Serum Glucocorticoid Kinase Research Articles

Localization and function of humanized F508del-CFTR in mouse intestine following activation of serum glucocorticoid kinase 1 and Trikafta

The CFTR modulator Trikafta has markedly improved lung disease for Cystic Fibrosis (CF) patients carrying the common delta F508 (F508del-CFTR) CFTR mutation. F508del-CFTR results in an apical trafficking defect and loss of function in CFTR-expressing epithelial cells. However, Trikafta has not resulted in improved gastrointestinal function in CF patients. A humanized mouse model of F508del-CFTR was recently generated to evaluate CFTR modulators and other compounds to treat human F508del-CFTR CF intestinal disease. Short-term (4 h) treatment of rats with Dexamethasone (Dex) potently activates serum glucocorticoid kinase 1 (SGK1) and increases CFTR apical traffic and ion transport in the native intestine. This study examined CFTR localization and ion transport in intestinal segments from humanized F508del-CFTR mice following treatment with Dex in the presence/absence of Trikafta. Dex treatment improved apical CFTR localization and function but was inconsistent along intestinal segments. Combined treatment with Dex and Trikafta was superior to Dex alone but inconsistently improved CFTR localization and function. These data suggest further optimization of humanized CF mouse models will be necessary to test the efficacy of compounds to treat human CF intestinal disease.

SGK1 is involved in doxorubicin-induced chronic cardiotoxicity and dysfunction through activation of the NFκB pathway

Breast cancer is the predominant cancer among women worldwide, and chemotherapeutic agents, such as doxorubicin (DOX), have the potential to significantly prolong survival, albeit at the cost of inducing severe cardiovascular toxicity. Inflammation has emerged as a crucial biological process contributing to the remodeling of cardiovascular toxicity. The role of serum glucocorticoid kinase 1 (SGK1) in various inflammatory diseases has been extensively investigated. Here, we studied the molecular mechanisms underlying the function of SGK1 in DOX-induced cardiotoxicity in HL-1 cardiomyocyte cell lines and in a tumor-bearing mouse model. SGK1 was upregulated in the DOX-induced cardiotoxicity model, accompanied by increased levels of inflammatory factors. Furthermore, inhibition of SGK1 suppresses the phosphorylation of nuclear factor-kappa B (NFκB) in cardiomyocytes, which inhibits the production of inflammatory factors and apoptosis of cardiomyocytes, and has cardioprotective effects. Simultaneously, small interfering RNA targeting SGK1 inhibited the proliferation of breast cancer cells. Conversely, overexpression of SGK1 increases the phosphorylation of NFκB and aggravates myocardial injury. In conclusion, our study demonstrates that SGK1 promotes DOX-induced cardiac inflammation and apoptosis by promoting NFκB activity. Our results indicate that inhibiting SGK1 might be an effective treatment strategy that can provide both tumor-killing and cardioprotective functions. Further in vivo research is needed to fully elucidate the effects and mechanisms of combination therapy with SGK1 inhibitors and DOX in breast cancer treatment.

Eicosanoid-Regulated Myeloid ENaC and Isolevuglandin Formation in Human Salt-Sensitive Hypertension.

The mechanisms by which salt increases blood pressure in people with salt sensitivity remain unclear. Our previous studies found that high sodium enters antigen-presenting cells (APCs) via the epithelial sodium channel and leads to the production of isolevuglandins and hypertension. In the current mechanistic clinical study, we hypothesized that epithelial sodium channel-dependent isolevuglandin-adduct formation in APCs is regulated by epoxyeicosatrienoic acids (EETs) and leads to salt-sensitive hypertension in humans. Salt sensitivity was assessed in 19 hypertensive subjects using an inpatient salt loading and depletion protocol. Isolevuglandin-adduct accumulation in APCs was analyzed using flow cytometry. Gene expression in APCs was analyzed using cellular indexing of transcriptomes and epitopes by sequencing analysis of blood mononuclear cells. Plasma and urine EETs were measured using liquid chromatography-mass spectrometry. Baseline isolevuglandin+ APCs correlated with higher salt-sensitivity index. Isolevuglandin+ APCs significantly decreased from salt loading to depletion with an increasing salt-sensitivity index. We observed that human APCs express the epithelial sodium channel δ subunit, SGK1 (salt-sensing kinase serum/glucocorticoid kinase 1), and cytochrome P450 2S1. We found a direct correlation between baseline urinary 14,15 EET and salt-sensitivity index, whereas changes in urinary 14,15 EET negatively correlated with isolevuglandin+ monocytes from salt loading to depletion. Coincubation with 14,15 EET inhibited high-salt-induced increase in isolevuglandin+ APC. Isolevuglandin formation in APCs responds to acute changes in salt intake in salt-sensitive but not salt-resistant people with hypertension, and this may be regulated by renal 14,15 EET. Baseline levels of isolevuglandin+ APCs or urinary 14,15 EET may provide diagnostic tools for salt sensitivity without a protocol of salt loading.

