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1.
Medicine (Baltimore) ; 99(33): e21355, 2020 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-32871987

RESUMO

RATIONALE: Coronary artery abnormalities are usually of major significance in clinical cardiology and cardiac surgery departments due to associated myocardial ischemia, myocardial infarction, and sudden cardiac death. Among them, anatomical malformations account for most coronary artery abnormalities. However, hypoplasia of the coronary artery is a rare type of coronary artery without any genetic screening information. PATIENT CONCERNS: A 10-year-old boy suffered severe chest pain, and a subsequent syncope occurred. DIAGNOSIS AND INTERVENTION: The boy complained of significant chest pain with syncope. Computerized tomography (CT) angiography scanning showed that the left coronary artery was dominated by abnormal origins and dramatically narrow artery lesions. Moreover, cardiac magnetic resonance imaging (MRI) confirmed myocardial ischemia. Cardiac catheterization confirmed that this was an extremely rare hypoplastic coronary case. Finally, a mutation was identified in NOTCH1 c.1023C>A for the first time. OUTCOMES: The boy was discharged after completing all examinations and was forbidden to play any kind of sport activity while waiting for heart transplantation. LESSONS: Hypoplastic coronary artery diseases have only been reported within very limited cases. This is the only report that has identified hypoplasia in 3 epicardial major coronary arteries. In addition, this is the first case to provide evidence between NOTCH1 genetic disorder and hypoplastic coronary artery disease in the clinic.


Assuntos
Anomalias dos Vasos Coronários/genética , Receptor Notch1/genética , Criança , Angiografia por Tomografia Computadorizada , Anomalias dos Vasos Coronários/diagnóstico por imagem , Anomalias dos Vasos Coronários/cirurgia , Transplante de Coração , Humanos , Imagem por Ressonância Magnética , Masculino , Mutação
2.
Arterioscler Thromb Vasc Biol ; 40(9): 2227-2243, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32640901

RESUMO

OBJECTIVE: Perivascular adipose tissue (PVAT) surrounding arteries supports healthy vascular function. During obesity, PVAT loses its vasoprotective effect. We study pathological conversion of PVAT, which involves molecular changes in protein profiles and functional changes in adipocytes. Approach and Results: C57BL6/J mice were fed a 60% high-fat diet for 12 weeks or a cardioprotective 30% calorie-restricted diet for 5 weeks. Proteomic analysis identified PVAT as a molecularly distinct adipose depot, and novel markers for thermogenic adipocytes, such as GRP75 (stress-70 protein, mitochondrial), were identified. High-fat diet increased the similarity of protein signatures in PVAT and brown adipose, suggesting activation of a conserved whitening pathway. The whitening phenotype was characterized by suppression of UCP1 (uncoupling protein 1) and increased lipid deposition, leptin, and inflammation, and specifically in PVAT, elevated Notch signaling. Conversely, PVAT from calorie-restricted mice had decreased Notch signaling and less lipid. Using the Adipoq-Cre strain, we constitutively activated Notch1 signaling in adipocytes, which phenocopied the changes in PVAT caused by a high-fat diet, even on a standard diet. Preadipocytes from mouse PVAT expressed Sca1, CD140a, Notch1, and Notch2, but not CD105, showing differences compared with preadipocytes from other depots. Inhibition of Notch signaling during differentiation of PVAT-derived preadipocytes reduced lipid deposition and adipocyte marker expression. CONCLUSIONS: PVAT shares features with other adipose depots, but has a unique protein signature that is regulated by dietary stress. Increased Notch signaling in PVAT is sufficient to initiate the pathological conversion of PVAT by promoting adipogenesis and lipid accumulation and may thus prime the microenvironment for vascular disease.


Assuntos
Adipócitos Brancos/metabolismo , Adipogenia , Tecido Adiposo Branco/metabolismo , Lipogênese , Obesidade/metabolismo , Receptores Notch/metabolismo , Adipócitos Brancos/patologia , Tecido Adiposo Branco/patologia , Adiposidade , Animais , Ataxina-1/metabolismo , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/patologia , Restrição Calórica , Dieta Hiperlipídica , Modelos Animais de Doenças , Endoglina/metabolismo , Feminino , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Obesidade/genética , Obesidade/patologia , Fenótipo , Pró-Proteína Convertase 9/genética , Pró-Proteína Convertase 9/metabolismo , Proteômica , Receptor Notch1/genética , Receptor Notch1/metabolismo , Receptor Notch2/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptores Notch/genética , Transdução de Sinais
3.
Anticancer Res ; 40(6): 3155-3161, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32487610

