Deletion Of Glucocorticoid Receptor Research Articles

A 5:2 intermittent fasting regimen ameliorates NASH and fibrosis and blunts HCC development via hepatic PPARα and PCK1

The role and molecular mechanisms of intermittent fasting (IF) in non-alcoholic steatohepatitis (NASH) and its transition to hepatocellular carcinoma (HCC) are unknown. Here, we identified that an IF 5:2 regimen prevents NASH development as well as ameliorates established NASH and fibrosis without affecting total calorie intake. Furthermore, the IF 5:2 regimen blunted NASH-HCC transition when applied therapeutically. The timing, length, and number of fasting cycles as well as the type of NASH diet were critical parameters determining the benefits of fasting. Combined proteome, transcriptome, and metabolome analyses identified that peroxisome-proliferator-activated receptor alpha (PPARα) and glucocorticoid-signaling-induced PCK1 act co-operatively as hepatic executors of the fasting response. In line with this, PPARα targets and PCK1 were reduced in human NASH. Notably, only fasting initiated during the active phase of mice robustly induced glucocorticoid signaling and free-fatty-acid-induced PPARα signaling. However, hepatocyte-specific glucocorticoid receptor deletion only partially abrogated the hepatic fasting response. In contrast, the combined knockdown of Ppara and Pck1 invivo abolished the beneficial outcomes of fasting against inflammation and fibrosis. Moreover, overexpression of Pck1 alone or together with Ppara invivo lowered hepatic triglycerides and steatosis. Our data support the notion that the IF 5:2 regimen is a promising intervention against NASH and subsequent liver cancer.

Abstract 5517: Chronic stress causes metastasis via neutrophil-mediated changes to the microenvironment

Abstract Chronic stress is associated with increased risk of metastasis and poor survival in cancer patients, yet the reasons are unclear. We show that chronic stress increased lung metastasis from disseminated cancer cells 2- to 4-fold in mice. Chronic stress significantly altered the lung microenvironment, with fibronectin accumulation, reduced T cell infiltration, and increased neutrophil infiltration. Depleting neutrophils abolished stress-induced metastasis, indicating their critical role. Chronic stress shifted neutrophils’ normal circadian rhythm, and, via glucocorticoid release, caused increased neutrophil extracellular trap (NET) formation. Importantly, in mice with neutrophil-specific glucocorticoid receptor deletion, chronic stress failed to increase NETs and metastasis. Furthermore, digesting NETs with deoxyribonuclease (DNase) I prevented chronic stress-induced metastasis. Together, our data show that glucocorticoids released during chronic stress cause NET formation and establish a metastasis-promoting microenvironment. Therefore, NETs could be targets for preventing metastatic recurrence in cancer patients, many of whom will experience chronic stress due to their disease. Citation Format: Xue-Yan He, Yuan Gao, David Ng, Evdokia Michalopoulou, Shanu George, Jose M. Adrover, Lijuan Sun, Jean Albrengues, Juliane Daßler-Plenker, Xiao Han, Ledong Wan, Xiaoli S. Wu, Longling S. Shui, Yu-Han Huang, Bodu Liu, Chang Su, David L. Spector, Christopher R. Vakoc, Linda VanAelst, Mikala Egeblad. Chronic stress causes metastasis via neutrophil-mediated changes to the microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5517.

Chronic stress increases metastasis via neutrophil-mediated changes to the microenvironment

Chronic stress is associated with increased risk of metastasis and poor survival in cancer patients, yet the reasons are unclear. We show that chronic stress increases lung metastasis from disseminated cancer cells 2- to 4-fold in mice. Chronic stress significantly alters the lung microenvironment, with fibronectin accumulation, reduced Tcell infiltration, and increased neutrophil infiltration. Depleting neutrophils abolishes stress-induced metastasis. Chronic stress shifts normal circadian rhythm of neutrophils and causes increased neutrophil extracellular trap (NET) formation via glucocorticoid release. In mice with neutrophil-specific glucocorticoid receptor deletion, chronic stress fails to increase NETs and metastasis. Furthermore, digesting NETs with DNase I prevents chronic stress-induced metastasis. Together, our data show that glucocorticoids released during chronic stress cause NET formation and establish a metastasis-promoting microenvironment. Therefore, NETs could be targets for preventing metastatic recurrence in cancer patients, many of whom will experience chronic stress due to their disease.

