APLNR Regulates IFN-γ signaling via ß-arrestin 1 mediated JAK-STAT1 pathway in melanoma cells.

The apelin receptor (APLNR) regulates many biological processes including metabolism, angiogenesis, circulating blood volume and cardiovascular function. Additionally, APLNR is overexpressed in various types of cancer and influences cancer progression. APLNR is reported to regulate tumor recognition during immune surveillance by modulating the IFN-γ response. However, the mechanism of APLNR cross-talk with intratumoral IFN-γ signaling remains unknown. Here, we show that activation of APLNR up-regulates IFN-γ signaling in melanoma cells through APLNR mediated ß-arrestin 1 but not ß-arrestin 2 recruitment. Our data suggests that ß-arrestin 1 directly interacts with STAT1 to inhibit STAT1 phosphorylation to attenuate IFN-γ signaling. The APLNR mutant receptor, I109A, which is deficient in ß-arrestins recruitment, is unable to enhance intratumoral IFN-γ signaling. While APLNR N112G, a constitutively active mutant receptor, increases intratumoral sensitivity to IFN-γ signaling by enhancing STAT1 phosphorylation upon IFN-γ exposure. We also demonstrate in a co-culture system that APLNR regulates tumor survival rate. Taken together, our findings reveal that APLNR modulates IFN-γ signaling in melanoma cells and suggest that APLNR may be a potential target to enhance the efficacy of immunotherapy.

(-)-Epicatechin Is a Biased Ligand of Apelin Receptor.

(-)-Epicatechin (EC) is part of a large family of biomolecules called flavonoids and is widely distributed in the plant kingdom. Several studies have shown the beneficial effects of EC consumption. Many of these reported effects are exerted by activating the signaling pathways associated with the activation of two specific receptors: the G protein-coupled estrogen receptor (GPER), a transmembrane receptor, and the pregnane X receptor (PXR), which is a nuclear receptor. However, the effects of EC are so diverse that these two receptors cannot describe the complete phenomenon. The apelin receptor or APLNR is classified within the G protein-coupled receptor (GPCR) family, and is capable of activating the G protein canonical pathways and the ß-arrestin transducer, which participates in the phenomenon of receptor desensitization and internalization. ß-arrestin gained interest in selective pharmacology and mediators of the so-called "biased agonism". With molecular dynamics (MD) and in vitro assays, we demonstrate how EC can recruit the ß-arrestin in the active conformation of the APLN receptor acting as a biased agonist.

Identification of potent pyrazole based APELIN receptor (APJ) agonists.

The apelinergic system comprises the apelin receptor and its cognate apelin and elabela peptide ligands of various lengths. This system has become an increasingly attractive target for pulmonary and cardiometabolic diseases. Small molecule regulators of this receptor with good drug-like properties are needed. Recently, we discovered a novel pyrazole based small molecule agonist 8 of the apelin receptor (EC50 = 21.5 µM, Ki = 5.2 µM) through focused screening which was further optimized to initial lead 9 (EC50 = 0.800 µM, Ki = 1.3 µM). In our efforts to synthesize more potent agonists and to explore the structural features important for apelin receptor agonism, we carried out structural modifications at N1 of the pyrazole core as well as the amino acid side-chain of 9. Systematic modifications at these two positions provided potent small molecule agonists exhibiting EC50 values of <100 nM. Recruitment of ß-arrestin as a measure of desensitization potential of select compounds was also investigated. Functional selectivity was a feature of several compounds with a bias towards calcium mobilization over ß-arrestin recruitment. These compounds may be suitable as tools for in vivo studies of apelin receptor function.

Apelin Controls Angiogenesis-Dependent Glioblastoma Growth.