Role of epithelial sodium channel-related inflammation in human diseases.

The epithelial sodium channel (ENaC) is a heterotrimer and is widely distributed throughout the kidneys, blood vessels, lungs, colons, and many other organs. The basic role of the ENaC is to mediate the entry of Na+ into cells; the ENaC also has an important regulatory function in blood pressure, airway surface liquid (ASL), and endothelial cell function. Aldosterone, serum/glucocorticoid kinase 1 (SGK1), shear stress, and posttranslational modifications can regulate the activity of the ENaC; some ion channels also interact with the ENaC. In recent years, it has been found that the ENaC can lead to immune cell activation, endothelial cell dysfunction, aggravated inflammation involved in high salt-induced hypertension, cystic fibrosis, pseudohypoaldosteronism (PHA), and tumors; some inflammatory cytokines have been reported to have a regulatory role on the ENaC. The ENaC hyperfunction mediates the increase of intracellular Na+, and the elevated exchange of Na+ with Ca2+ leads to an intracellular calcium overload, which is an important mechanism for ENaC-related inflammation. Some of the research on the ENaC is controversial or unclear; we therefore reviewed the progress of studies on the role of ENaC-related inflammation in human diseases and their mechanisms.

Genetic inhibition of serum glucocorticoid kinase 1 prevents obesity-related atrial fibrillation.

Obesity is an important risk factor for atrial fibrillation (AF), but a better mechanistic understanding of obesity-related atrial fibrillation is required. Serum glucocorticoid kinase 1 (SGK1) is a kinase positioned within multiple obesity-related pathways, and prior work has shown a pathologic role of SGK1 signaling in ventricular arrhythmias. We validated a mouse model of obesity-related AF using wild-type mice fed a high-fat diet. RNA sequencing of atrial tissue demonstrated substantial differences in gene expression, with enrichment of multiple SGK1-related pathways, and we showed upregulated of SGK1 transcription, activation, and signaling in obese atria. Mice expressing a cardiac specific dominant-negative SGK1 were protected from obesity-related AF, through effects on atrial electrophysiology, action potential characteristics, structural remodeling, inflammation, and sodium current. Overall, this study demonstrates the promise of targeting SGK1 in a mouse model of obesity-related AF.

SGK1 is essential for meiotic resumption in mammalian oocytes

In mammalian females, oocytes are stored in the ovary and meiosis is arrested at the diplotene stage of prophase I. When females reach puberty oocytes are selectively recruited in cycles to grow, overcome the meiotic arrest, complete the first meiotic division and become mature (ready for fertilization). At a molecular level, the master regulator of prophase I arrest and meiotic resumption is the maturation-promoting factor (MPF) complex, formed by the active form of cyclin dependent kinase 1 (CDK1) and Cyclin B1. However, we still do not have complete information regarding the factors implicated in MPF activation.In this study we document that out of three mammalian serum-glucocorticoid kinase proteins (SGK1, SGK2, SGK3), mouse oocytes express only SGK1 with a phosphorylated (active) form dominantly localized in the nucleoplasm. Further, suppression of SGK1 activity in oocytes results in decreased CDK1 activation via the phosphatase cell division cycle 25B (CDC25B), consequently delaying or inhibiting nuclear envelope breakdown. Expression of exogenous constitutively active CDK1 can rescue the phenotype induced by SGK1 inhibition. These findings bring new insights into the molecular pathways acting upstream of MPF and a better understanding of meiotic resumption control by presenting a new key player SGK1 in mammalian oocytes.

Immune Mechanisms of Dietary Salt-Induced Hypertension and Kidney Disease: Harry Goldblatt Award for Early Career Investigators 2020.

