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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Células Endoteliales , Hipertensión Pulmonar , Receptor Notch4 , Transducción de Señal , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Células Endoteliales/metabolismo , Células Endoteliales/patología , Hipertensión Pulmonar/metabolismo , Hipertensión Pulmonar/patología , Animales , Humanos , Receptor Notch4/metabolismo , Receptor Notch4/genética , Reprogramación Celular , Capilares/metabolismo , Capilares/patología , Ratones , Arteria Pulmonar/metabolismo , Arteria Pulmonar/patología , Arteria Pulmonar/citologíaRESUMEN
Internal tandem duplication mutations of FLT3 (FLT3/ITD) confer poor prognosis in AML. FLT3 tyrosine kinase inhibitors (TKIs) alone have limited and transient clinical efficacy thus calling for new targets for more effective combination therapy. In a loss-of-function RNAi screen, we identified NOTCH4 as one such potential target whose inhibition proved cytotoxic to AML cells, and also sensitized them to FLT3 inhibition. Further investigation found increased NOTCH4 expression in FLT3/ITD AML cell lines and primary patient samples. Inhibition of NOTCH4 by shRNA knockdown, CRISPR-Cas9-based knockout or γ-secretase inhibitors synergized with FLT3 TKIs to kill FLT3/ITD AML cells in vitro. NOTCH4 inhibition sensitized TKI-resistant FLT3/ITD cells to FLT3 TKI inhibition. The combination reduced phospho-ERK and phospho-AKT, indicating inhibition of MAPK and PI3K/AKT signaling pathways. It also led to changes in expression of genes involved in regulating cell cycling, DNA repair and transcription. A patient-derived xenograft model showed that the combination reduced both the level of leukemic involvement of primary human FLT3/ITD AML cells and their ability to engraft secondary recipients. In summary, these results demonstrate that NOTCH4 inhibition synergizes with FLT3 TKIs to eliminate FLT3/ITD AML cells, providing a new therapeutic target for AML with FLT3/ITD mutations.
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Leucemia Mieloide Aguda , Inhibidores de Proteínas Quinasas , Receptor Notch4 , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa 3 Similar a fms , Humanos , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Leucemia Mieloide Aguda/metabolismo , Animales , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Ratones , Receptor Notch4/genética , Mutación , Línea Celular Tumoral , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: Alzheimer's Disease (AD) is the most common form of dementia, affecting cognitive and behavioral functions. AD is a complex disease resulting from the modest effect of gene interaction and environmental factors, as a result of which the exact pathogenesis is still unknown. AIM: The aim of the present study was to investigate the association between variants of 98 targeted genes with Alzheimer's disease phenotype. METHOD: A total of 98 genes from 32 AD cases and 11 controls were genotyped using the Haloplex target enrichment method and the PCR-RFLP approach.Association analysis was performed using the PLINK tool to identify the variant significantly associated with AD. Functional enrichment analysis and network analysis was performed using ClueGo and String database respectively. The Expression Quantitative Trait Loci (eQTL) analysis using the Genotype Tissue Expression (GTEx) dataset to explore the possible implication of the variant on the expression of one or more genes in different brain regions and whole blood. RESULT: Association analysis showed significant association of 19 variant assigned to 16 genes with Alzheimer's with p-value < 0.05 with rs367398/NOTCH4 only variant that passed multiple test corrections. Functional enrichment analysis showed association of these genes with AD. ClueGo and network analysis utilizing the String database suggested that genes are directly and indirectly linked to the AD pathogenesis. eQTL analysis revealed that the rs367398/NOTCH4 and rs1799806/ACHE variant showed significant eQTL for the neighbouring genes. CONCLUSION: The present study showed the possible role of 16 genes in AD pathogenesis, especially highlighting the role of rs367398/NOTCH4 and rs1799806/ACHE. However further investigation with large cohort is required to study and validate the implication of these variants in the AD pathogenesis.