Glioblastoma (GBM) present with an abundant and aberrant tumor neo-vasculature. While rapid growth of solid tumors depends on the initiation of tumor angiogenesis, GBM also progress by infiltrative growth and vascular co-option. The angiogenic factor apelin (APLN) and its receptor (APLNR) are upregulated in GBM patient samples as compared to normal brain tissue. Here, we studied the role of apelin/APLNR signaling in GBM angiogenesis and growth. By functional analysis of apelin in orthotopic GBM mouse models, we found that apelin/APLNR signaling is required for in vivo tumor angiogenesis. Knockdown of tumor cell-derived APLN massively reduced the tumor vasculature. Additional loss of the apelin signal in endothelial tip cells using the APLN-knockout (KO) mouse led to a further reduction of GBM angiogenesis. Direct infusion of the bioactive peptide apelin-13 rescued the vascular loss-of-function phenotype specifically. In addition, APLN depletion massively reduced angiogenesis-dependent tumor growth. Consequently, survival of GBM-bearing mice was significantly increased when APLN expression was missing in the brain tumor microenvironment. Thus, we suggest that targeting vascular apelin may serve as an alternative strategy for anti-angiogenesis in GBM.

The sinus venosus contributes to coronary vasculature through VEGFC-stimulated angiogenesis.

Identifying coronary artery progenitors and their developmental pathways could inspire novel regenerative treatments for heart disease. Multiple sources of coronary vessels have been proposed, including the sinus venosus (SV), endocardium and proepicardium, but their relative contributions to the coronary circulation and the molecular mechanisms regulating their development are poorly understood. We created an ApjCreER mouse line as a lineage-tracing tool to map SV-derived vessels onto the heart and compared the resulting lineage pattern with endocardial and proepicardial contributions to the coronary circulation. The data showed a striking compartmentalization to coronary development. ApjCreER-traced vessels contributed to a large number of arteries, capillaries and veins on the dorsal and lateral sides of the heart. By contrast, untraced vessels predominated in the midline of the ventral aspect and ventricular septum, which are vessel populations primarily derived from the endocardium. The proepicardium gave rise to a smaller fraction of vessels spaced relatively uniformly throughout the ventricular walls. Dorsal (SV-derived) and ventral (endocardial-derived) coronary vessels developed in response to different growth signals. The absence of VEGFC, which is expressed in the epicardium, dramatically inhibited dorsal and lateral coronary growth but left vessels on the ventral side unaffected. We propose that complementary SV-derived and endocardial-derived migratory routes unite to form the coronary vasculature and that the former requires VEGFC, revealing its role as a tissue-specific mediator of blood endothelial development.

Bioactivity of the putative apelin proprotein expands the repertoire of apelin receptor ligands.

BACKGROUND: Apelin is a peptide ligand for a class A G-protein coupled receptor called the apelin receptor (AR or APJ) that regulates angiogenesis, the adipoinsular axis, and cardiovascular functions. Apelin has been shown to be bioactive as 13, 17, and 36 amino acid isoforms, C-terminal fragments of the putatively inactive 55-residue proprotein (proapelin or apelin-55). Although intracellular proprotein processing has been proposed, isolation of apelin-55 from colostrum and milk demonstrates potential for secretion prior to processing and the possibility of proapelin-AR interaction. METHODS: Apelin isoform activity and potency were compared by an In-Cell Western™ assay for ERK phosphorylation using a stably AR-transfected HEK293A cell line. Conformational comparison of apelin isoforms was carried out by circular dichroism and heteronuclear solution-state nuclear magnetic resonance spectroscopy. RESULTS: Apelin-55 is shown to activate the AR, with similar maximum ERK phophorylation response and potency to the shorter isoforms except for apelin-13, which exhibited a greater potency. Correlating to this shared activity, highly similar conformations are exhibited in all apelin isoforms for the shared C-terminal region responsible for receptor binding and activation. CONCLUSIONS: AR activation by all apelin isoforms likely hinges upon shared conformation and dynamics in the C-terminus, with apelin-55 providing an alternative bioactive isoform despite the addition of 19N-terminal residues relative to apelin-36. GENERAL SIGNIFICANCE: Beyond providing novel insight into the physiology of this system, re-annotation of proapelin to the bioactive apelin-55 isoform adds to the molecular toolkit for dissection of apelin-AR interactions and expands the repertoire of therapeutic targets for the apelinergic system.

