed to patients with inflammatory and autoimmune diseases. DEX is also used to induce osteogenic differentiation in vitro. Recently, it has been highlighted that DEX induces changes in the osteogenic differentiation of human mesenchymal stromal cells by downregulating the transcription factor SRY-box transcription factor 9 (SOX9) and upregulating the peroxisome proliferator-activated receptor γ (PPARG). SOX9 is fundamental in the control of chondrogenesis, but also in osteogenesis by acting as a dominant-negative of RUNX2. Many processes remain to be clarified during cell fate determination, such as the interplay between the key transcription factors. The main objective pursued by this work is to shed light on the interaction between GR and SOX9 in the presence and absence of DEX at an atomic level of resolution using molecular dynamics simulations. The outcome of this research could help the understanding of possible molecular interactions between GR and SOX9 and their role in the determination of cell fate. The results highlight the key residues at the interface between GR and SOX9 involved in the complexation process and shed light on the mechanism through which DEX modulates GR-SOX9 binding and exerts its biological activity.

REDD1 (regulated in development and DNA damage1) modulates the glucocorticoid receptor function in keratinocytes.

Glucocorticoids (GCs) are widely used for the treatment of inflammatory skin diseases despite significant adverse effects including skin atrophy. Effects of GCs are mediated by the glucocorticoid receptor (GR), a well-known transcription factor. Previously, we discovered that one of the GR target genes, REDD1, is causatively involved in skin atrophy. Here, we investigated its role in GR function using HaCaT REDD1 knockout (KO) keratinocytes. We found large differences in transcriptome of REDD1 KO and control Cas9 cells in response to glucocorticoid fluocinolone acetonide (FA): both the scope and amplitude of response were significantly decreased in REDD1 KO. The status of REDD1 did not affect GR stability/degradation during self-desensitization, and major steps in GR activation-its nuclear import and phosphorylation at activating Ser211. However, the amount of GR phosphorylated at Ser226 that may play negative role in GR signalling, was increased in the nuclei of REDD1 KO cells. GR nuclear import and transcriptional activity also depend on the composition of GR chaperone complex: exchange of chaperone FKBP51 (FK506-binding protein 5) for FKBP52 (FK506-binding protein 4) being a necessary step in GR activation. We found the increased expression and abnormal nuclear translocation of FKBP51 in both untreated and FA-treated REDD1 KO cells. Overall, our results suggest the existence of a feed-forward loop in GR signalling mediated by its target gene REDD1, which has translational potential for the development of safer GR-targeted therapies.

RNA binding by the glucocorticoid receptor attenuates dexamethasone-induced gene activation

The glucocorticoid receptor (GR) is a ligand-activated transcription factor that regulates a suite of genes through direct binding of GR to specific DNA promoter elements. GR also interacts with RNA, but the function of this RNA-binding activity remains elusive. Current models speculate that RNA could repress the transcriptional activity of GR. To investigate the function of the GR-RNA interaction on GR’s transcriptional activity, we generated cells that stably express a mutant of GR with reduced RNA binding affinity and treated the cells with the GR agonist dexamethasone. Changes in the dexamethasone-driven transcriptome were quantified using 4-thiouridine labeling of RNAs followed by high-throughput sequencing. We find that while many genes are unaffected, GR-RNA binding is repressive for specific subsets of genes in both dexamethasone-dependent and independent contexts. Genes that are dexamethasone-dependent are activated directly by chromatin-bound GR, suggesting a competition-based repression mechanism in which increasing local concentrations of RNA may compete with DNA for binding to GR at sites of transcription. Unexpectedly, genes that are dexamethasone-independent instead display a localization to specific chromosomal regions, which points to changes in chromatin accessibility or architecture. These results show that RNA binding plays a fundamental role in regulating GR function and highlights potential functions for transcription factor-RNA interactions.

B2 SINE RNA as a novel regulator of glucocorticoid receptor transcriptional activity

Glucocorticoids are a key component to the cellular response to stress. Glucocorticoids act via glucocorticoid receptors found ubiquitously in the brain and body. Glucocorticoid receptors can bind to response elements throughout the genome to elicit changes in transcription, an adaptation observed at the cellular level. Yet, the transcriptional changes as a consequence of glucocorticoid receptor activation are variable across brain regions, stress conditions and recurrent bouts of glucocorticoid exposure. Here we describe a non-coding RNA, B2 SINE, which is regulated by glucocorticoids and can in turn regulate glucocorticoid receptor transcriptional activity. We show that activated glucocorticoid receptors interact directly with B2 SINE RNA via a decoy response element contained within the transcript sequence and alter receptor binding to response elements in the genome and, subsequently, changes in loci expression.

Isosteviol Sodium (STVNA) Reduces Pro-Inflammatory Cytokine IL-6 and GM-CSF in an In Vitro Murine Stroke Model of the Blood-Brain Barrier (BBB).