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Enfermedad de Alzheimer , Humanos , Enfermedad de Alzheimer/genética , Epistasis Genética , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Genotipo , Polimorfismo de Nucleótido Simple , Sitios de Carácter Cuantitativo , Receptor Notch4/genéticaRESUMEN
Resistance to epidermal growth factor tyrosine kinase inhibitors (EGFR-TKI) remains one of the major challenges in lung adenocarcinoma (LUAD) therapy. Here, we find an increased frequency of the L12_16 amino acid deletion mutation in the signal peptide region of NOTCH4 (NOTCH4ΔL12_16) in EGFR-TKI-sensitive patients. Functionally, exogenous induction of NOTCH4ΔL12_16 in EGFR-TKI -resistant LUAD cells sensitizes them to EGFR-TKIs. This process is mainly mediated by the reduction of the intracellular domain of NOTCH4 (NICD4) caused by the NOTCH4ΔL12_16 mutation, which results in a lower localization of NOTCH4 in the plasma membrane. Mechanistically, NICD4 transcriptionally upregulates the expression of HES1 by competitively binding to the gene promoter relative to p-STAT3. Because p-STAT3 can downregulate the expression of HES1 in EGFR-TKI-resistant LUAD cells, the reduction of NICD4 induced by NOTCH4ΔL12_16 mutation leads to a decrease in HES1. Moreover, inhibition of the NOTCH4-HES1 pathway using inhibitors and siRNAs abolishes the resistance of EGFR-TKI. Overall, we report that the NOTCH4ΔL12_16 mutation sensitizes LUAD patients to EGFR-TKIs through transcriptional down-regulation of HES1 and that targeted blockade of this signaling cohort could reverse EGFR-TKI -resistance in LUAD, providing a potential approach to overcome resistance to EGFR-TKI -therapy.
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Adenocarcinoma del Pulmón , Neoplasias Pulmonares , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Receptores ErbB/metabolismo , Regulación hacia Abajo , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Adenocarcinoma del Pulmón/tratamiento farmacológico , Adenocarcinoma del Pulmón/genética , Mutación , Factor de Transcripción HES-1/genética , Factor de Transcripción HES-1/metabolismo , Receptor Notch4/genéticaRESUMEN
INTRODUCTION: Brain arteriovenous malformations (BAVMs) are high-flow intracranial vascular malformations characterized by the direct connection of arteries to veins without an intervening capillary bed. They are one of the main causes of intracranial hemorrhage and epilepsy, although morbidity is low. Angiogenesis, heredity, inflammation, and arteriovenous malformation syndromes play important roles in BAVM formation. Animal experiments and previous studies have confirmed that NOTCH4 may be associated with BAVM development. Our study identifies a connection between NOTCH4 gene polymorphisms and BAVM in a Chinese Han population. METHODS: We enrolled 150 patients with BAVMs confirmed by digital subtraction angiography (DSA) in the Department of Neurosurgery, Zhujiang Hospital, Southern Medical University from June 2017 to July 2019. Simultaneously, 150 patients without cerebrovascular disease were confirmed by computed tomography angiography/magnetic resonance angiography/DSA. DNA was extracted from peripheral blood and NOTCH4 genotypes were identified by PCR-ligase detection reaction. The χ2 test or Fisher's exact test was used to evaluate the differences in allele and genotype frequencies between the BAVM group, control group, bleeding group, and other complications. RESULTS: Two single-nucleotide polymorphisms (SNPs), rs443198 and rs438475, were significantly associated with BAVM. No SNP genotypes were significantly associated with hemorrhage or epilepsy. SNPs rs443198_AA-SNP and rs438475_AA-SNP may be associated with a lower risk of BAVM (p = 0.011, odds ratio (OR) = 0.459, 95% confidence interval (CI): 0.250-0.845; p = 0.033, OR = 0.759, 95% CI: 0.479-1.204). CONCLUSION: NOTCH4 gene polymorphisms were associated with BAVM and may be a risk factor in a Chinese Han population.
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Epilepsia , Malformaciones Arteriovenosas Intracraneales , Humanos , Polimorfismo de Nucleótido Simple , Pueblos del Este de Asia , Encéfalo/patología , Malformaciones Arteriovenosas Intracraneales/cirugía , Receptor Notch4/genéticaRESUMEN
Background: Tuberculosis (TB) is a serious public health problem to human health, but the pathogenesis of TB remains elusive. Methods: To identify novel candidate genes associated with TB susceptibility, we performed a population-based case control study to genotype 41SNPs spanning 21 genes in 435 pulmonary TB patients and 375 health donors from China. Results: We found Notch4 gene rs206018 and rs422951 polymorphisms were associated with susceptibility to pulmonary tuberculosis. The association was validated in another independent cohort including 790 TB patients and 1,190 healthy controls. Moreover, we identified that the rs206018 C allele was associated with higher level of Notch4 in PBMCs from pulmonary TB patients. Furthermore, Notch4 expression increased in TB patients and higher Notch4 expression correlated with the severer pulmonary TB. Finally, we explored the origin and signaling pathways involved in the regulation of Notch4 expression in response to Mycobacterium tuberculosis (Mtb) infection. We determine that Mtb induced Notch4 and its ligand Jagged1expression in macrophages, and Notch4 through TLR2/P38 signaling pathway and Jagged1 through TLR2/ERK signaling pathway. Conclusion: Our work further strengthens that Notch4 underlay an increased risk of TB in humans and is involved in the occurrence and development of TB, which could serve as a novel target for the host-targeted therapy of TB.