RESUMO

BACKGROUND/AIM: The deacetylase sirtuin1 (SIRT1) inhibits tumor suppressor p53 and may promote tumorigenesis; however, SIRT1 effects on leukemia cells are controversial. The aim of this study was to clarify the activity of SIRT1 in leukemia cells. MATERIALS AND METHODS: The effects of SIRT1 inhibition or activation and SIRT1 knockdown or overexpression were examined in two T cell acute lymphoblastic leukemia (T-ALL) cell lines carrying NOTCH1 mutations and three acute myeloid leukemia (AML) cell lines. RESULTS: The growth of T-ALL cells was promoted by SIRT1 inhibition and SIRT1 knockdown but was reduced by SIRT1 activation and overexpression; however, no effects were observed in AML cells. SIRT1 activation decreased NOTCH, NF-κB, and mTOR signaling and inhibited p53, suggesting that the possible mechanisms of T-ALL growth suppression by SIRT1 are independent of p53. CONCLUSION: SIRT1 activators acting through the down-regulation of NOTCH, NF-κB, and mTOR pathways can be novel targeted drugs for NOTCH1-mutated T-ALLs.


Assuntos
NF-kappa B/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Receptores Notch/metabolismo , Sirtuína 1/metabolismo , Carbazóis/farmacologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Humanos , Mutação , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Receptor Notch1/genética , Receptor Notch1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sirtuína 1/antagonistas & inibidores , Sirtuína 1/biossíntese , Sirtuína 1/genética , Serina-Treonina Quinases TOR/metabolismo , Transfecção
4.
NPJ Syst Biol Appl ; 6(1): 12, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32376854

RESUMO

The genomic platform that informs evolution of microRNA cascades remains unknown. Here we capitalised on the recent evolutionary trajectory of hominin-specific miRNA-4673, encoded in intron 4 of notch-1, to uncover the identity of one such precursor genomic element and the selective forces acting upon it. The miRNA targets genes that regulate Wnt/ß-catenin signalling cascade. Primary sequence of the microRNA and its target region in Wnt modulating genes evolved from homologous signatures mapped to homotypic cis-clusters recognised by TCF3/4 and TFAP2A/B/C families. Integration of homologous TFAP2A/B/C cis-clusters (short range inhibitor of ß-catenin) into the transcriptional landscape of Wnt cascade genes can reduce noise in gene expression. Probabilistic adoption of miRNA secondary structure by one such cis-signature in notch-1 reflected selection for superhelical curvature symmetry of precursor DNA to localise a nucleosome that overlapped the latter cis-cluster. By replicating the cis-cluster signature, non-random interactions of the miRNA with key Wnt modulator genes expanded the transcriptional noise buffering capacity via a coherent feed-forward loop mechanism. In consequence, an autonomous transcriptional noise dampener (the cis-cluster/nucleosome) evolved into a post-transcriptional one (the miRNA). The findings suggest a latent potential for remodelling of transcriptional landscape by miRNAs that capitalise on non-random distribution of genomic cis-signatures.


Assuntos
MicroRNAs/genética , MicroRNAs/metabolismo , Expressão Gênica , Redes Reguladoras de Genes , Genoma , Genômica , Humanos , Origem da Vida , Receptor Notch1/genética , Via de Sinalização Wnt/genética , beta Catenina
5.
BMC Med Genet ; 21(1): 93, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32375772

RESUMO

BACKGROUND: Pathogenic SLC6A1 variants have been reported in patients with myoclonic-atonic epilepsy (MAE). NOTCH1, encoding a member of the Notch family of proteins, is known to be associated with aortic valve disease. The PRIMPOL variant has only been identified in Chinese patients with high myopia. Exome sequencing analysis now allows the simultaneous detection of multiple genetic etiologies for patients with complicated clinical features. However, the presence of three Mendelian disorders in one patient supported by their respective pathogenic variants and clinical phenotypes is very rare. CASE PRESENTATION: Here, we report a 4-year-old Chinese boy who presented with MAE, delayed language, borderline intellectual disability (ID), mildly impaired social skills and attention deficit hyperactivity disorder (ADHD). He also had mild aortic valve stenosis and high myopia. Using whole-exome sequencing (WES), we identified three variants: (1) SLC6A1, NM_003042.4: c.881-883del (p.Phe294del), (2) NOTCH1, NM_017617.5:c.1100-2A > G and (3) PRIMPOL, NM_152683.4:c.265 T > G (p.Tyr89Asp). Parental Sanger sequencing confirmed that SLC6A1 and NOTCH1 variants were de novo, whereas the PRIMPOL variant was inherited from the father who also had high myopia. Furthermore, the PRIMPOL variant was absent from the genomes of the paternal grandparents, and thus was also a de novo event in the family. All three variants are classified as pathogenic. CONCLUSION: The SLC6A1 variant could explain the features of MAE, delayed language, borderline ID, impaired social skills and ADHD in this patient, whereas the features of aortic valve stenosis and high myopia of the patient may be explained by variants in NOTCH1 and PRIMPOL, respectively. This case demonstrated the utility of exome sequencing in uncovering the multiple pathogenic variants in a patient with complicated phenotypes due to the blending of three Mendelian disorders.


