Serum And Glucocorticoid-regulated Kinase 1 Research Articles

JAK/STAT signaling pathway affects CCR5 expression in human CD4+ T cells.

CCR5 serves as R5-tropic HIV co-receptor. Knocking out CCR5 in HIV patients, which has occurred <10 times, is believed important for cure. JAK/STAT inhibitors tofacitinib and ruxolitinib inhibit CCR5 expression in HIV+ viremic patients. We investigated the association of JAK/STAT signaling pathway with CCR5/CCR2 expression in human primary CD4+ T cells and confirmed its importance. Six of nine JAK/STAT inhibitors that reduced CCR5/CCR2 expression were identified. Inhibitor-treated CD4+ T cells were relatively resistant, specifically to R5-tropic HIV infection. Furthermore, single JAK2, STAT3, STAT5A, and STAT5B knockout and different combinations of JAK/STAT knockout significantly reduced CCR2/CCR5 expression of both RNA and protein levels, indicating that CCR5/CCR2 expression was positively regulated by JAK-STAT pathway in CD4+ T cells. Serum and glucocorticoid-regulated kinase 1 (SGK1) knockout affected CCR2/CCR5 gene expression, suggesting that SGK1 is involved in CCR2/CCR5 regulation. If cell surface CCR5 levels can be specifically and markedly down-regulated without adverse effects, that may have a major impact on the HIV cure agenda.

Combined Effects of Polystyrene Nanosphere and Homosolate Exposures on Estrogenic End Points in MCF-7 Cells and Zebrafish.

Micro- and nanoplastics (MNPs) and homosalate (HMS) are ubiquitous emerging environmental contaminants detected in human samples. Despite the well-established endocrine-disrupting effects (EDEs) of HMS, the interaction between MNPs and HMS and its impact on HMS-induced EDEs remain unclear. This study aimed to investigate the influence of MNPs on HMS-induced estrogenic effects and elucidate the underlying mechanisms in vitro and invivo. We assessed the impact of polystyrene nanospheres (PNSs; , ) on HMS-induced MCF-7 cell proliferation (HMS: , equivalent to ) using the E-SCREEN assay and explored potential mechanisms through transcriptomics. Adult zebrafish were exposed to HMS () with or without PNSs (, ) for 21 d. EDEs were evaluated through gonadal histopathology, fertility tests, steroid hormone synthesis, and gene expression changes in the hypothalamus-pituitary-gonad-liver (HPGL) axis. Coexposure of HMS and PNSs resulted in higher expression of estrogen receptor (ESR1) and the mRNAs of target genes (pS2, AREG, and PGR), a greater estrogen-responsive element transactivation activity, and synergistic stimulation on MCF-7 cell proliferation. Knockdown of serum and glucocorticoid-regulated kinase 1 (SGK1) rescued the MCF-7 cell proliferation induced by PNSs alone or in combination with HMS. In zebrafish, coexposure showed higher expression of SGK1 and promoted ovary development but inhibited spermatogenesis. In addition, coexposure led to lower egg hatchability, higher embryonic mortality, and greater larval malformation. Coexposure also modulated steroid hormone synthesis genes (cyp17a2, hsd17[Formula: see text]1, esr2b, vtg1, and vtg2), and resulted in higher () release in females. Conversely, males showed lower testosterone, , and gene expressions of cyp11a1, cyp11a2, cyp17a1, cyp17a2, and hsd17[Formula: see text]1. PNS exposure exacerbated HMS-induced estrogenic effects via SGK1 up-regulation in MCF-7 cells and disrupting the HPGL axis in zebrafish, with gender-specific patterns. This offers new mechanistic insights and health implications of MNP and contaminant coexposure. https://doi.org/10.1289/EHP13696.

Adipose c-Jun NH2-terminal kinase promotes angiotensin II-induced and deoxycorticosterone acetate salt-induced hypertension and vascular dysfunction by inhibition of adiponectin production and activation of SGK1 in mice.

Adipose c-Jun NH2-terminal kinase 1/2 (JNK1/2) is a central mediator involved in the development of obesity and its complications. However, the roles of adipose JNK1/2 in hypertension remain elusive. Here we explored the role of adipose JNK1/2 in hypertension. The roles of adipose JNK1/2 in hypertension were investigated by evaluating the impact of adipose JNK1/2 inactivation in both angiotensin II (Ang II)-induced and deoxycorticosterone acetate (DOCA) salt-induced hypertensive mice. Specific inactivation of JNK1/2 in adipocytes significantly alleviates Ang II-induced and DOCA salt-induced hypertension and target organ damage in mice. Interestingly, such beneficial effects are also observed in hypertensive mice after oral administration of JNK1/2 inhibitor SP600125. Mechanistically, adipose JNK1/2 acts on adipocytes to reduce the production of adiponectin (APN), then leads to promote serum and glucocorticoid-regulated kinase 1 (SGK1) phosphorylation and increases epithelial Na + channel α-subunit (ENaCα) expression in both renal cells and adipocytes, respectively, finally exacerbates Na + retention. In addition, chronic treatment of recombinant mouse APN significantly augments the beneficial effects of adipose JNK1/2 inactivation in DOCA salt-induced hypertension. By contrast, the blood pressure-lowering effects of adipose JNK1/2 inactivation are abrogated by adenovirus-mediated SGK1 overexpression in Ang II -treated adipose JNK1/2 inactivation mice. Adipose JNK1/2 promotes hypertension and targets organ impairment via fine-tuning the multiorgan crosstalk among adipose tissue, kidney, and blood vessels.