Differential susceptibility of male and female germ cells to glucocorticoid-mediated signaling

While physiologic stress has long been known to impair mammalian reproductive capacity through hormonal dysregulation, mounting evidence now suggests that stress experienced prior to or during gestation may also negatively impact the health of future offspring. Rodent models of gestational physiologic stress can induce neurologic and behavioral changes that persist for up to three generations, suggesting that stress signals can induce lasting epigenetic changes in the germline. Treatment with glucocorticoid stress hormones is sufficient to recapitulate the transgenerational changes seen in physiologic stress models. These hormones are known to bind and activate the glucocorticoid receptor (GR), a ligand-inducible transcription factor, thus implicating GR-mediated signaling as a potential contributor to the transgenerational inheritance of stress-induced phenotypes. Here, we demonstrate dynamic spatiotemporal regulation of GR expression in the mouse germline, showing expression in the fetal oocyte as well as the perinatal and adult spermatogonia. Functionally, we find that fetal oocytes are intrinsically buffered against changes in GR signaling, as neither genetic deletion of GR nor GR agonism with dexamethasone altered the transcriptional landscape or the progression of fetal oocytes through meiosis. In contrast, our studies revealed that the male germline is susceptible to glucocorticoid-mediated signaling, specifically by regulating RNA splicing within the spermatogonia, although this does not abrogate fertility. Together, our work suggests a sexually dimorphic function for GR in the germline, and represents an important step towards understanding the mechanisms by which stress can modulate the transmission of genetic information through the germline.

Differential susceptibility of male and female germ cells to glucocorticoid-mediated signaling.

While physiologic stress has long been known to impair mammalian reproductive capacity through hormonal dysregulation, mounting evidence now suggests that stress experienced prior to or during gestation may also negatively impact the health of future offspring. Rodent models of gestational physiologic stress can induce neurologic and behavioral changes that persist for up to three generations, suggesting that stress signals can induce lasting epigenetic changes in the germline. Treatment with glucocorticoid stress hormones is sufficient to recapitulate the transgenerational changes seen in physiologic stress models. These hormones are known to bind and activate the glucocorticoid receptor (GR), a ligand-inducible transcription factor, thus implicating GR-mediated signaling as a potential contributor to the transgenerational inheritance of stress-induced phenotypes. Here, we demonstrate dynamic spatiotemporal regulation of GR expression in the mouse germline, showing expression in the fetal oocyte as well as the perinatal and adult spermatogonia. Functionally, we find that fetal oocytes are intrinsically buffered against changes in GR signaling, as neither genetic deletion of GR nor GR agonism with dexamethasone altered the transcriptional landscape or the progression of fetal oocytes through meiosis. In contrast, our studies revealed that the male germline is susceptible to glucocorticoid-mediated signaling, specifically by regulating RNA splicing within the spermatogonia, although this does not abrogate fertility. Together, our work suggests a sexually dimorphic function for GR in the germline, and represents an important step towards understanding the mechanisms by which stress can modulate the transmission of genetic information through the germline.

Lesion level-dependent systemic muscle wasting after spinal cord injury is mediated by glucocorticoid signaling in mice.

An incomplete mechanistic understanding of skeletal muscle wasting early after spinal cord injury (SCI) precludes targeted molecular interventions. Here, we demonstrated systemic wasting that also affected innervated nonparalyzed (supralesional) muscles and emerged within 1 week after experimental SCI in mice. Systemic muscle wasting caused muscle weakness, affected fast type 2 myofibers preferentially, and became exacerbated after high (T3) compared with low (T9) thoracic paraplegia, indicating lesion level-dependent ("neurogenic") mechanisms. The wasting of nonparalyzed muscle and its rapid onset and severity beyond what can be explained by disuse implied unknown systemic drivers. Muscle transcriptome and biochemical analysis revealed a glucocorticoid-mediated catabolic signature early after T3 SCI. SCI-induced systemic muscle wasting was mitigated by (i) endogenous glucocorticoid ablation (adrenalectomy) and (ii) pharmacological glucocorticoid receptor (GR) blockade and was (iii) completely prevented after T3 relative to T9 SCI by genetic muscle-specific GR deletion. These results suggest that neurogenic hypercortisolism contributes to a rapid systemic and functionally relevant muscle wasting syndrome early after paraplegic SCI in mice.