Assuntos
Epilepsias Mioclônicas/genética , Epilepsia Generalizada/genética , Predisposição Genética para Doença , Miopia/genética , Estenose da Valva Aórtica/genética , Estenose da Valva Aórtica/patologia , Pré-Escolar , DNA Primase/genética , DNA Polimerase Dirigida por DNA/genética , Epilepsias Mioclônicas/patologia , Epilepsia Generalizada/patologia , Proteínas da Membrana Plasmática de Transporte de GABA/genética , Testes Genéticos , Humanos , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Masculino , Enzimas Multifuncionais/genética , Mutação/genética , Miopia/patologia , Receptor Notch1/genética , Sequenciamento Completo do Exoma
6.
Nat Genet ; 52(6): 604-614, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32424351

RESUMO

During aging, progenitor cells acquire mutations, which may generate clones that colonize the surrounding tissue. By middle age, normal human tissues, including the esophageal epithelium (EE), become a patchwork of mutant clones. Despite their relevance for understanding aging and cancer, the processes that underpin mutational selection in normal tissues remain poorly understood. Here, we investigated this issue in the esophageal epithelium of mutagen-treated mice. Deep sequencing identified numerous mutant clones with multiple genes under positive selection, including Notch1, Notch2 and Trp53, which are also selected in human esophageal epithelium. Transgenic lineage tracing revealed strong clonal competition that evolved over time. Clone dynamics were consistent with a simple model in which the proliferative advantage conferred by positively selected mutations depends on the nature of the neighboring cells. When clones with similar competitive fitness collide, mutant cell fate reverts towards homeostasis, a constraint that explains how selection operates in normal-appearing epithelium.


Assuntos
Esôfago/citologia , Mutação , Proteína ADAM10/genética , Secretases da Proteína Precursora do Amiloide/genética , Animais , Linhagem da Célula , Dietilnitrosamina/toxicidade , Epitélio/efeitos dos fármacos , Epitélio/patologia , Epitélio/fisiologia , Esôfago/fisiologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Masculino , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor Notch1/genética , Receptor Notch2/genética , Reprodutibilidade dos Testes , Proteína Supressora de Tumor p53/genética
7.
Toxicol Lett ; 327: 19-31, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32234357

RESUMO

Traditional Chinese Medicines (TCMs)-containing aconitine are popular and indispensable home remedies in Asia for thousands of years due to its excellent pharmaceutical effects. Accumulating evidence has identified that repeated-dose of aconitine could cause polymorphic ventricular arrhythmias. However, underlying molecular mechanisms are still not fully understood. Hence, the present study firstly investigated the potential role of Notch1 signaling in aconitine-induced cardiotoxicity, aiming to elaborate possible molecular mechanisms involved in aconitine triggered ventricular arrhythmias. Our results showed that aconitine increased Notch1 signaling and downstream KDM5A expression in human and rat cardiomyocytes at non-detectable cytotoxic doses. Furthermore, aconitine promoted the formation of a new regulatory complex containing NICD and KDM5A in a CK2αHI regime, which then targeted to HCN4 promoter and induced re-expression of HCN4 in mature cardiomyocytes. Ultimately, HCN4-mediated If current contributed to aconitine-caused alterations in beating rate of rat cardiomyocytes. All changes aforementioned were significantly ameliorated by Notch1 inhibitor, suggesting that Notch1-mediated epigenetic regulation of HCN4 contributes to aconitine-induced ventricular myocardial dysrhythmia. Thus, our findings provide a novel toxic mechanism and position Notch1/NICD/KDM5A/HCN4 toxicity pathway as a potential target for the treatments of repeated-dose of medicine containing aconitine induced ventricular arrhythmias.