Salt sensitivity of blood pressure is an independent risk factor for cardiovascular mortality not only in hypertensive but also in normotensive adults. The diagnosis of salt sensitivity of blood pressure is not feasible in the clinic due to lack of a simple diagnostic test, making it difficult to investigate therapeutic strategies. Most research efforts to understand the mechanisms of salt sensitivity of blood pressure have focused on renal regulation of sodium. However, salt retention or plasma volume expansion is not different between salt-sensitive and salt-resistant individuals. In addition, over 70% of extracellular fluid is interstitial and, therefore, not directly controlled by renal salt and water excretion. We discuss in this review how the seminal work by Harry Goldblatt paved the way for our attempts at understanding the mechanisms that underlie immune activation by salt in hypertension. We describe our findings that sodium, entering antigen-presenting cells via an epithelial sodium channel, triggers a PKC (protein kinase C)- and SGK1 (serum/glucocorticoid kinase 1)-stimulated activation of nicotinamide adenine dinucleotide phosphate oxidase, which, in turn, enhances lipid oxidation with generation of highly reactive isolevuglandins. Isolevuglandins adduct to proteins, with the potential to generate degraded peptide neoantigens. Activated antigen-presenting cells increase production of the TH17 polarizing cytokines, IL (interleukin)-6, IL-1β, and IL-23, which leads to differentiation and proliferation of IL-17A producing T cells. Our laboratory and others have shown that this cytokine contributes to hypertension. We also discuss where this sodium activation of antigen-presenting cells may occur in vivo and describe the multiple experiments, with pharmacological antagonists and knockout mice that we used to unravel this sequence of events in rodents. Finally, we describe experiments in mononuclear cells obtained from normotensive or hypertensive volunteers, which confirm that analogous processes of salt-induced immunity take place in humans.

B-AB18-04 SGK1 DEFICIENCY IS PROTECTIVE IN A MODEL OF SEPSIS-RELATED ATRIAL FIBRILLATION

Sepsis is associated with new onset atrial fibrillation (AF), and AF in septic patients is associated with worse outcomes. Serum glucocorticoid kinase 1 (SGK1), a serine/threonine kinase in the PI-3 kinase pathway, is involved in cardiac inflammation. Prior work has suggested a potential therapeutic role for SGK1 inhibition in cardiac arrhythmias, which we investigated in a model of bacterial sepsis-induced AF. We sought to determine if 1) CLP, a well established model of polymicrobial sepsis leads to an increased risk of AF, and 2) whether SGK1 inhibition is protective in this model. Cecal ligation and puncture was performed in male wild type C57/Bl6 (WT) and mice with cardiac specific knockdown of SGK1 (SGK1 DN). Approximately 7 (range 7-9) days after sepsis induction, the mice underwent terminal in vivo electrophysiology studies to determine AF inducibility and basic atrial/ventricular tissue parameters. Additional mice were used to perform optical mapping to determine EP properties and flow cytometry to determine cellular composition. CLP-induced sepsis in WT mice led to high AF inducibility (5 out of 7 mice), as well as AF frequency and burden during EP study. As compared with naïve controls, the AF phenotype was associated with significantly slowed right atrial conduction velocity (0.54±0.03 versus 0.62±0.08 m/s) along with a similar trend in the left atrium, as measured by optical mapping. Flow cytometry revealed that sepsis increased fibroblasts, macrophages, and neutrophils in the atrial wall when compared to control mice without CLP. When CLP was performed in WT and SGK1 DN littermates, SGK1 DN mice had significantly (p<0.05 by chi-squared) lower AF inducibility (3 out of 8) than the WT mice (7 out of 8). In addition, there was a significant decrease in AF frequency and a trend towards a decrease in total AF burden during the entirety of the EP study. Bacterial sepsis in mice remodels the atrial cell composition and increases AF inducibility. Genetic SGK1 inhibition protected septic mice against AF. These data motivate further investigation into how inflammatory remodeling of atria contributes to sepsis-induced AF, and the potential of SGK1 inhibitors for treatment of this entity.

Convergent genomic and pharmacological evidence of PI3K/GSK3 signaling alterations in neurons from schizophrenia patients

Human-induced pluripotent stem cells (hiPSCs) allow for the establishment of brain cellular models of psychiatric disorders that account for a patient’s genetic background. Here, we conducted an RNA-sequencing profiling study of hiPSC-derived cell lines from schizophrenia (SCZ) subjects, most of which are from a multiplex family, from the population isolate of the Central Valley of Costa Rica. hiPSCs, neural precursor cells, and cortical neurons derived from six healthy controls and seven SCZ subjects were generated using standard methodology. Transcriptome from these cells was obtained using Illumina HiSeq 2500, and differential expression analyses were performed using DESeq2 (|fold change|>1.5 and false discovery rate < 0.3), in patients compared to controls. We identified 454 differentially expressed genes in hiPSC-derived neurons, enriched in pathways including phosphoinositide 3-kinase/glycogen synthase kinase 3 (PI3K/GSK3) signaling, with serum-glucocorticoid kinase 1 (SGK1), an inhibitor of glycogen synthase kinase 3β, as part of this pathway. We further found that pharmacological inhibition of downstream effectors of the PI3K/GSK3 pathway, SGK1 and GSK3, induced alterations in levels of neurite markers βIII tubulin and fibroblast growth factor 12, with differential effects in patients compared to controls. While demonstrating the utility of hiPSCs derived from multiplex families to identify significant cell-specific gene network alterations in SCZ, these studies support a role for disruption of PI3K/GSK3 signaling as a risk factor for SCZ.