Stress aggravates imiquimod-induced psoriasiform inflammation by promoting M1 macrophage polarization

Psychological stress has long been considered to cause the aggravation and recurrence of psoriasis, but the underlying mechanism remains largely unknown. Here, we used a mouse model of restraint-induced stress and imiquimod (IMQ)-induced psoriasiform inflammation to investigate the crosstalk between stress and the skin immune system and their functions in the pathogenesis of psoriasis. We found that stress aggravated skin inflammation and elevated serum corticosterone (CORT) levels in mice. Stress also increased the number of macrophages and glucocorticoid receptor (GR) expression in IMQ-treated mouse skin. GR agonist CORT upregulated the phosphorylation of STAT1 to promote M1 macrophage polarization in vitro. Additionally, GR deletion in macrophages and pharmacologic inhibition of GR improved skin inflammation and reduced M1 macrophage polarization under stress. Taken together, these results indicate that stress aggravates psoriasiform inflammation by promoting CORT/GR signaling-induced M1 macrophage polarization, suggesting that blocking the GR signaling has great potential as an adjuvant treatment for psoriasis patients with chronic stress.

Deletion of the chondrocyte glucocorticoid receptor attenuates cartilage degradation through suppression of early synovial activation in murine posttraumatic osteoarthritis

Disruption of endogenous glucocorticoid signalling in bone cells attenuates osteoarthritis (OA) in aged mice, however, the role of endogenous glucocorticoids in chondrocytes is unknown. Here, we investigated whether deletion of the glucocorticoid receptor, specifically in chondrocytes, also alters OA progression. Knee OA was induced by surgical destabilisation of the medial meniscus (DMM) in male 22-week-old tamoxifen-inducible glucocorticoid receptor knockout (chGRKO) mice and their wild-type (WT) littermates (n=7-9/group). Mice were harvested 2, 4, 8 and 16 weeks after surgery to examine the spatiotemporal changes in molecular, cellular, and histological characteristics. At all time points following DMM, cartilage damage was significantly attenuated in chGRKO compared to WT mice. Two weeks after DMM, WT mice exhibited increased chondrocyte and synoviocyte hypoxia inducible factor (HIF)-2α expression resulting in extensive synovial activation characterised by synovial thickening and increased interleukin-1 beta expression. At 2 and 4weeks after DMM, WT mice displayed pronounced chondrocyte senescence and elevated catabolic signalling (reduced Yes-associated protein 1 (YAP1) and increased matrixmetalloprotease [MMP]-13 expression). Contrastingly, at 2 weeks after DMM, HIF-2α expression and synovial activation were much less pronounced in chGRKO than in WT mice. Furthermore, chondrocyte YAP1 and MMP-13 expression, as well as chondrocyte senescence were similar in chGRKO-DMM mice and sham-operated controls. Endogenous glucocorticoid signalling in chondrocytes promotes synovial activation, chondrocyte senescence and cartilage degradation by upregulation of catabolic signalling through HIF-2α in murine posttraumatic OA. These findings indicate that inhibition of glucocorticoid signalling early after injury may present a promising way to slow osteoarthritic cartilage degeneration.

The crucial role of muscle glucocorticoid signaling in accelerating obesity and glucose intolerance via hyperinsulinemia.

Metabolic crosstalk from skeletal muscle to multiple organs is important for maintaining homeostasis, and its dysregulation can lead to various diseases. Chronic glucocorticoid administration often induces muscle atrophy and metabolic disorders such as diabetes and central obesity; however, the detailed underlying mechanism remains unclear. We previously reported that the deletion of glucocorticoid receptor (GR) in skeletal muscle increases muscle mass and reduces fat mass through muscle-liver-fat communication under physiological conditions. In this study, we show that muscle GR signaling plays a crucial role in accelerating obesity through the induction of hyperinsulinemia. Fat accumulation in liver and adipose tissue, muscle atrophy, hyperglycemia, and hyperinsulinemia induced by chronic corticosterone (CORT) treatment improved in muscle-specific GR knockout (GRmKO) mice. Such CORT-induced fat accumulation was alleviated by suppressing insulin production (streptozotocin injection), indicating that hyperinsulinemia enhanced by muscle GR signaling promotes obesity. Strikingly, glucose intolerance and obesity in ob/ob mice without CORT treatment were also improved in GRmKO mice, indicating that muscle GR signaling contributes to obesity-related metabolic changes, regardless of systemic glucocorticoid levels. Thus, this study provides new insight for the treatment of obesity and diabetes by targeting muscle GR signaling.