A Study of GWAS-Supported Variants of rs9943582 in a Chinese Han Population with Ischemic Stroke: No Associations with Disease Onset and Clinical Outcomes.

BACKGROUND: The variant rs9943582 of APLNR (apelin receptor) was identified by a large-scale study to be associated with an increased risk of ischemic stroke in a Japanese population. We conducted this study to investigate the association between the variant and age of onset and clinical outcomes of ischemic stroke in a Chinese population. METHODS: Improved multiple ligase detection reaction was used to genotype the variant. We compared the mean age at ischemic stroke onset with one-way ANOVA. The Kaplan-Meier method, log-rank test, and Cox proportional hazards regression models were performed to analyze the association between the variant and clinical outcomes (recurrence and death). RESULTS: A total of 916 ischemic stroke patients were recruited for the study. For age at ischemic stroke onset, no significant association was identified with the variant in any genetic model. In addition, the variant was not strongly associated with recurrence and death risk of ischemic stroke, as shown by the results. CONCLUSIONS: The findings indicated that the variant rs9943582 was not associated with age at onset and clinical outcomes of ischemic stroke. However, evidence from well-designed studies with larger and in different ethnic populations are warranted to further explore the effects of APLNR on the ischemic stroke onset and clinical outcomes.

Polymorphisms of APLN-APLNR system are associated with essential hypertension in Mexican-Mestizo individuals.

The aim of this study was to evaluate if polymorphisms of APLN and APLNR genes may play a role as susceptibility markers for hypertension in a group of Mexican-Mestizo patients. A case-control study was carried out including normotensive and hypertensive individuals. For these, two polymorphisms of APLN (rs3761581 and rs56204867) and two of APLNR () genes were genotyped by 5' exonuclease TaqMan assay in 400 normotensive individuals and 383 patients. The results showed that, under an additive model adjusted by BMI, HDL, triglycerides, glucose and family history of essential hypertension, the rs7119375 and rs10501367 polymorphisms of APLNR gene were associated significantly with a decreased risk of essential hypertension (P=0.039 and P=0.029, respectively). Besides, the haplotypes analysis of these polymorphisms showed that H1 haplotype was associated with an increased risk of essential hypertension (P=0.026), while the H2 haplotype was associated with a decreased risk (P=0.032). Contrary, the rs3761581 and rs56204867 polymorphisms of APLN gene were not associated with essential hypertension (P=0.1707 and P=0.0769, respectively). The data suggest that APLNR rs7119375 and rs10501367 are associated with a decreased risk of essential hypertension in our Mexican-Mestizo studied group, but further studies are warranted.

Discovery of a novel small molecule agonist scaffold for the APJ receptor.

The apelinergic system includes a series of endogenous peptides apelin, ELABELA/TODDLER and their 7-transmembrane G-protein coupled apelin receptor (APJ, AGTRL-1, APLNR). The APJ receptor is an attractive therapeutic target because of its involvement in cardiovascular diseases and potentially other disorders including liver fibrosis, obesity, diabetes, and neuroprotection. To date, pharmacological characterization of the APJ receptor has been limited due to the lack of small molecule functional agonists or antagonists. Through focused screening we identified a drug-like small molecule agonist hit 1 with a functional EC50 value of 21.5±5µM and binding affinity (Ki) of 5.2±0.5µM. Initial structure-activity studies afforded compound 22 having a 27-fold enhancement in potency and the first sub-micromolar full agonist with an EC50 value of 800±0.1nM and Ki of 1.3±0.3µM. Preliminary SAR, synthetic methodology, and in vitro pharmacological characterization indicate this scaffold will serve as a favorable starting point for further refinement of APJ potency and selectivity.