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Tuberculosis Pulmonar , Tuberculosis , Humanos , Receptor Toll-Like 2/genética , Estudios de Casos y Controles , Predisposición Genética a la Enfermedad , Polimorfismo de Nucleótido Simple , Tuberculosis Pulmonar/microbiología , Tuberculosis/genética , Expresión Génica , Receptor Notch4/genéticaRESUMEN
The Notch signaling pathway is an important therapeutic target for the treatment of inflammatory diseases and cancer. We previously created ligand-specific inhibitors of Notch signaling comprised of Fc fusions to specific EGF-like repeats of the Notch1 extracellular domain, called Notch decoys, which bound ligands, blocked Notch signaling, and showed anti-tumor activity with low toxicity. However, the study of their function depended on virally mediated expression, which precluded dosage control and limited clinical applicability. We have refined the decoy design to create peptibody-based Notch inhibitors comprising the core binding domains, EGF-like repeats 10-14, of either Notch1 or Notch4. These Notch peptibodies showed high secretion properties and production yields that were improved by nearly 100-fold compared to previous Notch decoys. Using surface plasmon resonance spectroscopy coupled with co-immunoprecipitation assays, we observed that Notch1 and Notch4 peptibodies demonstrate strong but distinct binding properties to Notch ligands DLL4 and JAG1. Both Notch1 and Notch4 peptibodies interfere with Notch signaling in endothelial cells and reduce expression of canonical Notch targets after treatment. While prior DLL4 inhibitors cause hyper-sprouting, the Notch1 peptibody reduced angiogenesis in a 3-dimensional in vitro sprouting assay. Administration of Notch1 peptibodies to neonate mice resulted in reduced radial outgrowth of retinal vasculature, confirming anti-angiogenic properties. We conclude that purified Notch peptibodies comprising EGF-like repeats 10-14 bind to both DLL4 and JAG1 ligands and exhibit anti-angiogenic properties. Based on their secretion profile, unique Notch inhibitory activities, and anti-angiogenic properties, Notch peptibodies present new opportunities for therapeutic Notch inhibition.
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Inhibidores de la Angiogénesis , Células Endoteliales , Receptor Notch1 , Receptor Notch4 , Animales , Ratones , Inhibidores de la Angiogénesis/genética , Inhibidores de la Angiogénesis/metabolismo , Inhibidores de la Angiogénesis/farmacología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Inmunoprecipitación , Inflamación/tratamiento farmacológico , Inflamación/genética , Inflamación/metabolismo , Ligandos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Receptor Notch1/antagonistas & inhibidores , Receptor Notch1/genética , Receptor Notch1/metabolismo , Receptor Notch4/genética , Receptor Notch4/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Vasos Retinianos/efectos de los fármacos , Resonancia por Plasmón de SuperficieRESUMEN
Background: NRAS wildtype melanoma accounts for approximately 80% of melanomas. Previous studies have shown that NRAS wildtype melanoma had higher response rates and better prognoses than NRAS-mutant patients following immunotherapy, while as major actors in tumor cells and tumor microenvironment (TME), the association between NOTCH family genes and response to immunotherapy in NRAS wildtype melanoma remains indistinct. Objective: We aim to explore whether NOTCH family gene variation is associated with genomic factors in immune checkpoint inhibitor (ICI) response in NRAS wildtype melanoma and with clinical results in these patients. Method: This research used genomic data of 265 NRAS wildtype ICI-pretreatment samples from five ICI-treated melanoma cohorts to analyze the relationship between NOTCH family gene mutation and the efficacy of ICI therapy. Results: NRAS wildtype melanomas with NOTCH4-Mut were identified to be associated with prolonged overall survival (OS) in both the discovery (HR: 0.30, 95% CI: 0.11-0.83, P = 0.01) and validation cohorts(HR: 0.21, 95% CI: 0.07-0.68, P = 0.003). Moreover, NOTCH4-Mut melanoma had a superior clinical response in the discovery cohort (ORR, 40.0% vs 13.11%, P = 0.057) and validation cohort (ORR, 68.75% vs 30.07%, P = 0.004). Further exploration found that NOTCH4-Mut tumors had higher tumor mutation burden (TMB) and tumor neoantigen burden (TNB) (P <0.05). NOTCH4-Mut tumors had a significantly increased mutation in the DNA damage response (DDR) pathway. Gene set enrichment analysis revealed NOTCH4-Mut tumor enhanced anti-tumor immunity. Conclusion: NOTCH4 mutation may promote tumor immunity and serve as a biomarker to predict good immune response in NRAS wildtype melanoma and guide immunotherapeutic responsiveness.