Early treatment with glucocorticoids could help reduce both cytotoxic and vasogenic edema, leading to improved clinical outcome after stroke. In our previous study, isosteviol sodium (STVNA) demonstrated neuroprotective effects in an in vitro stroke model, which utilizes oxygen-glucose deprivation (OGD). Herein, we tested the hypothesis that STVNA can activate glucocorticoid receptor (GR) transcriptional activity in brain microvascular endothelial cells (BMECs) as previously published for T cells. STVNA exhibited no effects on transcriptional activation of the glucocorticoid receptor, contrary to previous reports in Jurkat cells. However, similar to dexamethasone, STVNA inhibited inflammatory marker IL-6 as well as granulocyte-macrophage colony-stimulating factor (GM-CSF) secretion. Based on these results, STVNA proves to be beneficial as a possible prevention and treatment modality for brain ischemia-reperfusion injury-induced blood–brain barrier (BBB) dysfunction.

Phosphodiesterase 5 inhibitor mirodenafil ameliorates Alzheimer-like pathology and symptoms by multimodal actions

BackgroundAlzheimer’s disease (AD) pathology is associated with complex interactions among multiple factors, involving an intertwined network of various signaling pathways. The polypharmacological approach is an emerging therapeutic strategy that has been proposed to overcome the multifactorial nature of AD by targeting multiple pathophysiological factors including amyloid-β (Aβ) and phosphorylated tau. We evaluated a blood-brain barrier penetrating phosphodiesterase 5 (PDE5) inhibitor, mirodenafil (5-ethyl-2-7-n-propyl-3,5-dihydrro-4H-pyrrolo[3,2-d]pyrimidin-4-one), for its therapeutic effects on AD with polypharmacological properties.MethodsTo evaluate the potential of mirodenafil as a disease-modifying AD agent, mirodenafil was administered to test its effects on the cognitive behaviors of the APP-C105 AD mouse model using the Morris water maze and passive avoidance tests. To investigate the mechanisms of action that underlie the beneficial disease-modifying effects of mirodenafil, human neuroblastoma SH-SY5Y cells and mouse hippocampal HT-22 cells were used to show mirodenafil-induced alterations associated with the cyclic guanosine monophosphate (cGMP)/cGMP-dependent protein kinase (PKG)/cAMP-responsive element-binding protein (CREB) pathway, apoptotic cell death, tau phosphorylation, amyloidogenesis, the autophagy-lysosome pathway, glucocorticoid receptor (GR) transcriptional activity, and the Wnt/β-catenin signaling.ResultsHere, mirodenafil is demonstrated to improve cognitive behavior in the APP-C105 mouse model. Mirodenafil not only reduced the Aβ and phosphorylated tau burdens in vivo, but also ameliorated AD pathology induced by Aβ through the modulation of the cGMP/PKG/CREB signaling pathway, glycogen synthase kinase 3β (GSK-3β) activity, GR transcriptional activity, and the Wnt/β-catenin signaling in neuronal cells. Interestingly, homodimerization and nuclear localization of GR were inhibited by mirodenafil, but not by other PDE5 inhibitors. In addition, only mirodenafil reduced the expression levels of the Wnt antagonist Dickkopf-1 (Dkk-1), thus activating the Wnt/β-catenin signaling.ConclusionsThese findings strongly suggest that the PDE5 inhibitor mirodenafil shows promise as a potential polypharmacological drug candidate for AD treatment, acting on multiple key signaling pathways involved in amyloid deposition, phosphorylated tau burden, the cGMP/PKG/CREB pathway, GSK-3β kinase activity, GR signaling, and the Wnt/β-catenin signaling. Mirodenafil administration to the APP-C105 AD mouse model also improved cognitive behavior, demonstrating the potential of mirodenafil as a polypharmacological AD therapeutic agent.

Glucocorticoids differentially regulate the skeletal muscle transcriptome in response to acute aerobic exercise at distinct times of day

Adaptations to the skeletal muscle following long term aerobic exercise training are mediated at least in part by transcriptional changes that occur in response to each bout of exercise. There is evidence that glucocorticoids induce transcriptional changes in the skeletal muscle in response to a bout of aerobic exercise, but the extent of these changes and whether time of day exercise affects the glucocorticoid transcriptional response is unknown. The purpose of this study was to define changes to the skeletal muscle glucocorticoid transcriptome in response to acute aerobic exercise at different times of day. A secondary purpose was to define the role of Regulated in Development and DNA Damage 1 (REDD1) in mediating these changes as REDD1 is not only a transcriptional target of the glucocorticoid receptor, but it can also modulate glucocorticoid receptor transcriptional activity. A list of glucocorticoid target genes whose expression is altered in the skeletal muscle at various times of day in response to acute exercise was generated by comparing published genomic data sets from rodents subjected to acute aerobic exercise in the light or dark cycles to published genomic data sets from C2C12 myotubes treated with Dexamethasone. The role of glucocorticoid receptor signaling to the genomic changes was assessed in exercised mice treated with RU‐486. The role of REDD1 in mediating the changes to the glucocorticoid transcriptome was assessed in mice lacking REDD1. Glucocorticoids modulated expression of numerous genes in the skeletal muscle in response to acute aerobic exercise in the light and dark cycles, including those that are a target of, or contribute to, ubiquitin conjugation. The extent of changes to the glucocorticoid transcriptome was greater when exercise was conducted in the dark cycle compared to the light cycle. Only some genes encoding proteins involved in ubiquitin conjugation were induced by exercise at both times of day. REDD1 is required for the induction of these genes, whereas those only induced in the light cycle did not require REDD1. The time of day at which aerobic exercise is conducted dictates the extent of changes to the glucocorticoid transcriptome in skeletal muscle. REDD1 is a transducer mediating the time‐of‐day changes to the glucocorticoid transcriptome in skeletal muscle.