Abstract P1162: Nedd4-2 Inactivation Underlies The Increase In Cardiac Sodium-dependent Glucose Cotransporter-1 In Diabetic Hearts

Rationale: Type-2 diabetes (T2D) is a major risk factor for developing heart failure. While the underlying mechanisms are poorly understood, recent evidence suggests that upregulation of the Na + /glucose cotransporter 1 (SGLT1) plays an important role, as it causes myocyte Na + overload. Objective: To identify the mechanisms involved in promoting an increase in SGLT1 protein level in diabetic hearts. Methods/Results: We compared hearts from patients with and without T2D and hearts from rats with late-onset T2D caused by overexpression of human amylin in pancreatic β-cells versus hearts from wild-type littermates. SGLT1 protein level was significantly higher in hearts from both humans and rats with T2D. In contrast, the SGLT1 mRNA levels were comparable in hearts from T2D and non-diabetic patients and rats. This result suggests that theT2D-induced increase in cardiac SGLT1 protein occurs at a post-transcriptional level. Co-immunoprecipitation of SGLT1 with ubiquitin was reduced in diabetic human and rat hearts, which indicates that SGLT1 degradation is impaired. The E3 ubiquitin ligase Nedd4-2 is responsible for ubiquitination of several cardiac membrane transporters. Pharmacological inhibition of Nedd4-2 with Heclin in HL-1 cells resulted in significantly higher SGLT1 protein levels, suggesting that Nedd4-2 also mediates SGLT1 degradation. Nedd4-2 is inactivated upon phosphorylation by several kinases, including the serum and glucocorticoid-regulated kinase-1 (SGK1). Using western blotting, we found increased Nedd4-2 phosphorylation in hearts from humans and rats with T2D compared to their non-diabetic counterparts. Moreover, hearts from T2D patients exhibited higher levels of SGK1 and its activated phosphorylated form compared to their non-diabetic counterparts. Conclusion/New Hypothesis: The increase in SGLT1 protein in the diabetic heart is regulated post-transcriptionally by an impairment in Nedd4-2 dependent ubiquitination and degradation of SGLT1.

SGK1 Target Genes Involved in Heart and Blood Vessel Functions in PC12 Cells.

Serum and glucocorticoid-regulated kinase 1 (SGK1) is expressed in neuronal cells and involved in the pathogenesis of hypertension and metabolic syndrome, regulation of neuronal function, and depression in the brain. This study aims to identify the cellular mechanisms and signaling pathways of SGK1 in neuronal cells. In this study, the SGK1 inhibitor GSK650394 is used to suppress SGK1 expression in PC12 cells using an in vitro neuroscience research platform. Comparative transcriptomic analysis was performed to investigate the effects of SGK1 inhibition in nervous cells using mRNA sequencing (RNA-seq), differentially expressed genes (DEGs), and gene enrichment analysis. In total, 12,627 genes were identified, including 675 and 2152 DEGs at 48 and 72 h after treatment with GSK650394 in PC12 cells, respectively. Gene enrichment analysis data indicated that SGK1 inhibition-induced DEGs were enriched in 94 and 173 genes associated with vascular development and functional regulation and were validated using real-time PCR, Western blotting, and GEPIA2. Therefore, this study uses RNA-seq, DEG analysis, and GEPIA2 correlation analysis to identify positive candidate genes and signaling pathways regulated by SGK1 in rat nervous cells, which will enable further exploration of the underlying molecular signaling mechanisms of SGK1 and provide new insights into neuromodulation in cardiovascular diseases.