Hypothalamic AgRP neurons exert top-down control on systemic TNF-α release during endotoxemia

Loss of appetite and negative energy balance are common features of endotoxemia in all animals and are thought to have protective roles by reducing nutrient availability to host and pathogen metabolism. Accordingly, fasting and caloric restriction have well-established anti-inflammatory properties. However, in response to reduced nutrient availability at the cellular and organ levels, negative energy balance also recruits distinct energy-sensing brain circuits, but it is not known whether these neuronal systems have a role in its anti-inflammatory effects. Here, we report that hypothalamic AgRP neurons-a critical neuronal population for the central representation of negative energy balance-have parallel immunoregulatory functions. We found that when endotoxemia occurs in fasted mice, the activity of AgRP neurons remains sustained, but this activity does not influence feeding behavior and endotoxemic anorexia. Furthermore, we found that endotoxemia acutely desensitizes AgRP neurons, which also become refractory to inhibitory signals. Mimicking this sustained AgRP neuron activity in fed mice by chemogenetic activation-a manipulation known to recapitulate core behavioral features of fasting-results in reduced acute tumor necrosis factor alpha (TNF-α) release during endotoxemia. Mechanistically, we found that endogenous glucocorticoids play an important role: glucocorticoid receptor deletion from AgRP neurons prevents their endotoxemia-induced desensitization, and importantly, it counteracts the fasting-induced suppression of TNF-α release, resulting in prolonged sickness. Together, these findings provide evidence directly linking AgRP neuron activity to the acute response during endotoxemia, suggesting that these neurons are a functional component of the immunoregulatory effects associated with negative energy balance and catabolic metabolism.

Genetic deletion of the glucocorticoid receptor in Cx3cr1+ myeloid cells is neuroprotective and improves motor recovery after spinal cord injury

Glucocorticoid receptors (GRs), part of the nuclear receptor superfamily of transcription factors (TFs), are ubiquitously expressed in all cell types and regulate cellular responses to glucocorticoids (e.g., cortisol in humans; corticosterone in rodents). In myeloid cells, glucocorticoids binding to GRs can enhance or repress gene transcription, thereby imparting distinct and context-dependent functions in macrophages at sites of inflammation. In experimental models and in humans, glucocorticoids are widely used as anti-inflammatory treatments to promote recovery of function after SCI. Thus, we predicted that deleting GR in mouse myeloid lineage cells (i.e., microglia and monocyte-derived macrophages) would enhance inflammation at the site of injury and worsen functional recovery after traumatic spinal cord injury (SCI). Contrary to our prediction, the intraspinal macrophage response to a moderate (75 kdyne) spinal contusion SCI was reduced in Cx3cr1-Cre;GRf/f conditional knockout mice (with GR specifically deleted in myeloid cells). This phenotype was associated with improvements in hindlimb motor recovery, myelin sparing, axon sparing/regeneration, and microvascular protection/plasticity relative to SCI mice with normal myeloid cell GR expression. Further analysis revealed that macrophage GR deletion impaired lipid and myelin phagocytosis and foamy macrophage formation. Together, these data reveal endogenous GR signaling as a key pathway that normally inhibits mechanisms of macrophage-mediated repair after SCI.