Acute Hyperglycemia Aggravates Lung Injury via Activation of the SGK1-NKCC1 Pathway.

Acute lung injury (ALI) is characterized by severe hypoxemia and has significantly high mortality rates. Acute hyperglycemia occurs in patients with conditions such as sepsis or trauma, among others, and it results in aggravated inflammation and induces damage in patients with ALI. Regulation of alveolar fluid is essential for the development and resolution of pulmonary edema in lung injury. Pulmonary sodium-potassium-chloride co-transporter 1 (NKCC1) regulates the net influx of ions and water into alveolar cells. The activation of with-no-lysine kinase 4 (WNK4), STE20/SPS1-related proline/alanine rich kinase (SPAK) and the NKCC1 pathway lead to an increase in the expression of NKCC1 and aggravation of ALI. Moreover, hyperglycemia is known to induce NKCC1 expression via the activation of the serum-glucocorticoid kinase 1 (SGK1)–NKCC1 pathway. We aim to evaluate the influence of acute hyperglycemia on the SGK1–NKCC1 pathway in ALI. ALI was induced using a high tidal volume for four hours in a rat model. Acute hyperglycemia was induced by injection with 0.5 mL of 40% glucose solution followed by continuous infusion at 2 mL/h. The animals were divided into sham, sham+ hyperglycemia, ALI, ALI + hyperglycemia, ALI + inhaled bumetanide (NKCC1 inhibitor) pretreatment, ALI + hyperglycemia + inhalational bumetanide pretreatment, and ALI + hyperglycemia + post-ALI inhalational bumetanide groups. Severe lung injury along with pulmonary edema, alveolar protein leakage, and lung inflammation was observed in ALI with hyperglycemia than in ALI without hyperglycemia. This was concurrent with the higher expression of pro-inflammatory cytokines, infiltration of neutrophils and alveolar macrophages (AM) 1, and NKCC1 expression. Inhalational NKCC1 inhibitor significantly inhibited the SGK1–NKCC1, and WNK4–SPAK–NKCC1 pathways. Additionally, it reduced pulmonary edema, inflammation, levels of pro-inflammatory cytokines, neutrophils and AM1 and increased AM2. Therefore, acute hyperglycemia aggravates lung injury via the further activation of the SGK1–NKCC1 pathway. The NKCC1 inhibitor can effectively attenuate lung injury aggravated by acute hyperglycemia.

Low Circulating Levels of GR, FKBP5, and SGK1 in Medicated Patients With Depression Are Not Altered by Electroconvulsive Therapy.

Hypothalamic-pituitary-adrenal axis dysregulation is frequently observed in patients with depression, with increased levels of the glucocorticoid (GC) cortisol commonly reported. Hypothalamic-pituitary-adrenal axis dysregulation may be a consequence of impaired feedback inhibition due to GC receptor (GR) impairments or dysfunction, termed "glucocorticoid resistance." Here, our objective was to assess mRNA levels of GC-related markers (GR, FKBP5, serum glucocorticoid kinase 1 [SGK1]) in patients with depression versus controls and in patient samples after electroconvulsive therapy (ECT). We also examined the relationship between these GC-related markers and 24-item Hamilton Depression Rating Scale (HAM-D24) scores to assess the utility of using them as biological markers for depression or the therapeutic response to ECT. GR, FKBP5, and SGK1 mRNA levels were examined in whole blood samples from 88 medicated patients with depression pre-/post-ECT and 63 controls using quantitative real-time polymerase chain reaction. Exploratory subgroup correlational analyses were performed to determine the relationship between GR, FKBP5, and SGK1 and 24-item Hamilton Depression Rating Scale scores. GR, FKBP5, and SGK1 mRNA levels were significantly lower in medicated patients with depression compared with controls (P < 0.001, P = 0.03, P < 0.001, respectively), but ECT did not alter their levels (all P > 0.05). There was no relationship between GR, FKBP5, or SGK1 and 24-item Hamilton Depression Rating Scale scores. GR, FKBP5, and SGK1 do not seem to be involved in the peripheral molecular response to ECT and do not represent useful biomarkers for predicting the therapeutic response to ECT in a real-world clinical setting.