An electrostatically conjugated-functional MNK1 aptamer reverts the intrinsic antitumor effect of polyethyleneimine-coated iron oxide nanoparticles in vivo in a human triple-negative cancer xenograft

BackgroundTriple-negative breast cancer (TNBC) remains a difficult breast cancer subtype to treat as it exhibits a particularly aggressive behavior. The dysregulation of distinct signaling pathways underlies this aggressive behavior, with an overactivation of MAP kinase interacting kinases (MNKs) promoting tumor cell behavior, and driving proliferation and migration. Therefore, MNK1 is an excellent target to impair the progression of TNBC and indeed, an MNK1-specific aptamer has proved to be efficient in inhibiting TBNC cell proliferation in vitro. Although polyethyleneimine-coated iron oxide nanoparticles (PEI–IONPs) have been used as transfection and immunomodulating agents, no study has yet addressed the benefits of using these nanoparticles as a magnetic carrier for the delivery of a functional aptamer.ResultsHere, we tested the antitumor effect of a PEI–IONP complexed to the functional MNK1b-specific aptamer in vitro and in vivo. We demonstrated that these apMNKQ2@PEI–IONP nanoconjugates delivered three times more apMNKQ2 to MDA-MB-231 cells than the aptamer alone, and that this enhanced intracellular delivery of the aptamer had consequences for MNK1 signaling, reducing the amount of MNK1 and its target the phospho(Ser209)-eukaryotic initiation factor 4E (eIF4E). As a result, a synergistic effect of the apMNKQ2 and PEI–IONPs was observed that inhibited MDA-MB-231 cell migration, probably in association with an increase in the serum and glucocorticoid-regulated kinase-1 (SGK1) and the phospho(Thr346)-N-myc down-regulated gene 1 (NDRG1). However, intravenous administration of the apMNKQ2 alone did not significantly impair tumor growth in vivo, whereas the PEI–IONP alone did significantly inhibit tumor growth. Significantly, tumor growth was not inhibited when the apMNKQ2@PEI–IONP nanocomplex was administered, possibly due to fewer IONPs accumulating in the tumor. This apMNKQ2-induced reversion of the intrinsic antitumor effect of the PEI–IONPs was abolished when an external magnetic field was applied at the tumor site, promoting IONP accumulation.ConclusionsElectrostatic conjugation of the apMNKQ2 aptamer with PEI–IONPs impedes the accumulation of the latter in tumors, which appears to be necessary for PEI–IONPs to exert their antitumor activity.Graphical

Early Interleukin-6 Mediated Activation of SGK1 is Primarily via Variant 6

Hypertension is associated with inflammation, including elevated interleukin-6 (IL-6) levels. IL-6 is an inflammatory cytokine, which we have previously shown to activate the mineralocorticoid receptor (MR) and the downstream sodium (Na) transporters- Na-chloride cotransporter (NCC) and the epithelial Na channel (ENaC). However, how IL-6 activates the MR is still unknown. IL-6 is also linked to Serum and Glucocorticoid-regulated Kinase 1 (SGK1), which is a downstream effector of the MR. SGK1 regulates both NCC and ENaC, increasing activity and tubular reabsorption of Na. This increased Na reabsorption favors water retention, thus increasing blood pressure. Previously, IL-6 has also been linked with SGK1 activation. We hypothesize that IL-6 increases SGK1 mRNA and protein expression in renal epithelial cells. To test this, we used in vitro renal epithelial cell models of both the late distal convoluted tubule (DCT2) and inner medullary collecting duct (IMCD), mDCT15 and IMCD3, respectively. Cells were treated with an IL-6 time-course (100ng/ml for 1, 3 and 6 hours). There are 6 known transcriptional variants for SGK1. We designed variant-specific PCR primers and, a pilot study based on regular PCR concluded that expression levels of only variants 5 and 6 (V5, V6) are significant. Using Real Time quantitative RT-PCR, we were then able to quantify transcript levels of both SGK1 V5 and V6 variants. With IL-6 treatment, we observed a 160%-211% increase at early time points (30mins-1hr) in SKG1 V6 and a lesser response (26%-88% increase) with V5 as compared to vehicle treated mDCT15 cells. Since SGK1 has been observed in the membrane fraction and in the cytoplasm, protein extracts from cells were fractionated (mPER kit) to assess membrane and cytosolic SGK1 protein levels using immunoblots. Densitometric analysis of immunoblots demonstrated a 5.5- and 3.1-fold increase in SGK1 protein expression at 1hr in mDCT15 and IMCD3 cells, respectively. These data suggest that IL-6 induces increases in SGK1 transcription and translation. Interestingly, we also found that majority of SGK1 was in the membrane fraction and the level was consistent. Together, these data suggest that SGK1 V6 may be responsible for the increased sodium reabsorption in response to IL-6 stimulation in renal epithelial cells. NIH funding This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Berberine attenuates sunitinib-induced cardiac dysfunction by normalizing calcium regulation disorder via SGK1 activation.