Corticosteroid sensitization drives opioid addiction.

The global crisis of opioid overdose fatalities has led to an urgent search to discover the neurobiological mechanisms of opioid use disorder (OUD). A driving force for OUD is the dysphoric and emotionally painful state (hyperkatifeia) that is produced during acute and protracted opioid withdrawal. Here, we explored a mechanistic role for extrahypothalamic stress systems in driving opioid addiction. We found that glucocorticoid receptor (GR) antagonism with mifepristone reduced opioid addiction-like behaviors in rats and zebrafish of both sexes and decreased the firing of corticotropin-releasing factor neurons in the rat amygdala (i.e., a marker of brain stress system activation). In support of the hypothesized role of glucocorticoid transcriptional regulation of extrahypothalamic GRs in addiction-like behavior, an intra-amygdala infusion of an antisense oligonucleotide that blocked GR transcriptional activity reduced addiction-like behaviors. Finally, we identified transcriptional adaptations of GR signaling in the amygdala of humans with OUD. Thus, GRs, their coregulators, and downstream systems may represent viable therapeutic targets to treat the "stress side" of OUD.

Glucocorticoid Receptor Overexpression in the Dorsal Hippocampus Attenuates Spatial Learning and Synaptic Plasticity Deficits after Pediatric Traumatic Brain Injury.

Traumatic brain injury (TBI) in children <4 years of age leads to long-term deficits in cognitive and learning abilities that can persist or even worsen as these children age into adolescence. In this study, the role of glucocorticoid receptor (GR) function in the dorsal hippocampus (DH) in hippocampal-dependent cognitive function and synaptic plasticity were assessed following injury to the 11-day-old rat. Brain injury produced significant impairments in spatial learning and memory in the Morris water maze in male and female rats at 1-month post-injury (adolescence), which was accompanied by impairments in induction and maintenance of long-term potentiation (LTP) in the CA1 region of the DH. Brain injury resulted in a significant decrease in the expression of the glucocorticoid-inducible gene, serum- and glucocorticoid-kinase 1 (sgk1), suggestive of an impairment in GR transcriptional activity within the hippocampus. Lentiviral transfection of the human GR (hGR) in the DH improved spatial learning and memory in the Morris water maze and attenuated LTP deficits following TBI. GR overexpression in the DH was also associated with a significant increase in the mRNA expression levels of sgk1, and the glutamate receptor subunits GluA1 and GluA2 within the hippocampus. Overall, these findings support an important role for dorsal hippocampal GR function in learning and memory deficits following pediatric TBI and suggest that these effects may be related to the regulation of glutamate receptor subunit expression in the DH.

Class 3 phosphoinositide 3-kinase promotes hepatic glucocorticoid receptor stability and transcriptional activity.

AimLipid kinase class 3 phosphoinositide 3‐kinase (PI3K) and nuclear receptor transcription factor glucocorticoid receptor (GR) play essential physiological roles in metabolic adaptation to fasting by activating lysosomal degradation by autophagy and metabolic gene expression, yet their functional interaction is unknown. The requirement of class 3 PI3K for GR function was investigated in liver tissue.MethodsInactivation of class 3 PI3K was achieved through deletion of its essential regulatory subunit Vps15, by expressing Cre‐recombinase in the livers of Vps15f/f mice. The response to both 24‐h fasting and synthetic GR ligand, dexamethasone (DEX) was evaluated in control and mutant mice. Liver tissue was analysed by immunoblot, RT‐qPCR, and LC‐MS.ResultsVps15 mutant mice show decreased transcript levels of GR targets, coupled with lower nuclear levels of total and phosphorylated on Ser211, GR protein. Acute DEX treatment and 24‐h fasting both failed to re‐activate expression of GR targets in the livers of Vps15 mutant mice to the levels observed in controls. Decreased levels of endogenous GR ligand corticosterone and lower expression of 11β‐hydroxysteroid dehydrogenase 1 (11β‐HSD1), a metabolic enzyme that controls corticosterone availability, were found in the livers of Vps15 mutants. Hepatic Vps15 depletion resulted in the activation of nuclear Akt1 signalling, which was paralleled by increased polyubiquitination of GR.ConclusionIn the liver, class 3 PI3K is required for corticosterone metabolism and GR transcriptional activity.