Clinical characteristics of colorectal cancer patients and anti-neoplasm activity of genistein

BackgroundEpidemiologically, the disease incidence of colorectal cancer (CRC) ranks the third among all malignant tumors, and its mortality is the second following lung cancer. If unmanaged, CRC will develop fatal invasiveness and metastasis. However, existing chemotherapy is limitedly effective to treat metastatic CRC. Genistein, a functional phytoestrogen, is found with potent pharmacological activity against cancer cells. Therefore, this study was designed to characterize the clinical signatures of human CRC and to conduct anti-CRC experiments using genistein. MethodsBriefly, the plasma, tumor, non-tumor samples of CRC patients were harvested for biological experiments, followed by analysis of clinical data. A pharmacological study in vitro of genistein for treating CRC cells was conducted accordingly. ResultsIn diagnostic data, molecular tumor biomarkers in CRC patients were detected in plasma samples, consistent with pathological and imaging diagnoses of CRC. Notably, carcinomatous expressions of miR-95, serum glucocorticoid kinase 1 (SGK1), B-cell lymphoma-2 (Bcl-2), extracellular regulated protein kinase 1 (Erk1) in human CRC were notably elevated when compared to those in non-tumor controls. In pharmacological experiments using cell culture model, genistein-treated CRC cells resulted in reduced cellular viability, elevated lactate dehydrogenase (LDH) content, increased apoptotic cells and TdT mediated dUTP nick end labeling (TUNEL)-positive cells following a dose-dependent manner. Interestingly, down-regulated expressions of endogenous miR-95, SGK1, Bcl-2, Erk1 were observed after genistein treatments in a dose-dependent way. ConclusionsCollectively, the current clinical data indicate pathological markers of miR-95, SGK1, Erk1 in human CRC cases, and further experimental findings reveal that anti-CRC pharmacological mechanism using genistein was implicated in suppression of cellular miR-95, SGK1, Erk1 expressions. Together, genistein may be a promising bioactive compound for treating CRC.

High Salt Activates CD11c+ Antigen-Presenting Cells via SGK (Serum Glucocorticoid Kinase) 1 to Promote Renal Inflammation and Salt-Sensitive Hypertension.

Salt-sensing mechanisms in hypertension involving the kidney, vasculature, and central nervous system have been well studied; however, recent studies suggest that immune cells can sense sodium (Na+). Antigen-presenting cells (APCs) including dendritic cells critically modulate inflammation by activating T cells and producing cytokines. We recently found that Na+ enters dendritic cells through amiloride-sensitive channels including the α and γ subunits of the epithelial sodium channel (ENaC) and mediates nicotinamide adenine dinucleotide phosphate oxidase-dependent formation of immunogenic IsoLG (isolevuglandin)-protein adducts leading to inflammation and hypertension. Here, we describe a novel pathway in which the salt-sensing kinase SGK1 (serum/glucocorticoid kinase 1) in APCs mediates salt-induced expression and assembly of ENaC-α and ENaC-γ and promotes salt-sensitive hypertension by activation of the nicotinamide adenine dinucleotide phosphate oxidase and formation of IsoLG-protein adducts. Mice lacking SGK1 in CD11c+ cells were protected from renal inflammation, endothelial dysfunction, and developed blunted hypertension during the high salt feeding phase of the N-Nitro-L-arginine methyl ester hydrochloride/high salt model of salt-sensitive hypertension. CD11c+ APCs treated with high salt exhibited increased expression of ENaC-γ which coimmunoprecipitated with ENaC-α. This was associated with increased activation and expression of various nicotinamide adenine dinucleotide phosphate oxidase subunits. Genetic deletion or pharmacological inhibition of SGK1 in CD11c+ cells prevented the high salt-induced expression of ENaC and nicotinamide adenine dinucleotide phosphate oxidase. These studies indicate that expression of SGK1 in CD11c+ APCs contributes to the pathogenesis of salt-sensitive hypertension.

Serum Glucocorticoid Kinase 1 (SGK1) Expression in Dendritic Cells Contributes to Salt‐Induced Hypertension in Mice