Sunitinib (SNT)-induced cardiotoxicity is associated with abnormal calcium regulation caused by phosphoinositide 3 kinase inhibition in the heart. Berberine (BBR) is a natural compound that exhibits cardioprotective effects and regulates calcium homeostasis. We hypothesized that BBR ameliorates SNT-induced cardiotoxicity by normalizing the calcium regulation disorder via serum and glucocorticoid-regulated kinase 1 (SGK1) activation. Mice, neonatal rat cardiomyocytes (NRVMs), and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were used to study the effects of BBR-mediated SGK1 activity on the calcium regulation disorder caused by SNT as well as the underlying mechanism. BBR offered prevention against SNT-induced cardiac systolic dysfunction, QT interval prolongation, and histopathological changes in mice. After the oral administration of SNT, the Ca2+ transient and contraction of cardiomyocytes was significantly inhibited, whereas BBR exhibited an antagonistic effect. In NRVMs, BBR was significantly preventive against the SNT-induced reduction of calcium transient amplitude, prolongation of calcium transient recovery, and decrease in SERCA2a protein expression; however, SGK1 inhibitors resisted the preventive effects of BBR. In hiPSC-CMs, BBR pretreatment significantly prevented SNT from inhibiting the contraction, whereas coincubation with SGK1 inhibitors antagonized the effects of BBR. These findings indicate that BBR attenuates SNT-induced cardiac dysfunction by normalizing the calcium regulation disorder via SGK1 activation.

SGK1 inhibition attenuated the action potential duration in patient- and genotype-specific re-engineered heart cells with congenital long QT syndrome

Long QT syndrome (LQTS) stems from pathogenic variants in KCNQ1 (LQT1), KCNH2 (LQT2), or SCN5A (LQT3) and is characterized by action potential duration (APD) prolongation. Inhibition of serum and glucocorticoid regulated kinase-1 (SGK1) is proposed as a novel therapeutic for LQTS. The study sought to test the efficacy of novel, selective SGK1 inhibitors in induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) models of LQTS. The mexiletine (MEX)-sensitive SCN5A-P1332L iPSC-CMs were tested initially compared with a CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 SCN5A-P1332L variant-corrected isogenic control (IC). The SGK1-I1 therapeutic efficacy, compared with MEX, was tested for APD at 90% repolarization (APD90) shortening in SCN5A-P1332L, SCN5A-R1623Q, KCNH2-G604S, and KCNQ1-V254M iPSC-CMs using FluoVolt. The APD90 was prolonged in SCN5A-P1332L iPSC-CMs compared with its IC (646 ± 7 ms vs 482 ± 23 ms; P < .0001). MEX shortened the APD90 to 560 ± 7 ms (52% attenuation, P < .0001). SGK1-I1 shortened the APD90 to 518 ± 5 ms (78% attenuation, P < .0001) but did not shorten the APD90 in the IC. SGK1-I1 shortened the APD90 of the SCN5A-R1623Q iPSC-CMs (753 ± 8 ms to 475 ± 19 ms compared with 558 ± 19 ms with MEX), the KCNH2-G604S iPSC-CMs (666 ± 10 ms to 574 ± 18 ms vs 538 ± 15 ms after MEX), and the KCNQ1-V254M iPSC-CMs (544 ± 10 ms to 475 ± 11ms; P = .0004). Therapeutically inhibiting SGK1 effectively shortens the APD in human iPSC-CM models of the 3 major LQTS genotypes. These preclinical data support development of SGK1 inhibitors as novel, first-in-class therapy for patients with congenital LQTS.

SGK1 inhibition attenuates the action potential duration in reengineered heart cell models of drug-induced QT prolongation

Drug-induced QT prolongation (DI-QTP) is a clinical entity in which administration of a human ether-à-go-go-related gene/rapid delayed rectifier potassium current blocker such as dofetilide prolongs the cardiac action potential duration (APD) and the QT interval on the electrocardiogram. Inhibition of serum and glucocorticoid regulated kinase-1 (SGK1) reduces the APD at 90% repolarization (APD90) in induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) derived from patients with congenital long QT syndrome. Here, we test the efficacy of 2 novel SGK1 inhibitors-SGK1-I1 and SGK1-I2-in iPSC-CM models of dofetilide-induced APD prolongation. Normal iPSC-CMs were treated with dofetilide to produce a DI-QTP iPSC-CM model. SGK1-I1's and SGK1-I2's therapeutic efficacy for shortening the dofetilide-induced APD90 prolongation was compared to mexiletine. The APD90 values were recorded 4hours after treatment using a voltage-sensing dye. The APD90 was prolonged in normal iPSC-CMs treated with dofetilide (673 ± 8 ms vs 436 ± 4 ms; P < .0001). While 10 mM mexiletine shortened the APD90 of dofetilide-treated iPSC-CMs from 673 ± 4 to 563 ± 8 ms (46% attenuation; P < .0001), 30 nM of SGK1-I1 shortened the APD90 from 673 ± 8 to 502 ± 7 ms (72% attenuation; P < .0001). Additionally, 300 nM SGK1-I2 shortened the APD90 of dofetilide-treated iPSC-CMs from 673 ± 8 to 460 ± 7 ms (90% attenuation; P < .0001). These novel SGK1-Is substantially attenuated the pathological APD prolongation in a human heart cell model of DI-QTP. These preclinical data support the development of this therapeutic strategy to counter and neutralize DI-QTP, thereby increasing the safety profile for patients receiving drugs with torsadogenic potential.