Assuntos
Aconitina/farmacologia , Arritmias Cardíacas/induzido quimicamente , Ventrículos do Coração/efeitos dos fármacos , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Canais de Potássio/metabolismo , Receptor Notch1/metabolismo , Animais , Animais Recém-Nascidos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Histonas , Humanos , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/genética , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Canais de Potássio/genética , Ratos , Receptor Notch1/genética , Superóxidos/metabolismo
8.
Nat Commun ; 11(1): 2018, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32332750

RESUMO

Gene regulation and metabolism are two fundamental processes that coordinate the self-renewal and differentiation of neural precursor cells (NPCs) in the developing mammalian brain. However, little is known about how metabolic signals instruct gene expression to control NPC homeostasis. Here, we show that methylglyoxal, a glycolytic intermediate metabolite, modulates Notch signalling to regulate NPC fate decision. We find that increased methylglyoxal suppresses the translation of Notch1 receptor mRNA in mouse and human NPCs, which is mediated by binding of the glycolytic enzyme GAPDH to an AU-rich region within Notch1 3'UTR. Interestingly, methylglyoxal inhibits the enzymatic activity of GAPDH and engages it as an RNA-binding protein to suppress Notch1 translation. Reducing GAPDH levels or restoring Notch signalling rescues methylglyoxal-induced NPC depletion and premature differentiation in the developing mouse cortex. Taken together, our data indicates that methylglyoxal couples the metabolic and translational control of Notch signalling to control NPC homeostasis.


Assuntos
Encéfalo/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Neurais/metabolismo , Aldeído Pirúvico/metabolismo , Receptor Notch1/metabolismo , Regiões 3' não Traduzidas , Animais , Encéfalo/citologia , Encéfalo/metabolismo , Diferenciação Celular , Linhagem Celular , Feminino , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Células HEK293 , Humanos , Camundongos , Neurogênese/genética , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Receptor Notch1/genética , Transdução de Sinais/genética
9.
Arterioscler Thromb Vasc Biol ; 40(6): e153-e165, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32295422

RESUMO

OBJECTIVE: Macrophages have been described in calcific aortic valve disease, but it is unclear if they promote or counteract calcification. We aimed to determine how macrophages are involved in calcification using the Notch1+/- model of calcific aortic valve disease. Approach and Results: Macrophages in wild-type and Notch1+/- murine aortic valves were characterized by flow cytometry. Macrophages in Notch1+/- aortic valves had increased expression of MHCII (major histocompatibility complex II). We then used bone marrow transplants to test if differences in Notch1+/- macrophages drive disease. Notch1+/- mice had increased valve thickness, macrophage infiltration, and proinflammatory macrophage maturation regardless of transplanted bone marrow genotype. In vitro approaches confirm that Notch1+/- aortic valve cells promote macrophage invasion as quantified by migration index and proinflammatory phenotypes as quantified by Ly6C and CCR2 positivity independent of macrophage genotype. Finally, we found that macrophage interaction with aortic valve cells promotes osteogenic, but not dystrophic, calcification and decreases abundance of the STAT3ß isoform. CONCLUSIONS: This study reveals that Notch1+/- aortic valve disease involves increased macrophage recruitment and maturation driven by altered aortic valve cell secretion, and that increased macrophage recruitment promotes osteogenic calcification and alters STAT3 splicing. Further investigation of STAT3 and macrophage-driven inflammation as therapeutic targets in calcific aortic valve disease is warranted.


Assuntos
Estenose da Valva Aórtica/patologia , Valva Aórtica/patologia , Calcinose/patologia , Macrófagos/fisiologia , Fator de Transcrição STAT3/fisiologia , Animais , Valva Aórtica/imunologia , Valva Aórtica/fisiopatologia , Estenose da Valva Aórtica/imunologia , Estenose da Valva Aórtica/fisiopatologia , Transplante de Medula Óssea , Calcinose/imunologia , Calcinose/fisiopatologia , Movimento Celular , Óxidos S-Cíclicos/farmacologia , Modelos Animais de Doenças , Expressão Gênica , Genótipo , Humanos , Inflamação/patologia , Macrófagos/química , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteogênese , Receptor Notch1/análise , Receptor Notch1/genética , Receptor Notch1/fisiologia , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética
10.
DNA Cell Biol ; 39(5): 783-789, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32129674