Small proteins, big roles: the signaling protein Apela extends the complexity of developmental pathways in the early zebrafish embryo.

The identification of molecules controlling embryonic patterning and their functional analysis has revolutionized the fields of Developmental and Cell Biology. The use of new sequence information and modern bioinformatics tools has enriched the list of proteins that could potentially play a role in regulating cell behavior and function during early development. The recent application of efficient methods for gene knockout in zebrafish has accelerated the functional analysis of many proteins, some of which have been overlooked due to their small size. Two recent publications report on the identification of one such protein and its role in zebrafish embryogenesis. The protein, currently designated Apela, was shown to act as a secreted protein whose absence adversely affected various early developmental processes. Additional signaling proteins that have been identified in one of the studies are likely to open the way to unraveling hitherto unknown developmental pathways and have the potential to provide a more comprehensive understanding of known developmental processes.

Apelin promotes diabetic nephropathy by inducing podocyte dysfunction via inhibiting proteasome activities.

Podocyte injuries are associated with progression of diabetic nephropathy (DN). Apelin, an adipocyte-derived peptide, has been reported to be a promoting factor for DN. In this study, we aim to determine whether apelin promotes progression of DN by inducing podocyte dysfunction. kk-Ay mice were used as models for DN. Apelin and its antagonist, F13A were intraperitoneally administered for 4 weeks, respectively. Renal function and foot process proteins were analysed to evaluate the effects of apelin on kk-Ay mice and podocytes. Apelin increased albuminuria and decreased podocyte foot process proteins expression in kk-Ay mice, which is consistent with the results that apelin receptor (APLNR) levels increased in glomeruli of patients or mice with DN. In cultured podocytes, high glucose increased APLNR expression and apelin administration was associated with increased permeability and decreased foot process proteins levels. All these dysfunctions were associated with decreased 26S proteasome activities and increased polyubiquitinated proteins in both kk-Ay mice and cultured podocytes, as demonstrated by 26S proteasome activation with cyclic adenosine monophosphate (cAMP) or oleuropein. These effects seemed to be related to endoplasmic reticulum (ER) stress, as apelin increased C/EBP homologous protein (CHOP) and peiFα levels while cAMP or oleuropein reduced it in high glucose and apelin treated podocytes. These results suggest that apelin induces podocyte dysfunction in DN through ER stress which was induced by decreased proteasome activities in podocytes.

The apelin receptor: physiology, pathology, cell signalling, and ligand modulation of a peptide-activated class A GPCR.

The apelin receptor (AR or APJ) is a class A (rhodopsin-like) G-protein-coupled receptor with wide distribution throughout the human body. Activation of the AR by its cognate peptide ligand, apelin, induces diverse physiological effects including vasoconstriction and dilation, strengthening of heart muscle contractility, angiogenesis, and regulation of energy metabolism and fluid homeostasis. Recently, another endogenous peptidic activator of the AR, Toddler/ELABELA, was identified as having a crucial role in zebrafish (Danio rerio) embryonic development. The AR is also implicated in pathologies including cardiovascular disease, diabetes, obesity, and cancer, making it a promising therapeutic target. Despite its established importance, the precise roles of AR signalling remain poorly understood. Moreover, little is known about the mechanisms of peptide-AR activation. Additional complexity arises from modulation of the AR by 2 endogenous peptide ligands, both with multiple bioactive isoforms of variable length and distribution. The various apelin and Toddler/ELABELA isoforms may also produce distinct cellular effects. Further complexity arises through formation of functionally distinct heterodimers between the AR and other G-protein-coupled receptors. This minireview outlines key (patho)physiological actions of the AR, addresses what is known about signal transduction downstream of AR activation, and concludes by discussing unique properties of the endogenous peptidic ligands of the AR.

The Role of Aplnr Signaling in the Developmental Regulation of Mesenchymal Stem Cell Differentiation from Human Pluripotent Stem Cells.