Serum and glucocorticoid-regulated kinase 1: Structure, biological functions, and its inhibitors.

Serum and glucocorticoid-regulated kinase 1 (SGK1) is a serine/threonine kinase belonging to the protein kinase A, G, and C (AGC) family. Upon initiation of the phosphoinositide 3-kinase (PI3K) signaling pathway, mammalian target of rapamycin complex 2 (mTORC2) and phosphoinositide-dependent protein kinase 1 (PDK1) phosphorylate the hydrophobic motif and kinase domain of SGK1, respectively, inducing SGK1 activation. SGK1 modulates essential cellular processes such as proliferation, survival, and apoptosis. Hence, dysregulated SGK1 expression can result in multiple diseases, including hypertension, cancer, autoimmunity, and neurodegenerative disorders. This review provides a current understanding of SGK1, particularly in sodium transport, cancer progression, and autoimmunity. In addition, we summarize the developmental status of SGK1 inhibitors, their structures, and respective potencies evaluated in pre-clinical experimental settings. Collectively, this review highlights the significance of SGK1 and proposes SGK1 inhibitors as potential drugs for treatment of clinically relevant diseases.

Abstract 11027: SGK1 Inhibition and Attenuation of the Action Potential Duration in Re-Engineered Heart Cell Models of Drug-Induced QT Prolongation

Background: Drug-induced QT prolongation (DI-QTP) is a clinical entity in which administration of a HERG/I Kr blocker such as dofetilide prolongs the cardiac action potential duration (APD) at the cellular level and the QT interval clinically, and increases the risk for a potentially lethal, QT-triggered ventricular arrhythmia. There may be a role for inhibition of I NaL to counter DI-QTP. Recently, we have shown that inhibition of serum and glucocorticoid regulated kinase-1 (SGK1) reduces the APD90 in induced pluripotent stem cell derived cardiomyocytes (iPSC-CM) from patients with LQT3 and ameliorates increases in I NaL . Objective: To test the efficacy of novel SGK1 inhibitors (SGK-I) in re-engineered cardiomyocyte models of dofetilide-induced APD prolongation. Methods: Normal iPSC-CMs were treated with dofetilide [5 nM] to produce a DI-QTP iPSC-CM model. The SGK1-I’s therapeutic efficacy for shortening the dofetilide-induced APD90 prolongation was compared to mexiletine. The APD90 values were recorded 4 hours after treatment using FluoVolt, a voltage-sensitive fluorescent dye. Results: The APD90 was significantly prolonged in normal iPSC-CMs treated with dofetilide (673 ± 8 ms with dofetilide vs 436 ± 4 ms with DMSO, p&lt;0.0001). While 10 μM mexiletine shortened the average APD90 of dofetilide-treated normal iPSC-CMs from 673 ± 4 ms to 563 ± 8 ms (46% attenuation, p&lt;0.0001), 30 nM of SGK-I shortened the APD90 from 673 ± 4 ms to 478 ± 10 ms (82% attenuation, p&lt;0.0001). Conclusions: Therapeutically inhibiting SGK1 effectively shortens the cardiomyocyte APD in a human heart cell model of DI-QTP. The novel SGK1 inhibitor normalized the pathological APD prolongation almost fully (&gt; 80%) in the iPSC-CM model treated with dofetilide. This pre-clinical data supports further development of this therapeutic strategy to counter and neutralize the threat of DI-QTP.

Abstract 10986: SGK1 Inhibition Attenuated the Action Potential Duration In-Patient and Genotype-Specific Re-Engineered Heart Cells With Congenital Long QT Syndrome