Conditional deletion of the glucocorticoid receptor in vascular smooth muscle disrupts the circadian rhythm of vascular reactivity and reduces blood pressure

Glucocorticoids synchronise peripheral clocks with the master clock in the suprachiasmatic nucleus in response to the ambient light cycle. In humans and mice, arrhythmic glucocorticoids induce non‐dipping blood pressure and vascular dysfunction. The mechanisms of this are poorly understood. We hypothesize that arrhythmic activation of the glucocorticoid receptor attenuates the circadian clock signalling in the vasculature leading to changes in blood pressure. To test this hypothesis, we assessed vascular function and blood pressure rhythms in control mice and mice with smooth muscle specific deletion of the glucocorticoid receptor (SMGRKO).Renal and mesenteric arteries were isolated at 7am (ZT0; lights on) and 7pm (ZT12; lights off), mounted on a wire myograph and the responses to increasing doses of phenylephrine or sodium nitroprusside (SNP) assessed. In wild‐type mice, the sensitivity of the renal artery to phenylephrine‐mediated contraction was greater at ZT12 than at ZT0, by comparing the half maximal effective concentration of each group, (logEC50: 7.54±0.26 versus 7.29±0.17 respectively; p=0.04). The sensitivity to SNP‐mediated relaxation was also greater when measured at ZT12 compared to ZT0 (7.64±0.3 versus 7.09±0.5; p=0.03). Similar responses were also detected in the mesenteric arteries. In SMGRKO mice, the temporal differences in renal and mesenteric artery vascular reactivity were absent. We used radiotelemetry to assess the effect of this altered rhythm of vascular reactivity on blood pressure. SMGRKO had significantly lower systolic blood pressure throughout the 24h cycle compared to wild‐type littermates, both when lights were off (107.9 ± 22.13 versus 136.3 ± 17.17 mmHg respectively; p=0.015) and during the lights on period (99.6 ± 13.16 versus 118 ± 15.9 mmHg; p=0.014). Blood pressure rhythmicity was not altered in SMGRKO mice.These data show that the glucocorticoid receptor signalling in vascular smooth muscle contributes both to the diurnal variation of vascular reactivity and to blood pressure, but loss of the receptor does not affect blood pressure rhythm. Whether glucocorticoid receptor deletion in smooth muscle protects against non‐dipping blood pressure induced by glucocorticoids needs further investigation.

The glucocorticoid receptor associates with RAS complexes to inhibit cell proliferation and tumor growth.

Mutations that activate members of the RAS family of GTPases are associated with various cancers and drive tumor growth. The glucocorticoid receptor (GR), a member of the nuclear receptor family, has been proposed to interact with and inhibit the activation of components of the PI3K-AKT and MAPK pathways downstream of RAS. In the absence of activating ligands, we found that GR was present in cytoplasmic KRAS-containing complexes and inhibited the activation of wild-type and oncogenic KRAS in mouse embryonic fibroblasts and human lung cancer A549 cells. The DNA binding domain of GR was involved in the interaction with KRAS, but GR-dependent inhibition of RAS activation did not depend on the nuclear translocation of GR. The addition of ligand released GR-dependent inhibition of RAS, AKT, the MAPK p38, and the MAPKK MEK. CRISPR-Cas9-mediated deletion of GR in A549 cells enhanced tumor growth in xenografts in mice. Patient samples of non-small cell lung carcinomas showed lower expression of NR3C1, the gene encoding GR, compared to adjacent normal tissues and lower NR3C1 expression correlated with a worse disease outcome. These results suggest that glucocorticoids prevent the ability of GR to limit tumor growth by inhibiting RAS activation, which has potential implications for the use of glucocorticoids in patients with cancer.

Glucocorticoid Receptor–Dependent Astrocytes Mediate Stress Vulnerability

BackgroundMajor depressive disorder is a devastating psychiatric illness that affects approximately 17% of the population worldwide. Astrocyte dysfunction has been implicated in its pathophysiology. Traumatic experiences and stress contribute to the onset of major depressive disorder, but how astrocytes respond to stress is poorly understood. MethodsUsing Western blotting analysis, we identified that stress vulnerability was associated with reduced astrocytic glucocorticoid receptor (GR) expression in mouse models of depression. We further investigated the functions of astrocytic GRs in regulating depression and the underlying mechanisms by using a combination of behavioral studies, fiber photometry, biochemical experiments, and RNA sequencing methods. ResultsGRs in astrocytes were more sensitive to stress than those in neurons. GR absence in astrocytes induced depressive-like behaviors, whereas restoring astrocytic GR expression in the medial prefrontal cortex prevented the depressive-like phenotype. Furthermore, we found that GRs in the medial prefrontal cortex affected astrocytic Ca2+ activity and dynamic ATP (adenosine 5′-triphosphate) release in response to stress. RNA sequencing of astrocytes isolated from GR deletion mice identified the PI3K-Akt (phosphoinositide 3-kinase–Akt) signaling pathway, which was required for astrocytic GR–mediated ATP release. ConclusionsThese findings reveal that astrocytic GRs play an important role in stress response and that reduced astrocytic GR expression in the stressed subject decreases ATP release to mediate stress vulnerability.