Stem cells are invaluable resources for personalized medicine. Mesenchymal stem cells (MSCs) have received great attention as therapeutic tools due to being a safe, ethical, and accessible option with immunomodulatory and controlled differentiation properties. Apelin receptor (Aplnr) signaling is reported to be involved in biological events, including gastrulation, mesoderm migration, proliferation of MSCs. However, the knowledge about the exact role and mechanism of Aplnr signaling during mesoderm and MSCs differentiation is still primitive. The current study aims to unveil the role of Aplnr signaling during mesoderm and MSC differentiation from pluripotent stem cells (PSCs) through peptide/small molecule activation, overexpression, knock down or CRISPR/Cas9 mediated knock out of the pathway components. Morphological changes, gene and protein expression analysis, including antibody array, LC/MS, mRNA/miRNA sequencing, reveal that Aplnr signaling promotes mesoderm commitment possibly via EGFR and TGF-beta signaling pathways and enhances migration of cells during mesoderm differentiation. Moreover, Aplnr signaling positively regulates MSCs differentiation from hPSCs and increases MSC characteristics and differentiation capacity by regulating pathways, such as EGFR, TGFß, Wnt, PDGF, and FGF. Osteogenic, chondrogenic, adipogenic, and myogenic differentiations are significantly enhanced with Aplnr signaling activity. This study generates an important foundation to generate high potential MSCs from PSCs to be used in personalized cell therapy.

Blocking Palmitoylation of Apelin Receptor Alleviates Morphine Tolerance in Neuropathic Cancer Pain.

Neuropathic cancer pain (NCP) is an important symptom in patients with cancer. However, significant analgesic tolerance and other side effects critically hamper the administration of morphine. Protein palmitoylation mediated by the DHHC family may be involved in the glial activation and inflammatory responses underlying organ failure. In this study, we investigated the key role of protein palmitoylation in cancer pain and sought to target palmitoylation to suppress morphine tolerance. We found that long-term use of morphine led to the accumulation of the morphine metabolite, morphine-3-glucuronide, in vivo and activated ERK1/2 and microglia to release inflammatory factors through the apelin receptor APLNR. Palmitoyltransferase ZDHHC9 was upregulated in NCP, and APLNR was palmitylated to protect it from lysosomal degradation and to maintain its stability. We also designed competitive inhibitors of APLNR palmitoylation to inhibit the development of NCP, release of inflammatory factors, and attenuation of morphine tolerance. Therefore, targeting APLNR palmitoylation in combination with morphine is a potent method for cancer pain treatment. Our data provide a basis for the future clinical use of related drugs combined with morphine for the treatment of cancer-related pain.

Adaptive enzyme-responsive self-assembling multivalent apelin ligands for targeted myocardial infarction therapy.

Microvascular dysfunction following myocardial infarction exacerbates coronary flow obstruction and impairs the preservation of ventricular function. The apelinergic system, known for its pleiotropic effects on improving vascular function and repairing ischemic myocardium, has emerged as a promising therapeutic target for myocardial infarction. Despite its potential, the natural apelin peptide has an extremely short circulating half-life. Current apelin analogs have limited receptor binding efficacy and poor targeting, which restricts their clinical applications. In this study, we utilized an enzyme-responsive peptide self-assembly technique to develop an enzyme-responsive small molecule peptide that adapts to the expression levels of matrix metalloproteinases in myocardial infarction lesions. This peptide is engineered to respond to the high concentration of matrix metalloproteinases in the lesion area, allowing for precise and abundant presentation of the apelin motif. The changes in hydrophobicity allow the apelin motif to self-assemble into a supramolecular multivalent peptide ligand-SAMP. This self-assembly behavior not only prolongs the residence time of apelin in the myocardial infarction lesion but also enhances the receptor-ligand interaction through increased receptor binding affinity due to multivalency. Studies have demonstrated that SAMP significantly promotes angiogenesis after ischemia, reduces cardiomyocyte apoptosis, and improves cardiac function. This novel therapeutic strategy offers a new approach to restoring coronary microvascular function and improving damaged myocardium after myocardial infarction.