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GTP Fosfohidrolasas , Melanoma , Proteínas de la Membrana , Receptor Notch4 , Biomarcadores , GTP Fosfohidrolasas/genética , Humanos , Factores Inmunológicos/uso terapéutico , Inmunoterapia/métodos , Melanoma/tratamiento farmacológico , Melanoma/genética , Melanoma/terapia , Proteínas de la Membrana/genética , Mutación , Receptor Notch4/genética , Microambiente Tumoral/genéticaRESUMEN
The early onset of colorectal cancer (CRC) in individuals younger than 50 years is an emerging phenomenon, and obesity is a strong risk factor. Inflammatory mechanisms are mediated by immune cells, with macrophages and their phenotypical changes playing a significant role in CRC. Obesity-related hormones, such as leptin and adiponectin, affect macrophage polarization and cytokine expression. Macrophage metabolism, and therefore polarization, directly affects tumor progression and survival in patients with CRC. Altered obesity-related hormone levels induce phosphoinositide kinase-3 (PI3K)/serine-threonine-protein kinase (AKT) activation in colon cancer, causing increased cell survival, hyperplasia, and proliferation. Investigating the effects of obesity-related mechanisms on PI3K/Akt signaling can provide new insights for targeting mechanisms in CRC and obesity among the young. Central molecules for the control of cell proliferation, differentiation, and tumorigenesis within the gastrointestinal tract include downstream targets of the PI3K/AKT pathway, such as Neurogenic locus notch homolog 4 (Notch4) and GATA binding proteins (GATA). Leptin and adiponectin both alter gene expression within this pathway, thereby affecting TAM-mediated CRC progression. Our goal is to introduce the NOTCH4-GATA4-IRG1 axis as a link between inflammation and sporadic CRC and to discuss this pathway as a new potential immunotherapeutic target in individuals affected with obesity and early-onset CRC.
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Neoplasias Colorrectales , Leptina , Adiponectina/metabolismo , Edad de Inicio , Carboxiliasas/metabolismo , Línea Celular Tumoral , Proliferación Celular , Factor de Transcripción GATA4/genética , Factor de Transcripción GATA4/metabolismo , Regulación Neoplásica de la Expresión Génica , Humanos , Leptina/metabolismo , Obesidad/complicaciones , Obesidad/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/genética , Receptor Notch4/genética , Receptor Notch4/metabolismoRESUMEN
Endothelial cell (EC) sensing of disturbed blood flow triggers atherosclerosis, a disease of arteries that causes heart attack and stroke, through poorly defined mechanisms. The Notch pathway plays a central role in blood vessel growth and homeostasis, but its potential role in sensing of disturbed flow has not been previously studied. Here, we show using porcine and murine arteries and cultured human coronary artery EC that disturbed flow activates the JAG1-NOTCH4 signaling pathway. Light-sheet imaging revealed enrichment of JAG1 and NOTCH4 in EC of atherosclerotic plaques, and EC-specific genetic deletion of Jag1 (Jag1ECKO) demonstrated that Jag1 promotes atherosclerosis at sites of disturbed flow. Mechanistically, single-cell RNA sequencing in Jag1ECKO mice demonstrated that Jag1 suppresses subsets of ECs that proliferate and migrate. We conclude that JAG1-NOTCH4 sensing of disturbed flow enhances atherosclerosis susceptibility by regulating EC heterogeneity and that therapeutic targeting of this pathway may treat atherosclerosis.