Salt‐sensing mechanisms in hypertension involving the kidney have been well studied; however, recent studies suggest that immune cells can sense Na+. Antigen presenting cells like dendritic cells (DCs) critically modulate inflammation by activating T cells and producing cytokines. We recently found that sodium enters DCs through amiloride‐sensitive channels including the alpha and gamma subunits of the epithelial sodium channel (ENaCαγ) and mediates NADPH oxidase‐dependent formation of immunogenic isolevuglandin (IsoLG)‐protein adducts leading to inflammation and hypertension. Here we describe a novel pathway in which the salt sensing kinase serum/glucocorticoid kinase 1 (SGK1) in DCs mediates salt‐induced assembly of ENaCαγ and promotes salt‐sensitive hypertension by activation of the NADPH oxidase and formation of IsoLG‐protein adducts. Mice lacking SGK1 in CD11c+ cells were protected from renal inflammation, endothelial dysfunction and developed blunted hypertension during the high salt feeding phase of the N‐Nitro‐L‐arginine methyl ester hydrochloride (L‐NAME)/high salt model of salt‐sensitive hypertension. DCs treated with high salt exhibited increased expression of ENaCγ which co‐immunoprecipitated with ENaCα. This was associated with increased activation and expression of various NADPH oxidase subunits. Genetic deletion or pharmacological inhibition of SGK1 in DCs prevented the high salt induced expression of ENaCγ (0.04±0.04, WT, 0.05±0.04, WT plus HS and 0.06±0.05, KO plus HS, p&lt;0.01) and ENaCα (0.29±0.20, for WT, 0.28±0.19, for WT plus HS and 0.30±0.24, for KO plus HS, p&lt;0.01). These results indicate that expression of SGK1 in DCs contributes to the pathogenesis of salt‐sensitive hypertension by stabilizing expression of ENaCαγ leading to activation of NADPH oxidase.Support or Funding InformationThis work was supported by the American Heart Association grants POST290900 and SFRN204200, and National Institutes of Health grants K01HL130497, R01HL125865, and R01HL039006.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Axonal and Myelin Neuroprotection by the Peptoid BN201 in Brain Inflammation.

The development of neuroprotective therapies is a sought-after goal. By screening combinatorial chemical libraries using in vitro assays, we identified the small molecule BN201 that promotes the survival of cultured neural cells when subjected to oxidative stress or when deprived of trophic factors. Moreover, BN201 promotes neuronal differentiation, the differentiation of precursor cells to mature oligodendrocytes in vitro, and the myelination of new axons. BN201 modulates several kinases participating in the insulin growth factor 1 pathway including serum-glucocorticoid kinase and midkine, inducing the phosphorylation of NDRG1 and the translocation of the transcription factor Foxo3 to the cytoplasm. In vivo, BN201 prevents axonal and neuronal loss, and it promotes remyelination in models of multiple sclerosis, chemically induced demyelination, and glaucoma. In summary, we provide a new promising strategy to promote neuroaxonal survival and remyelination, potentially preventing disability in brain diseases.

The role of dietary sodium in autoimmune diseases: The salty truth.

Autoimmune diseases are a group of heterogeneous condition that occur secondary to the intrinsic loss of tolerance to self- antigens. In genetically susceptible individuals, the complex interplay of environmental factors and epigenetic deregulations have been proposed to drive disease etiopathogenesis. Various environmental variables have been identified including viral infections, exposure to pollutants, stress and dietary factors. Sodium, a major constituent of salt is essential for mammalian physiology. However, high salt intake may play a role in the development of autoimmune diseases. Several lines of evidence point toward the role of high sodium intake in reversing the suppressive effects of Regulatory T cells (Tregs) and instead promoting cellular shift toward T-helper (Th)-1 and Th17 pro-inflammatory phenotypes. These effects have been attributed to cascade of events that ultimately results in downstream activation of serum glucocorticoid kinase 1 (Sgk1). In vivo, various autoimmune animal models have confirmed the role of high sodium diet in the emergence and the exacerbation of autoimmune conditions including for instance Experimental Autoimmune Encephalomyelitis model for multiple sclerosis, MRL/lpr mouse model for lupus nephritis, collagen induced arthritis model for rheumatoid arthritis, and dextran sulfate sodium induced colitis, and TNBS-induced colitis models for Crohn's disease. Clinical epidemiological studies are scarce. High sodium intake was associated with increased risk of rheumatoid arthritis disease emergence. In multiple sclerosis, some studies suggest a relation to clinical exacerbation rates however other studies did not corroborate these results. Taken together, high dietary salt intake plays a role in the spectrum of autoimmune disease etiology. Further research is warranted to better characterize such relationship and assist in identifying individuals that would benefit from dietary salt restriction.

Abstract P214: Serum Glucocorticoid Kinase 1 and Sympathetic Nerve Activity in Salt-Sensitive Hypertension.