APLNR inhibited nasopharyngeal carcinoma growth and immune escape by downregulating PD-L1.

BACKGROUND: APLNR is a G protein-coupled receptor and our previous study had revealed that APLNR could inhibit nasopharyngeal carcinoma (NPC) growth and metastasis. However, the role of APLNR in regulating PD-L1 expression and immune escape in NPC is unknown. METHODS: We analyzed the expression and correlation of APLNR and PD-L1 in NPC tissues and cells. We investigated the effect of APLNR on PD-L1 expression and the underlying mechanism in vitro and in vivo. We also evaluated the therapeutic potential of targeting APLNR in combination with PD-L1 antibody in a nude mouse xenograft model. RESULTS: We found that APLNR was negatively correlated with PD-L1 in NPC tissues and cells. APLNR could inhibit PD-L1 expression by binding to the FERM domain of JAK1 and blocking the interaction between JAK1 and IFNGR1, thus suppressing IFN-γ-mediated activation of the JAK1/STAT1 pathway. APLNR could also inhibit NPC immune escape by enhancing IFN-γ secretion and CD8+ T-cell infiltration and reducing CD8+ T-cell apoptosis and dysfunction. Moreover, the best effect was achieved in inhibiting NPC growth in nude mice when APLNR combined with PD-L1 antibody. CONCLUSIONS: Our study revealed a novel mechanism of APLNR regulating PD-L1 expression and immune escape in NPC and suggested that APLNR maybe a potential therapeutic target for NPC immunotherapy.

The vascular footprint in cardiac homeostasis and hypertensive heart disease-A link between apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase.

Recent studies have suggested a connection between disturbances of the apelin system and various cardiac pathologies, including hypertension, heart failure, and atherosclerosis. Vascular endothelial growth factor is crucial for cardiac homeostasis as a critical molecule in cardiac angiogenesis. Neuronal nitric oxide synthase is an essential enzyme producing nitric oxide, a key regulator of vascular tone. The present study aims to shed light upon the complex interactions between these three vital signaling molecules and examine their changes with the progression of hypertensive heart disease. We used two groups of spontaneously hypertensive rats and age-matched Wistar rats as controls. The expression of the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase were assessed immunohistochemically. We used capillary density and cross-sectional area of the cardiomyocytes as quantitative parameters of cardiac hypertrophy. Immunoreactivity of the molecules was more potent in both ventricles of spontaneously hypertensive rats compared with age-matched controls. However, capillary density was lower in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. In addition, the cross-sectional area of the cardiomyocytes was higher in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. Our study suggests a potential link between the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase in cardiac homeostasis and the hypertensive myocardium. Nevertheless, further research is required to better comprehend these interactions and their potential therapeutic implications.

Single-Cell RNA-Sequencing Analyses Identify APLNR, INS-IGF2, RGCC Genes May Be Involved in the Pathogenesis of Systemic Sclerosis Skin.

Background: Systemic sclerosis represents a persistent autoimmune disorder marked with fibrosis affecting both skin and other organs, which leads to a diminished quality of life and increased mortality. The affected skin provides a valuable opportunity to explore the pathogenesis of systemic sclerosis. Nevertheless, the roles of various cell populations within scleroderma remain intricate. Methods: We conducted a comprehensive reanalysis of recently published single-cell RNA-sequencing data from skin tissue cells in scleroderma. Through the utilization of Seurat, irGSEA, AUCell packages, and WGCNA analysis, we aimed to unveil crucial genes associated with the disease's etiological factors. Our investigation involved the characterization of heterogeneous pathway activities in both healthy and SSc-affected skin. Furthermore, we employed immunofluorescence techniques to validate the expression patterns of hub genes and differentially expressed genes. Results: The Endothelial-to-Mesenchymal Transition (EndMT) pathway was upregulated in SSc skin. Notably, the M4 module within Endothelial cell subpopulation 1 exhibited a strong association with EndMT. Furthermore, we identified three overexpressed genes (APLNR, INS-IGF2, RGCC) that demonstrated a significant correlation with EndMT. Importantly, their expression levels were markedly higher in skin of individuals with SSc when compared to healthy controls. Conclusion: APLNR, INS-IGF2 and RGCC serve as potential key players in the pathogenesis of SSc skin through EndMT-dependent mechanisms.