Background: Long QT syndrome (LQTS) often stems from pathogenic variants in KCNQ1 (LQT1), KCNH2 (LQT2), or SCN5A (LQT3) and is characterized cellularly by prolongation of the cardiac action potential duration (APD). We have shown that inhibition of serum and glucocorticoid regulated kinase-1 (SGK1) reduces I NaL and is proposed as a novel therapeutic strategy for LQT3. Objective: To test the efficacy of a potent and selective SGK1 inhibitor (SGK1-I) in human cardiomyocyte models of LQT1, LQT2, and LQT3. Methods: Induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) were generated from patients with either LQT1, LQT2, or LQT3. The mexiletine (MEX)-sensitive SCN5A-P1332L iPSC-CMs were tested initially. A CRISPR/Cas9 P1332L variant-corrected isogenic control (IC) was used as a control. The SGK1-I’s therapeutic efficacy was compared to MEX and to a commercially available SGK1 inhibitor (EMD638683). The novel SGK-I was then tested for APD shortening in SCN5A-R1623Q, KCNH2-G604S and KCNQ1-V254M iPSC-CMs. The APD90 values were recorded 4 hours after treatment using FluoVolt. Results: The APD90 was significantly prolonged in SCN5A-P1332L iPSC-CMs compared to its IC (646 ± 7 ms vs 482 ± 23 ms, p&lt;0.0001). MEX shortened the average APD90 to 560 ± 7 ms (52% attenuation). Interestingly, SGK1-I significantly shortened the APD to 518 ± 5 ms (78% attenuation) but did not further shorten the APD in the IC. SGK1-I also shortened the APD90 of the SCN5A-R1623Q iPSC-CMs from 753 ± 8 ms to 475 ± 19 ms compared to 558 ± 19 ms with MEX. The SGK1-I shortened the APD90 in KCNH2-G604S (666 ± 10ms to 574 ± 18ms for SGK1-I versus 538 ± 15 ms after MEX). EMD had limited APD shortening effects in SCN5A and KCNH2 iPSC-CMs. Interestingly, while neither MEX nor EMD reduced the APD90 in the KCNQ1-V254M iPSC-CMs, the novel SGK1-I reduced the APD90 from 544 ± 10 ms to 475 ± 11ms (p=0.0004). Conclusions: Therapeutically inhibiting SGK1 effectively shortens the cardiomyocyte APD in human heart cell models of the 3 major LQTS genotypes. The novel SGK1-I normalized the pathological APD prolongation almost fully (&gt; 70%) in the patient-derived SCN5A-P1332L iPSC-CM model. These pre-clinical data support further development of SGK1-I as a novel, first-in-class therapy for patients with congenital LQTS.

Long non-coding RNA VPS9D1-AS1 enhances proliferation, invasion, and epithelial-mesenchymal transition in endometrial cancer via miR-377-3p/SGK1.

Endometrial cancer (EC) is a kind of gynecologic malignancy with a rising incidence rate. This study aimed to explore the role of VPS9D1 antisense RNA1 (VPS9D1-AS1) in EC. The expression of VPS9D1-AS1, microRNA (miR)-377-3p, and serum and glucocorticoid-regulated kinase 1 (SGK1) was detected by Quantitative Real-Time PCR (qRT-PCR). Cell proliferation, invasion and epithelial-mesenchymal transition (EMT) were determined by cell counting kit-8 (CCK-8), 5-Ethynyl-2'-Deoxyuridine (EdU) transwell, and western bolt. VPS9D1-AS1 was predicted to sponge miR-377-3p via Starbase, and verified by luciferase reporter, RNA binding protein immunoprecipitation (RIP), and RNA pull-down experiments. The clinical characteristics of VPS9D1-AS1, miR-377-3p, and SGK1 were analyzed. The role of VPS9D1-AS1 on EC tumorigenesis was assessed in xenografted nude mice. VPS9D1-AS1 was upregulated in EC cells and tissues. Interference of VPS9D1-AS1 inhibited growth, invasion, and EMT of EC cells. Mechanically, VPS9D1-AS1 was a molecular sponge of miR-377-3p, and overexpression of miR-377-3p reversed VPS9D1-AS1-induced EC cells proliferation, invasion, and EMT. Moreover, SGK1 was confirmed to bind with miR-377-3p. Furthermore, overexpression of SGK1 alleviated sh-VPS9D1-AS1-caused effects on EC cells. High level of VPS9D1-AS1 and SGK1, or low miR-377-3p expression predicted a poor prognosis. The expression of the three genes was correlated with lymph node metastasis, pathological stage, and International Federation of Gynecology and Obstetrics (FIGO) stage, but not associated with age, ER, and PR expression. Interestingly, knockdown of VPS9D1-AS1 suppressed EC tumor growth in mice. VPS9D1-AS1 promoted cell invasion, proliferation, and EMT via modulating miR-377-3p/SGK1 axis, which provided new options for therapeutic strategies of EC.