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Aterosclerosis , Proteína Jagged-1 , Placa Aterosclerótica , Receptor Notch4 , Animales , Aterosclerosis/genética , Aterosclerosis/metabolismo , Vasos Coronarios/metabolismo , Células Endoteliales/metabolismo , Humanos , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Ratones , Placa Aterosclerótica/metabolismo , Receptor Notch4/genética , Receptor Notch4/metabolismo , Transducción de Señal , PorcinosRESUMEN
PURPOSE: Reliable biomarkers to predict the outcome and treatment response of estrogen receptor (ER)-negative breast cancer (BC) are urgently needed. Since immune-related signaling plays an important role in the tumorigenesis of ER-negative BC, we asked whether Notch genes, alone or in combination with other immune genes, can be used to predict the clinical outcome and immune checkpoint blockade (ICB) for this type of cancer. METHODS: We analyzed transcriptome data of 6918 BC samples from five independent cohorts, 81 xenograft triple-negative BC tumors that respond differently to ICB treatment and 754 samples of different cancer types from patients treated with ICB agents. RESULTS: We found that among four Notch genes, the expression levels of NOTCH1 and NOTCH4 were positively associated with recurrence of ER-negative BC, and that combined expression of these two genes (named Notch14) further enhanced this association, which was comparable with that of the Notch pathway signature. Analysis of 1182 immune-related genes revealed that the expression levels of most HLA genes, particularly HLA-DMA and -DRA, were reversely associated with recurrence in ER-negative BC with low, but not high Notch14 expression. A combined expression signature of NOTCH1, NOTCH4, HLA-DMA and HLA-DRA was more prognostic for ER-negative and triple-negative BCs than previously reported immune-related signatures. Furthermore, we found that the expression levels of these four genes were also synergistically associated with T cell exclusion score, infiltration of specific T cells and ICB efficacy in ER-negative BC, thereby providing a potential molecular mechanism for the synergistic effect of these genes on BC. CONCLUSIONS: Our data indicate that a gene signature composed of NOTCH1, NOTCH4, HLA-DMA and HLA-DRA may serve as a potential promising biomarker for predicting ICB therapy efficacy and recurrence in ER-negative/triple-negative BCs.
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Cadenas alfa de HLA-DR , Receptor Notch1 , Receptor Notch4 , Linfocitos T , Neoplasias de la Mama Triple Negativas , Biomarcadores de Tumor/biosíntesis , Biomarcadores de Tumor/genética , Femenino , Cadenas alfa de HLA-DR/biosíntesis , Cadenas alfa de HLA-DR/genética , Cadenas alfa de HLA-DR/inmunología , Humanos , Receptor Notch1/biosíntesis , Receptor Notch1/genética , Receptor Notch1/inmunología , Receptor Notch4/biosíntesis , Receptor Notch4/genética , Receptor Notch4/inmunología , Linfocitos T/inmunología , Linfocitos T/patología , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/inmunología , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
BACKGROUND: Hypoxic pulmonary hypertension (HPH) is a chronic progressive advanced disorder pathologically characterized by pulmonary vascular remodeling. Notch4 as a cell surface receptor is critical for vascular development. However, little is known about the role and mechanism of Notch4 in the development of hypoxic vascular remodeling. METHODS: Lung tissue samples were collected to detect the expression of Notch4 from patients with HPH and matched controls. Human pulmonary artery smooth muscle cells (HPASMCs) were cultured in hypoxic and normoxic conditions. Real-time quantitative PCR and western blotting were used to examine the mRNA and protein levels of Notch4. HPASMCs were transfected with small interference RNA (siRNA) against Notch4 or Notch4 overexpression plasmid, respectively. Cell viability, cell proliferation, apoptosis, and migration were assessed using Cell Counting Kit-8, Edu, Annexin-V/PI, and Transwell assay. The interaction between Notch4 and ERK, JNK, P38 MAPK were analyzed by co-immunoprecipitation. Adeno-associated virus 1-mediated siRNA against Notch4 (AAV1-si-Notch4) was injected into the airways of hypoxic rats. Right ventricular systolic pressure (RVSP), right ventricular hypertrophy and pulmonary vascular remodeling were evaluated. RESULTS: In this study, we demonstrate that Notch4 is highly expressed in the media of pulmonary vascular and is upregulated in lung tissues from patients with HPH and HPH rats compared with control groups. In vitro, hypoxia induces the high expression of Delta-4 and Notch4 in HPASMCs. The increased expression of Notch4 promotes HPASMCs proliferation and migration and inhibits cells apoptosis via ERK, JNK, P38 signaling pathways. Furthermore, co-immunoprecipitation result elucidates the interaction between Notch4 and ERK/JNK/P38. In vivo, silencing Notch4 partly abolished the increase in RVSP and pulmonary vascular remodeling caused by hypoxia in HPH rats. CONCLUSIONS: These findings reveal an important role of the Notch4-ERK/JNK/P38 MAPK axis in hypoxic pulmonary remodeling and provide a potential therapeutic target for patients with HPH.