DNMT1 mediates proliferation, migration and invasion of extravillous trophoblasts by regulating the methylation level of APLNR.

INTRODUCTION: Proliferation, migration and invasion of extravillous trophoblasts (EVTs) play an important role in the progression of preeclampsia (PE). The purpose of this study was to investigate the molecular mechanism by which DNA methylase regulates the transcription level of APLNR and affects the phenotypic function of EVTs. MATERIALS AND METHODS: PE mice model and H/R model in HTR8/Svneo cells were constructed. Clinical samples of normal pregnant women and PE patients were collected. Expression and methylation level of APLNR in vivo and in vitro were detected. ChIP-qPCR was used to detect the binding of DNA methyltransferase at the APLNR promoter. The expression of DNA methyltransferase 1 (DNMT1), NO and eNOS in vitro were detected. EVTs proliferation, migration and invasion in vitro were detected. RESULTS: In placental tissues or HTR8/Svneo cells of the PE model group, the expression of APLNR was reduced and APLNR methylation level was up-regulated. There was no significant difference in the APLNR expression in placental tissues between normal pregnant women and PE patients. H/R conditions only promote the binding of DNMT1 at the APLNR promoter. DNMT1 interference decreased the enrichment degree of DNMT1 in APLNR promoter region and up-regulated the mRNA and protein levels of APLNR in vivo and in vitro. The activation of APLNR by Elabela (ELA) can promote eNOS transcription, thereby promoting cell proliferation and NO level, while eNOS inhibitor can reverse this effect. DNMT1 down-regulation inhibted APLNR methylation level, promoted eNOS transcription, and promoted EVTs proliferation, migration and invasion, which could be revised by the interference of APLNR. DISCUSSION: DNMT1 promotes eNOS transcription by inhibting APLNR methylation level, and promotes EVTs proliferation, migration and invasion, thus providing a new and broad application prospect for PE treatment.

Variations in protein levels of the apelinergic system in adipose tissue of hypertensive individuals with class 3 obesity.

The aim of this study was to investigate the expression of apelin (APLN) and its receptor (APLNR) in visceral adipose tissue (VAT), and its effect on the downstream expression of endothelial nitric oxide synthase (eNOS) in individuals with class 3 obesity, with or without hypertension. Seventy-five unrelated individuals presenting obesity class 3 with or without hypertension were included. Gene expression of APLN, and APLNR were analyzed in VAT, by reverse transcription quantitative polymerase chain reaction. The APLN, APLNR and eNOS (total and phosphorylated) levels in VAT were evaluated by Western blot. Analysis of differences between groups of APLN, APLNR and eNOS were performed by a logistic regression adjusting by confounding factors. Forty-five individuals with hypertension formed the case group, and 30 individuals constituted the control group. The APLN mRNA and protein levels were higher in the group of individuals with hypertension versus individuals without hypertension (p = 0.027 and p = 0.036, respectively). Meanwhile, APLNR mRNA and protein levels in subjects with hypertension were lower versus the group of subjects without hypertension (p = 0.001 and p = 0.008, respectively). Further, the group with hypertension presented a lower level of phosphorylation of eNOS Ser1177, compared to the control group (p = 0.002). In conclusion, individuals with class 3 obesity and hypertension present a modified APLN/APLNR expression in visceral adipose tissue, which could be secondary to reduced eNOS phosphorylation.