Salt-sensitive hypertension affects nearly 50% of the population and reducing salt intake decreases blood pressure and cardiovascular events in the general population. The precise mechanism of how dietary salt contributes to blood pressure (BP) elevation, renal injury, and cardiovascular disease remains unclear. Substantial evidence supports a role of increased sympathetic output in salt-dependent hypertension and this promotes renal inflammation and dysfunction. It has been shown that salt accumulates in the interstitium of hypertensive humans and animals and drives immune cells toward a proinflammatory phenotype through the salt sensing kinase serum/glucocorticoid kinase 1 (SGK1). We have also shown an important role of monocytes and monocyte-derived dendritic cells in hypertension. In this study, we tested the hypothesis that SGK1 in myeloid CD11c + cells promote salt-sensitive hypertension by increasing sympathetic outflow. To test this hypothesis, we created mice lacking SGK1 in CD11c + cells (SGK1 CreCD11c mice) and used SGK1 fl/fl mice as controls. To induce salt-sensitivity, mice received 0.5 mg/ml of N-Nitro-L-arginine methyl ester hydrochloride (L-NAME) in the drinking water for 2 weeks. This was followed by a 2-week washout period and then a 4% high salt diet for 3 weeks. BP was monitored using telemetry. We found that BP elevation during high salt feeding was significantly attenuated in SGK1 CreCD11c mice compared to SGK1 fl/fl mice (125 ± 1 vs. 141 ± 1 mmHg; p &lt; 0.01). SGK1 CreCD11c mice had a significant reduction in CD45 + , CD3 + , CD4 + CD8 + and CD19 + cells in the kidney compared to SGK1 fl/fl mice as assessed by flow cytometry (p &lt; 0.05). Interestingly, on high salt, SGK1 CreCD11c mice had a significant reduction in heart rate (HR) compared to SGK1 fl/fl mice (528 ± 2 vs. 592 ± 7 bpm; p = 0.006). Microglial cells of the brain express CD11c, and we found that the reduction in HR was associated with a marked reduction in HR variability in SGK1 CreCD11c mice compared to SGK1 fl/fl mice (1.65 ± 0.38 vs. 3.00 ± 0.57; p = 0.0018), indicating a reduction in sympathetic outflow. Our data indicate that SGK1 in CD11c + myeloid cells and likely those of the central nervous system, modulate BP and sympathetic outflow promoting the pathogenesis of salt-sensitive hypertension.

Abstract 127: Axl + Siglec6 + Dendritic Cells: the Role of Salt, Stretch, and Hypertension

We have shown that monocyte-derived dendritic cells (DCs) are activated in hypertension to produce large amounts of cytokines and to activate T cells. DCs from hypertensive mice can convey hypertension when adoptively transferred to recipients. Single cell sequencing has recently identified a novel subset of DCs in humans that express Axl and Sigelc6 + (AS DCs). These cells have been reported to potently drive T cell proliferation and to produce large amounts of IL-8 and IL12. The role of AS DCs in hypertension remains unknown. We isolated total peripheral blood mononuclear cells (PBMCs) from normotensive (n=23) and hypertensive (n=12) patients and assessed DC populations, including AS DCs, using flow cytometry. We found a significant increase in the AS DCs in hypertensive compared to normotensive patients (297 ± 73 vs. 108 ± 26/ml; P=0.0304). In contrast, there were no differences in CD1c + DCs (3398 ± 776 vs. 5245 ± 122/ml) or CD141 + DCs (164 ± 20 vs. 218 ± 49/ml) between normotensive and hypertensive subjects. To investigate the mechanism by which AS DCs are formed in hypertension, we used two in vitro hypertensive stimuli: exposure to salt and hypertensive stretch of adjacent human endothelial cells. Human PBMCs were cultured in either normal NaCl (NS, 150 mM) or high NaCl (HS, 190 mM) for 48 hours. Flow cytometry indicated a striking increase in AS DCs by exposure to HS compared to NS (516 ± 181 vs 201 ± 57/ml) and this was prevented by co-treatment of cells with the salt-sensing Serum Glucocorticoid Kinase 1 inhibitor GSK650394. As a second approach, we co-cultured human aortic endothelial cells (HAECs) with PBMCs from normotensive donors and exposed the HAEC monolayer to either normal (5%) or hypertensive cyclical stretch (10%) for 24 hours. Co-culture of PBMCs with HAECs exposed to 10% stretch doubled AS DCs as compared to PBMCs cultured with HAECs undergoing 5% stretch (1.4 ± 0.5 vs 0.7 ± 0.3%; P=0.0217). These data show that AS DC population are increased in hypertensive patients and that known hypertensive stimuli in vitro promote formation of AS DCs. Thus, AS DCs seem to be an important immune cell subset in human hypertension and might be a novel therapeutic target for this disease.

Could Sodium Chloride be an Environmental Trigger for Immune-Mediated Diseases? An Overview of the Experimental and Clinical Evidence.