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Regulación de la Expresión Génica , Hipertensión Pulmonar/genética , Hipoxia/complicaciones , Miocitos del Músculo Liso/metabolismo , Receptor Notch4/genética , Remodelación Vascular/genética , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Animales , Apoptosis , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/patología , Hipoxia/genética , Hipoxia/metabolismo , Sistema de Señalización de MAP Quinasas , Masculino , Miocitos del Músculo Liso/patología , Arteria Pulmonar/metabolismo , Arteria Pulmonar/patología , Arteria Pulmonar/fisiopatología , Ratas , Ratas Sprague-Dawley , Receptor Notch4/biosíntesis , Transducción de Señal , Regulación hacia Arriba , Proteínas Quinasas p38 Activadas por Mitógenos/biosíntesisRESUMEN
NOTCH4 is a member of the NOTCH family of receptors whose expression is intensively induced in macrophages after their activation by Toll-like receptors (TLR) and/or interferon-γ (IFN-γ). In this work, we show that this receptor acts as a negative regulator of macrophage activation by diminishing the expression of proinflammatory cytokines, such as IL-6 and IL-12, and costimulatory proteins, such as CD80 and CD86. We have observed that NOTCH4 inhibits IFN-γ signaling by interfering with STAT1-dependent transcription. Our results show that NOTCH4 reprograms the macrophage response to IFN-γ by favoring STAT3 versus STAT1 phosphorylation without affecting their expression levels. This lower activation of STAT1 results in diminished transcriptional activity and expression of STAT1-dependent genes, including IRF1, SOCS1 and CXCL10. In macrophages, NOTCH4 inhibits the canonical NOTCH signaling pathway induced by LPS; however, it can reverse the inhibition exerted by IFN-γ on NOTCH signaling, favoring the expression of NOTCH-target genes, such as Hes1. Indeed, HES1 seems to mediate, at least in part, the enhancement of STAT3 activation by NOTCH4. NOTCH4 also affects TLR signaling by interfering with NF-κB transcriptional activity. This effect could be mediated by the diminished activation of STAT1. These results provide new insights into the mechanisms by which NOTCH, TLR and IFN-γ signal pathways are integrated to modulate macrophage-specific effector functions and reveal NOTCH4 acting as a new regulatory element in the control of macrophage activation that could be used as a target for the treatment of pathologies caused by an excess of inflammation.
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Interferón gamma/metabolismo , Activación de Macrófagos/genética , Macrófagos Peritoneales/inmunología , Receptor Notch4/metabolismo , Transducción de Señal/genética , Receptor Toll-Like 4/metabolismo , Animales , Donantes de Sangre , Humanos , Lipopolisacáridos/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Monocitos/metabolismo , Células RAW 264.7 , Receptor Notch4/genética , Transducción de Señal/efectos de los fármacos , TransfecciónRESUMEN
Previous genome-wide association studies revealed multiple common variants involved in eczema but the role of rare variants remains to be elucidated. Here, we investigate the role of rare variants in eczema susceptibility. We meta-analyze 21 study populations including 20,016 eczema cases and 380,433 controls. Rare variants are imputed with high accuracy using large population-based reference panels. We identify rare exonic variants in DUSP1, NOTCH4, and SLC9A4 to be associated with eczema. In DUSP1 and NOTCH4 missense variants are predicted to impact conserved functional domains. In addition, five novel common variants at SATB1-AS1/KCNH8, TRIB1/LINC00861, ZBTB1, TBX21/OSBPL7, and CSF2RB are discovered. While genes prioritized based on rare variants are significantly up-regulated in the skin, common variants point to immune cell function. Over 20% of the single nucleotide variant-based heritability is attributable to rare and low-frequency variants. The identified rare/low-frequency variants located in functional protein domains point to promising targets for novel therapeutic approaches to eczema.
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Fosfatasa 1 de Especificidad Dual/genética , Eccema/diagnóstico , Eccema/genética , Receptor Notch4/genética , Intercambiadores de Sodio-Hidrógeno/genética , Subunidad beta Común de los Receptores de Citocinas , Fosfatasa 1 de Especificidad Dual/química , Fosfatasa 1 de Especificidad Dual/metabolismo , Expresión Génica , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Proteínas de Unión a la Región de Fijación a la Matriz , Polimorfismo de Nucleótido Simple , Enfermedades Raras/genética , Receptor Notch4/química , Receptor Notch4/metabolismo , Intercambiadores de Sodio-Hidrógeno/química , Intercambiadores de Sodio-Hidrógeno/metabolismoRESUMEN
Breast cancer (BC) is a heterogeneous disease and is one of the most common malignancy affecting women worldwide while colorectal cancer (CRC) is estimated to be the third common cancer and second leading cause of cancer related death globally. Both BC and CRC involve multiple genetic and epigenetic alterations in genes belonging to various signaling pathways including NOTCH that has been implicated in the development of these cancers. We investigated four single nucleotide polymorphisms, each in genes encoding NOTCH1-4 receptors for their role in susceptibility to breast and colorectal cancers in Saudi population. In this case-control study, TaqMan genotypic analysis of rs3124591 in NOTCH1 and rs3820041 in NOTCH4 did not exhibit association with breast as well as colorectal cancers. However, a strong association of rs11249433 which is in close proximity to NOTCH2 was observed with breast cancer susceptibility especially with those having an early onset of the disease. Interestingly, the rs1043994 located in NOTCH3 showed gender preference and was found to be significantly associated with colorectal cancers in males. Validation of these findings in bigger populations of different ethnicities may prove beneficial in identifying rs11249433 and rs1043994 as genetic screening markers for early detection of breast and colorectal carcinomas, respectively.