Deletion of hippocampal Glucocorticoid receptors unveils sex-biased microRNA expression and neuronal morphology alterations in mice

Sex differences in the brain have prompted many researchers to investigate the underlying molecular actors, such as the glucocorticoid receptor (GR). This nuclear receptor controls gene expression, including microRNAs (miRNAs), in non-neuronal cells. Here, we investigated sex-biased effects of GR on hippocampal miRNA expression and neuronal morphology by generating a neuron-specific GR knockout mouse (Emx1-Nr3c1−/−). The levels of 578 mature miRNAs were assessed using NanoString technology and, in contrast to males, female Emx1-Nr3c1−/− mice showed a substantially higher number of differentially expressed miRNAs, confirming a sex-biased effect of GR ablation. Based on bioinformatic analyses we identified several transcription factors potentially involved in miRNA regulation. Functional enrichment analyses of the miRNA-mRNA interactions revealed pathways related to neuronal arborization and both spine morphology and density in both sexes. Two recognized regulators of dendritic morphology, CAMKII-α and GSK-3β, increased their protein levels by GR ablation in female mice hippocampus, without changes in males. Additionally, sex-specific effects of GR deletion were observed on CA1 neuronal arborization and dendritic spine features. For instance, a reduced density of mushroom spines in apical dendrites was evidenced only in females, while a decreased length in basal dendrites was noted only in males. However, length and arborization of apical dendrites were reduced by GR ablation irrespective of the sex. Overall, our study provides new insights into the sex-biased GR actions, especially in terms of miRNAs expression and neuronal morphology in the hippocampus.

P.374 Glucocorticoid receptor deletion in microglia alters neuroinflammatory response to stress

P.374 Glucocorticoid receptor deletion in microglia alters neuroinflammatory response to stress

MON-LB130 Glucocorticoid Receptor Signaling and Stress in the Developing Mammalian Germline

While the consequences of stress on organs such as the heart and immune system have long been established, it has only recently been shown in mice that stress experienced during pregnancy can cause molecular and behavioral changes in the exposed fetus, and even their offspring. The transmission of such phenotypes across generations implies that fetal germ cells can sense and maintain epigenetic memories of stress experienced in the womb. However, how the fetal germ cells within the developing embryo detect and respond to these stress signals is unknown. In response to stress, the major circulating hormone released is cortisol, which is the ligand for the nuclear hormone receptor known as the glucocorticoid receptor (GR). GR induces highly cell type-specific transcriptional changes in many cell types, although its function in germ cells has yet to be explored. Our preliminary studies have shown, for the first time, the robust expression and dynamic localization of GR in fetal germ cells, suggesting its activity during a crucial window of epigenetic reprogramming in male and female germ cell development. Fetal germ cells naturally undergo genome-wide demethylation of somatic and parental imprints, which leads to the de-repression of transposons that are normally silenced by methylation at their promoters. If not properly re-silenced, these transposons will generate large numbers of DNA double strand breaks during transposition, posing a serious risk to germline integrity. When GR was genetically deleted from all cells of the mouse body, we discovered elevated expression of the retrotransposon, Line1 (L1), specifically in the developing germ cells. Surprisingly, mice with a conditional deletion of GR specifically in germ cells were unable to phenocopy the full body knockout, implying that GR-mediated repression of L1 in fetal germ cells is a non-cell autonomous effect. Rather, conditional deletion of GR from the surrounding somatic cells (e.g. granulosa cells of the ovary) demonstrated increased germ cell L1 expression, strongly suggesting that GR signaling through the somatic niche is indirectly regulating L1 expression in neighboring germ cells. We therefore hypothesize that GR signaling from the somatic cell niche is functioning to indirectly suppress L1 expression within the developing fetal germ cells, which holds broad implications for how stress signals mediated by GR signaling can influence germline integrity and thus the health of future generations.