Dapagliflozin attenuates diabetes-induced diastolic dysfunction and cardiac fibrosis by regulating SGK1 signaling

BackgroundRecent studies have reported improved diastolic function in patients administered sodium-glucose cotransporter 2 inhibitors (SGLT2i). We aimed to investigate the effect of dapagliflozin on left ventricular (LV) diastolic function in a diabetic animal model and to determine the molecular and cellular mechanisms underlying its function.MethodsA total of 30 male New Zealand white rabbits were randomized into control, diabetes, or diabetes+dapagliflozin groups (n = 10/per each group). Diabetes was induced by intravenous alloxan. Cardiac function was evaluated using echocardiography. Myocardial samples were obtained for histologic and molecular evaluation. For cellular evaluation, fibrosis-induced cardiomyoblast (H9C2) cells were obtained, and transfection was performed for mechanism analysis (serum and glucocorticoid-regulated kinase 1 (SGK1) signaling analysis).ResultsThe diabetes+dapagliflozin group showed attenuation of diastolic dysfunction compared with the diabetes group. Dapagliflozin inhibited myocardial fibrosis via inhibition of SGK1 and epithelial sodium channel (ENaC) protein, which was observed both in myocardial tissue and H9C2 cells. In addition, dapagliflozin showed an anti-inflammatory effect and ameliorated mitochondrial disruption. Inhibition of SGK1 expression by siRNA decreased and ENaC and Na+/H+ exchanger isoform 1 (NHE1) expression was confirmed as significantly reduced as siSGK1 in the diabetes+dapagliflozin group.ConclusionsDapagliflozin attenuated left ventricular diastolic dysfunction and cardiac fibrosis via regulation of SGK1 signaling. Dapagliflozin also reduced macrophages and inflammatory proteins and ameliorated mitochondrial disruption.

Identification of Novel Inhibitors Targeting SGK1 via Ensemble-Based Virtual Screening Method, Biological Evaluation and Molecular Dynamics Simulation.

Serum and glucocorticoid-regulated kinase 1 (SGK1), as a serine threonine protein kinase of the AGC family, regulates different enzymes, transcription factors, ion channels, transporters, and cell proliferation and apoptosis. Inhibition of SGK1 is considered as a valuable approach for the treatment of various metabolic diseases. In this investigation, virtual screening methods, including pharmacophore models, Bayesian classifiers, and molecular docking, were combined to discover novel inhibitors of SGK1 from the database with 29,158 compounds. Then, the screened compounds were subjected to ADME/T, PAINS and drug-likeness analysis. Finally, 28 compounds with potential inhibition activity against SGK1 were selected for biological evaluation. The kinase inhibition activity test revealed that among these 28 hits, hit15 exhibited the highest inhibition activity against SGK1, which gave 44.79% inhibition rate at the concentration of 10 µM. In order to further investigate the interaction mechanism of hit15 and SGK1 at simulated physiological conditions, a molecular dynamics simulation was performed. The molecular dynamics simulation demonstrated that hit15 could bind to the active site of SGK1 and form stable interactions with key residues, such as Tyr178, ILE179, and VAL112. The binding free energy of the SGK1-hit15 was −48.90 kJ mol−1. Therefore, the identified hit15 with novel scaffold may be a promising lead compound for development of new SGK1 inhibitors for various diseases treatment.

Estrogen-increased SGK1 Promotes Endometrial Stromal Cell Invasion in Adenomyosis by Regulating with LPAR2.

Adenomyosis is an estrogen-dependent gynecological disorder. The abnormal migration and invasion of the eutopic endometrium is thought to be the primary role in the pathogenesis of adenomyosis. However, the exact underlying mechanism remains unclear. This study investigated involvement of serum and glucocorticoid-regulated kinase 1 (SGK1) in the pathogenesis of adenomyosis. The SGK1 expression level was higher in the eutopic endometrium of adenomyosis. Upregulation of SGK1 can promote the migration, invasion of human stromal endometrial cells (HESC). Through RNA sequencing and other technical methods, we found that SGK1 regulates the expression of the important downstream molecule Lysophosphatidic acid receptor 2 (LPAR2), and ultimately regulates the expression level of functional proteins such as matrix metalloproteinase 2 and matrix metalloproteinase 9, which are related to migration and invasion. Then, we found that 17β-estradiol (E2) upregulated the expression of SGK1 in endometrial cells in a dose-dependent manner. Furthermore, SGK1 shRNA significantly suppressed the migration and invasion induced by E2 in endometrial cells, as well as the related factors. Our study revealed the possible role of SGK1 in the migration and invasion in the development of adenomyosis.

Identification of serum and glucocorticoid-regulated kinase 1 as a regulator of signal transducer and activator of transcription 3 signaling

Signal transducer and activator of transcription 3 (STAT3) plays key roles in cancer cell proliferation, invasion, and immunosuppression. In many human cancer cells, STAT3 is hyperactivated, which leads to tumor progression and drug resistance, and therefore STAT3 and its modulators are considered effective drug targets. However, the complex regulatory mechanisms of STAT3 have made it difficult to develop potent anticancer drugs that suppress its activity. Here, we report serum and glucocorticoid-regulated kinase 1 (SGK1) as a novel regulator of STAT3 signaling and an effective target for combination therapy with Janus kinase (JAK) inhibitors. We screened small molecules using a gain-of-function mutant of STAT3 resistant to JAK inhibition and found that an SGK1 inhibitor suppressed the constitutive activation of STAT3. Importantly, our results revealed that SGK1 also mediated the activation of wild-type STAT3. Further examination suggested that the tuberous sclerosis complex 2 and mammalian target of rapamycin signaling pathway were involved in STAT3 activation by SGK1. Finally, we demonstrated that SGK1 inhibition enhanced the inhibitory effect of a JAK inhibitor on STAT3 phosphorylation and cancer cell proliferation. Our findings provide new insights into the molecular mechanisms of STAT3 activation and suggest SGK1 as a potential target for STAT3-targeted combination cancer therapy.