Immune mediated diseases (IMDs) are complex chronic inflammatory diseases involving genetic and environmental factors. Salt intake has been proposed as a diet factor that can influence the immune response. Indeed, experimental data report the influence of sodium chloride on the differentiation of naive CD4+ T cells into IL-17 secreting T helper (Th) cells (Th17 cells), by a mechanism involving the serum glucocorticoid kinase-1 (SGK1) that promotes the expression of the IL-23 receptor (IL-23R). The IL-23/IL-23R is critical for pathogenic inflammatory Th17 cell differentiation. Experimental data in murine models of arthritis, colitis and encephalomyelitis corroborate these findings. This manuscript reviews the current knowledge on the effects of sodium chloride on innate and adaptive immunity. We also performed a systematic literature review for clinical studies examining the relationships between salt consumption and the development or the activity/severity of the most common IMDs mediated by the IL-23/Th17 pathway, i.e., rheumatoid arthritis (RA), multiple sclerosis (MS), and Crohn's disease (CD). Nine studies were found, 4 in RA, 4 in MS and 1 in CD. An association was found between developments of anti-citrullinated protein antibody (ACPA) positive RA in smokers and salt intake, but these results were not confirmed in another study. For MS, no association was observed in pediatric subjects while in adult patients, a link was found between salt intake and disease activity. However, this result was not confirmed in another study. These conflicting results highlight the fact that further evaluation in human IMDs is required. Moreover, physicians need to develop clinical trials with diet interventions to evaluate the impact of low salt intake on disease activity/severity of IMDs.

The Role of Salt, Serum Glucocorticoid Kinase 1, and NADPH Oxidase in Salt‐Sensitive Hypertension

Hypertension is a major modifiable risk for death and cardiovascular disease, yet its causes are largely unknown. Salt‐sensitive hypertension affects nearly 50% of the population and reducing salt intake decreases blood pressure and cardiovascular events in the general population. The precise mechanism of how dietary salt contributes to blood pressure elevation and cardiovascular disease remains unclear. We have previously shown that renal and vascular inflammation contributes to the development of hypertension and that NADPH oxidase‐dependent formation of immunogenic isolevuglandin (isoLG)‐protein adducts in dendritic cells (DCs) plays an important role in this hypertensive inflammation. Moreover, it has been shown that salt often accumulates in the interstitium of hypertensive humans and animals, and that such high salt concentrations drive immune cells toward a proinflammatory phenotype through the salt sensing kinase serum/glucocorticoid kinase 1 (SGK1). In the current study, we tested the hypothesis that activation of SGK1 by salt drives NADPH oxidase activation and promotes hypertension. To determine the effects of salt on DCs, mouse splenocytes were cultured in either normal salt (NS, 150 mM NaCl), high salt (HS, 190 mM NaCl), or HS + SGK1 inhibitor for 24 hours. Western blot of protein extracts from DCs indicated a striking increase in all NADPH oxidase subunits by exposure to HS and that these effects were prevented by inhibition of SGK1. To test whether SGK1 in DCs plays a role in N‐Nitro‐L‐arginine methyl ester hydrochloride (L‐NAME)/HS feeding, a model of salt‐sensitive hypertension, we created mice lacking SGK1 in CD11c+ cells (SGK1CreCD11c mice) and used SGK1fl/fl mice as controls. Mice were implanted with radiotelemeters to measure mean arterial pressure (MAP) and 10 days later placed on L‐NAME/HS feeding protocol. Briefly, mice are treated with L‐NAME (0.5 mg/ml) in the drinking water for two weeks. Mice received normal drinking water for 2 weeks, and were finally placed on a 4% HS diet for 3 weeks. Our preliminary data show that the % change in MAP after L‐NAME/HS feeding was significantly attenuated in SGK1CreCD11c mice compared to the SGK1fl/fl mice (115.7 ± 3.5 vs 135.4). Using flow cytometry, we assessed the effects of genetic deletion of SGK1 in DCs on vascular and renal inflammation. Preliminary data show that SGK1CreCD11c had a trend for a reduction in both aortic and renal inflammation compare to SGK1fl/fl, indicating a critical role pro‐inflammatory role for SGK1 in DCs during salt‐sensitive hypertension. Taken together, our data indicate that DCs can activated by exposure to a high salt environment which often exists in hypertension, and that this likely involves NADPH oxidase‐dependent formation of superoxide. Moreover, SGK1 in DCs may be important in the development and pathogenesis of salt‐sensitive hypertension. These studies define a novel role of SGK1 in DCs in salt‐sensitive hypertension.Support or Funding InformationThis work was supported by American Heart Association grants POST290900 and SFRN204200, National Institutes of Health grants K01HL130497, R01HL125865, R01HL039006, and 5T32HL069765‐13, and the National Center for Advancing Translational Sciences ( UL1TR000445 )This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.