Selective effects of dorsal raphé nucleus glucocorticoid receptor deletion on depression-like behavior in female C57BL/6J mice

We have shown differing effects of glucocorticoid receptor (GR) deletion from the dorsal raphé nucleus (DRN) and locus coeruleus (LC) on depression-relevant behavior in male mice, but DRN GR deletion has not been tested in female mice. Female floxed GR mice were given DRN injections of AAV2/9 pseudotype viral vectors transducing Cre recombinase to produce DRN GR gene deletion (Cre) and compared with mice receiving DRN injections of AAV2/9 transducing green fluorescent protein (GFP). Social interaction, a measure of depression-like withdrawal, was unaffected by DRN GR deletion, but forced swim immobility, a measure of despair-like passivity, was reduced in female Cre vs. GFP mice. Behavioral effects were not attributable to changes in basal corticosterone or LC GR deletion. Combined with our prior studies, the current findings suggest that DRN GR have sex-independent effects to promote forced swim immobility, but influence social interaction only in male mice. Differential effects of DRN GR deletion in female mice may provide insight into the greater incidence of depression and specific depression symptoms in women.

Deletion of the serine protease CAP2/Tmprss4 leads to dysregulated renal water handling upon dietary potassium depletion

The kidney needs to adapt daily to variable dietary K+ contents via various mechanisms including diuretic, acid-base and hormonal changes that are still not fully understood. In this study, we demonstrate that following a K+-deficient diet in wildtype mice, the serine protease CAP2/Tmprss4 is upregulated in connecting tubule and cortical collecting duct and also localizes to the medulla and transitional epithelium of the papilla and minor calyx. Male CAP2/Tmprss4 knockout mice display altered water handling and urine osmolality, enhanced vasopressin response leading to upregulated adenylate cyclase 6 expression and cAMP overproduction, and subsequently greater aquaporin 2 (AQP2) and Na+-K+-2Cl− cotransporter 2 (NKCC2) expression following K+-deficient diet. Urinary acidification coincides with significantly increased H+,K+-ATPase type 2 (HKA2) mRNA and protein expression, and decreased calcium and phosphate excretion. This is accompanied by increased glucocorticoid receptor (GR) protein levels and reduced 11β-hydroxysteroid dehydrogenase 2 activity in knockout mice. Strikingly, genetic nephron-specific deletion of GR leads to the mirrored phenotype of CAP2/Tmprss4 knockouts, including increased water intake and urine output, urinary alkalinisation, downregulation of HKA2, AQP2 and NKCC2. Collectively, our data unveil a novel role of the serine protease CAP2/Tmprss4 and GR on renal water handling upon dietary K+ depletion.

Glucocorticoid signaling regulates CD8+T cell responses in cancer

Abstract Identifying signals in the tumor microenvironment (TME) that promote CD8+T cell dysfunction can inform the development of novel approaches to improve anti-tumor T cell responses. Here, we identified Nr3c1, the gene encoding the glucocorticoid receptor (GR), as one of the genes preferentially expressed by dysfunctional CD8+TILs. Glucocorticoids (GCs) are steroid hormones and ligands for the GR. Despite their widespread use, surprisingly little is known regarding the molecular circuitry by which glucocorticoids suppress immune responses. We demonstrate that glucocorticoids are synthesized de novo in the TME and activation of glucocorticoid signaling promotes features of dysfunction, including increased expression of checkpoint receptors like Tim3, PD1, Lag3 and TIGIT, and dampened effector function including decreased production of pro-inflammatory cytokines in both murine and human CD8+T cells. Deletion of GR in CD8+T cells in vivo resulted in loss of dysfunctional phenotype in CD8+TILs concomitant with improved tumor growth control. We thus identified glucocorticoid signaling as a component of immune suppression within the TME and uncovered a molecular mechanism by which it suppresses T cell responses. Glucocorticoids are the mainstay for treating the immune-related adverse events (IRAEs) that develop in patients receiving immune checkpoint blockade. Thus, having a better understanding of the mechanisms downstream of GCs could inform the application of glucocorticoids for suppressing immune responses.