Upregulation of serum and glucocorticoid-regulated kinase 1 (SGK1) ameliorates doxorubicin-induced cardiotoxic injury, apoptosis, inflammation and oxidative stress by suppressing glucose regulated protein 78 (GRP78)-mediated endoplasmic reticulum stress

ABSTRACT The clinical application of doxorubicin (Dox) in tumor chemotherapy is limited by time-dependent and dose-dependent cardiotoxicity. Hence, there is an urgent need to elucidate doxorubicin cardiotoxicity and to solve the difficult problem in clinical application. It has been verified that serum and glucocorticoid-regulated kinase 1 (SGK1) possess cardioprotective effects. Here, H9c2 cells were treated with 1 μM doxorubicin for 24 h to establish doxorubicin cardiotoxicity, so as to determine the biological role of SGK1 in doxorubicin cardiomyopathy and to elucidate the underlying molecular mechanism. SGK1 level in doxorubicin-treated H9c2 cells was assessed by performing Western blot assay and RT-qPCR. CCK-8 assay and TUNEL staining were employed to evaluate the cell viability and cell apoptosis. Besides, apoptosis-related proteins were measured by Western blot assay to analyze cell apoptosis. Additionally, the release of TNF-α, IL-1β, IL-6, and IL-10 and the levels of ROS, MDA, and SOD were detected to reflect inflammation and oxidative stress. Moreover, Western blot assay was adopted for determination of ERS-associated proteins. Results revealed that SGK1 was downregulated in doxorubicin-treated H9c2 cells. Upregulation of SGK1 alleviated doxorubicin-induced cardiotoxic injury, cell apoptosis, inflammation and oxidative stress in H9c2 cells. Moreover, SGK1 overexpression mitigated doxorubicin-induced ERS in H9c2 cells. The suppressing effects of SGK1 on doxorubicin-induced cardiotoxic injury, apoptosis, inflammation, oxidative stress and ERS in H9c2 cells were partially abolished upon GRP78 overexpression. To conclude, upregulation of SGK1 may alleviate doxorubicin cardiotoxicity by repressing GRP78-mediated ERS.

SGK1.1 isoform is involved in nociceptive modulation, offering a protective effect against noxious cold stimulus in a sexually dimorphic manner

The serum and glucocorticoid-regulated kinase 1 (SGK1) is a widely expressed protein in the Central Nervous System (CNS), involved in regulating the activity of a wide variety of ion channels and transporters and physiological functions, such as neuronal excitability. SGK1.1 is a neuronal splice isoform of SGK1, expressed exclusively in the CNS, distributed in brain and cerebellum, that decreases neuronal excitability via up-regulation of M-current, linked to Kv7.2/3 potassium channels. Strategies to maintain increased SGK1.1 activity could be helpful in decreasing neuronal hyperexcitability, as occurs in neuropathic pain. Transgenic mice overexpressing SGK1.1 (B6.Tg.sgk1) offer a particularly relevant opportunity to assess the physiological involvement of this protein in nociception.Behavior and physiological nociception were evaluated in male and female B6.Tg.sgk1 and wild-type mice (B6.WT), characterizing nociceptive thresholds to different nociceptive stimuli (thermal, chemical and mechanical), as well as the electrophysiological properties of cutaneous sensory Aδ-fibres isolated from the saphenous nerve. The acute antinociceptive effect of morphine was also evaluated.Compared with B6.WT animals, male and female B6.Tg.sgk1 mice showed increased spontaneous locomotor activity. Regarding nociception, there were no differences between transgenic and wild-type mice in heat, chemical and mechanical thresholds, but interestingly, male B6.Tg.sgk1 mice were less sensitive to cold stimulus; B6.Tg.sgk1 animals showed lower sensitivity to morphine. Electrophysiological properties of cutaneous primary afferent fibres were maintained.This is the first demonstration that the SGK1.1 isoform is involved in nociceptive modulation, offering a protective effect against noxious cold stimulus in a sexually dimorphic manner. B6.Tg.sgk1 mice offer a particularly relevant opportunity to further analyze the involvement of this protein in nociception, and studies in models of chronic, neuropathic pain are warranted.