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Biomarcadores de Tumor/genética , Neoplasias de la Mama/patología , Neoplasias Colorrectales/patología , Receptor Notch1/genética , Receptor Notch2/genética , Receptor Notch3/genética , Receptor Notch4/genética , Neoplasias de la Mama/epidemiología , Neoplasias de la Mama/genética , Estudios de Casos y Controles , Neoplasias Colorrectales/epidemiología , Neoplasias Colorrectales/genética , Femenino , Estudios de Seguimiento , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , Pronóstico , Arabia Saudita/epidemiologíaRESUMEN
A cardinal feature of COVID-19 is lung inflammation and respiratory failure. In a prospective multi-country cohort of COVID-19 patients, we found that increased Notch4 expression on circulating regulatory T (Treg) cells was associated with disease severity, predicted mortality, and declined upon recovery. Deletion of Notch4 in Treg cells or therapy with anti-Notch4 antibodies in conventional and humanized mice normalized the dysregulated innate immunity and rescued disease morbidity and mortality induced by a synthetic analog of viral RNA or by influenza H1N1 virus. Mechanistically, Notch4 suppressed the induction by interleukin-18 of amphiregulin, a cytokine necessary for tissue repair. Protection by Notch4 inhibition was recapitulated by therapy with Amphiregulin and, reciprocally, abrogated by its antagonism. Amphiregulin declined in COVID-19 subjects as a function of disease severity and Notch4 expression. Thus, Notch4 expression on Treg cells dynamically restrains amphiregulin-dependent tissue repair to promote severe lung inflammation, with therapeutic implications for COVID-19 and related infections.
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Interacciones Huésped-Patógeno , Inmunidad Celular , Neumonía Viral/etiología , Neumonía Viral/metabolismo , Receptor Notch4/metabolismo , Transducción de Señal , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Anfirregulina/farmacología , Animales , Biomarcadores , Citocinas/metabolismo , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Interacciones Huésped-Patógeno/inmunología , Humanos , Inmunohistoquímica , Inmunomodulación/efectos de los fármacos , Mediadores de Inflamación/metabolismo , Virus de la Influenza A/fisiología , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/patología , Pulmón/virología , Ratones , Ratones Transgénicos , Neumonía Viral/patología , Receptor Notch4/antagonistas & inhibidores , Receptor Notch4/genética , Índice de Severidad de la EnfermedadRESUMEN
BACKGROUND AND AIMS: Direct-acting antivirals (DAAs) usually lead to improvement/remission of cryoglobulinemic vasculitis (CV), although symptoms may persist/recur after a sustained virological response (SVR). We evaluated hematological and genetic markers in patients with HCV-SVR vasculitis with and without persisting/recurring symptoms to early predict the CV outcome. APPROACH AND RESULTS: Ninety-eight patients with HCV-CV were prospectively enrolled after a DAA-induced SVR: Group A: 52 with complete clinical response; Group B: 46 with symptom maintenance/recurrence. Monoclonal B-cell lymphocytosis, t(14;18) translocation, and abnormal free light chains κ/λ ratios were detected by flow cytometry or nested-PCR or nephelometry in 4% Group A versus 17% Group B (P = 0.04) patients, 17% Group A versus 40% Group B patients (P = 0.02), and 17% Group A versus 47% Group B (P = 0.003) patients, respectively. At least 1 out of 3 clonality markers was altered/positive in 29% of Group A versus 70% of Group B patients (P < 0.0001). When available, pretherapy samples were also tested for t(14;18) translocation (detected in 12/37 [32%] Group A and 21/38 [55%] Group B) and κ/λ ratios (abnormal in 5/35 [14%] Group A and 20/38 [53%] Group B) (P = 0.0006), whereas at least one clonality marker was detected/altered in 16/37 (43%) Group A and 30/38 (79%) Group B (P = 0.002). CV-associated single-nucleotide polymorphisms were tested by real-time PCR. Among them, notch4 rs2071286 T minor allele and TT genotype showed a higher frequency in Group B versus Group A (46% vs. 29%, P = 0.01, and 17% vs. 2%, P = 0.006, respectively). CONCLUSIONS: Hematological or genetic analyses could be used to foresee the CV clinical response after DAA therapy and could be valuable to assess a rational flowchart to manage CV during follow-up.
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Antivirales/uso terapéutico , Crioglobulinemia/sangre , Hepatitis C Crónica/tratamiento farmacológico , Vasculitis/sangre , Anciano , Cromosomas Humanos Par 14/genética , Cromosomas Humanos Par 18/genética , Crioglobulinemia/genética , Femenino , Hepatitis C Crónica/sangre , Hepatitis C Crónica/genética , Humanos , Cadenas kappa de Inmunoglobulina/sangre , Cadenas lambda de Inmunoglobulina/sangre , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , Pronóstico , Receptor Notch4/genética , Recurrencia , Respuesta Virológica Sostenida , Translocación Genética , Vasculitis/genéticaRESUMEN
Several comprehensive studies have demonstrated that the NOTCH pathway is altered in a bimodal manner in head and neck squamous cell carcinoma (HNSCC). In a previous study, it was found that the NOTCH4/HEY1 pathway was specifically upregulated in HNSCC and promoted epithelialmesenchymal transition (EMT), and that HEY1 activation supported SOX2 expression. However, the interactions in this pathway have not yet been fully elucidated. The present study investigated the NOTCH4/HEY1/SOX2 axis in HNSCC using in vitro models and the Cancer Genome Atlas (TCGA) database. To explore the association, reporter and ChIP RTqPCR assays using SOX2overexpressing (SOX2OE) cells were performed. The association between NOTCH4 and HEY1 was examined in the same manner using HEY1overexpressing (HEY1OE) cells. The results of the in vitro experiments indicated that HEY1 promoted EMT in the HNSCC cells. Furthermore, the overexpression of HEY1 also promoted sphere formation and increased murine xenograft tumorigenicity. Reporter assays and ChIP RTqPCR experiments indicated that SOX2 regulated HEY1 expression via direct binding of the HEY1 promoter. HEY1 expression significantly correlated with SOX2 expression in primary lung SCC and other SCCs using the TCGA database. HEY1 also regulated NOTCH4 expression to create a positive reciprocal feedback loop. On the whole, the present study demonstrates that HEY1 expression in HNSCC is regulated via the promotion of SOX2 and promotes EMT. The NOTCH4/HEY1 pathway is specifically upregulated via a positive reciprocal feedback loop mediated by the HEY1medaited regulation of NOTCH4 transcription, and SOX2 correlates with HEY1 expression in SCC from other primary sites.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Proteínas de Ciclo Celular/genética , Transición Epitelial-Mesenquimal/genética , Receptor Notch4/genética , Factores de Transcripción SOXB1/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Conjuntos de Datos como Asunto , Retroalimentación Fisiológica , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Receptor Notch4/metabolismo , Transducción de Señal/genética , Esferoides Celulares , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Regulación hacia Arriba , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The association between NOTCH4 and schizophrenia has been repeatedly reported. However, the results from different genetic studies are inconsistent, and the role of NOTCH4 in schizophrenia pathogenesis remains unknown. Here, we provide convergent lines of evidence that support NOTCH4 as a schizophrenia risk gene. We first performed a meta-analysis and found that a genetic variant (rs2071287) in NOTCH4 was significantly associated with schizophrenia (a total of 125 848 subjects, p=8.31×10-17), with the same risk allele across all tested samples. Expression quantitative trait loci (eQTL) analysis showed that rs2071287 was significantly associated with NOTCH4 expression (p=1.08×10-14) in human brain tissues, suggesting that rs2071287 may confer schizophrenia risk through regulating NOTCH4 expression. Sherlock integrative analysis using a large-scale schizophrenia GWAS and eQTL data from human brain tissues further revealed that NOTCH4 was significantly associated with schizophrenia (p=4.03×10-7 in CMC dataset and p=3.06×10-6 in xQTL dataset), implying that genetic variants confer schizophrenia risk through modulating NOTCH4 expression. Consistently, we found that NOTCH4 was significantly downregulated in brains of schizophrenia patients compared with controls (p=2.53×10-3), further suggesting that dysregulation of NOTCH4 may have a role in schizophrenia. Finally, we showed that NOTCH4 regulates proliferation, self-renewal, differentiation and migration of neural stem cells, suggesting that NOTCH4 may confer schizophrenia risk through affecting neurodevelopment. Our study provides convergent lines of evidence that support the involvement of NOTCH4 in schizophrenia. In addition, our study also elucidates a possible mechanism for the role of NOTCH4 in schizophrenia pathogenesis.