RESUMO

Adams-Oliver syndrome (AOS) is a rare hereditary disorder characterized by aplasia cutis congenita (ACC) and terminal transverse limb defects. The etiology of AOS has remained largely unknown, although mutations in the notch receptor 1 (NOTCH1) gene are most common genetic alteration associated with this disease. In this study, we aimed to identify the case of a 6-year-old boy, who presented with large ACC of the scalp and aortic valve stenosis, suggesting the possibility of AOS. Whole-exome sequencing identified a novel, de novo, in-frame deletion in the NOTCH1 gene (NOTCH1 c.1292_1294del, p.Asn431del) in the patient. The p.Asn431del variant was evaluated by several in silico analyses, which predicted that the mutant was likely to be pathogenic. In addition, molecular modeling with the PyMOL Molecular Graphics System suggested that the NOTCH1-N431del destabilizes calcium ion chelation, leading to decreased receptor-ligand binding efficiency. Quantitative reverse transcription PCR showed further significant downregulation of the Notch target genes, hes-related family bHLH transcription factor with YRPW motif 1 (HEY1) and hes family bHLH transcription factor 1 (HES1), suggesting that this mutation causes disease through dysregulation of the Notch signaling pathway. Our study provides evidence that the NOTCH1-N431del mutation is responsible for this case of AOS. To our knowledge, this is the first report of a patient with AOS caused by NOTCH1 mutation in Asia, and this information will be useful for providing the family with genetic counseling that can help to guide their future plans.


Assuntos
Displasia Ectodérmica/genética , Mutação da Fase de Leitura , Deformidades Congênitas dos Membros/genética , Receptor Notch1/genética , Dermatoses do Couro Cabeludo/congênito , Sequência de Aminoácidos , Animais , Sequência de Bases , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Criança , China , Humanos , Masculino , Modelos Moleculares , Conformação Proteica , Receptor Notch1/química , Proteínas Repressoras/genética , Dermatoses do Couro Cabeludo/genética , Fatores de Transcrição HES-1/genética
11.
World Neurosurg ; 138: e17-e25, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32142948

RESUMO

BACKGROUND: Eccrine carcinoma involvement of the central nervous system (CNS) is exceedingly rare. The prognosis and response to treatment of this pathology remain poorly characterized. METHODS: A retrospective case series and literature review were conducted. RESULTS: CNS-invading eccrine carcinoma was diagnosed in 3 patients (2 male and 1 female; age range, 60-79 years), including 2 cases of brain metastases and 1 case of brain-invading skull metastasis with subsequent spinal metastasis. The interval from primary tumor to CNS invasion was 18-51 months. All patients received multimodal therapy following diagnosis of CNS involvement. One patient who harbored a NOTCH1 mutation demonstrated a durable oncologic response after treatment with the immune checkpoint inhibitor pembrolizumab and lived 39 months after CNS invasion. The other 2 patients were discharged to hospice care within 1 month after the diagnosis of eccrine carcinoma brain metastasis. Including this case series, 23 cases of eccrine carcinoma invasion or metastasis to the CNS have been reported, with survival after diagnosis of CNS involvement ranging from a few weeks to 4 years. CONCLUSIONS: We present 3 cases of eccrine carcinoma metastatic to the CNS, including the first reported case to our knowledge of eccrine carcinoma treated with immunotherapy. This case, harboring a NOTCH1 mutation, demonstrated the longest durable oncologic response reported in this rare disease. Genomic and molecular testing may play increasingly important roles in the evaluation of these metastases.


Assuntos
Neoplasias Encefálicas/secundário , Carcinoma/secundário , Glândulas Écrinas , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias Cranianas/secundário , Neoplasias da Coluna Vertebral/secundário , Neoplasias das Glândulas Sudoríparas/patologia , Parede Abdominal , Idoso , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Carcinoma/tratamento farmacológico , Carcinoma/genética , Carcinoma Ductal/secundário , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Receptor Notch1/genética , Couro Cabeludo , Neoplasias Cranianas/diagnóstico por imagem , Neoplasias da Coluna Vertebral/diagnóstico por imagem
12.
Artigo em Chinês | MEDLINE | ID: mdl-32086902

RESUMO

Objective:To investigate the effect of Notch1 gene on radiosensitivity of nasopharyngeal carcinoma cells and its molecular mechanism. Method:A Notch1-knockout CNE-2 cell line was constructed using CRISPR/Cas9 system, and the expression of Notch1 gene was detected by RT-PCR and Western blot. After treatment with different doses of radiation, the survival fraction (SF) of each group was calculated, and used the GraphPad Prism 6.0 software and the Linear quadratic model were used to calculate the fitted dose survival curve and the sensitivity enhancement ratio(SER). Taking 6 Gy as radiation dose, the experiment was divided into four groups: Notch1(+) group, Notch1(-) group, IR+Notch1(+) group and IR+Notch1(-) group. CCK-8 assay was used to detect cell proliferation in each group. Annexin V-FITC/PI double staining assay was used to detect the changes of apoptosis in each group. The expression of H2AX, CyclinD1, Bax, Bcl-2 and GAPDH proteins were detected by Western blot. Result:The CNE-2 cell line with Notch1 gene knockout was successfully constructed. The clonogenic assay showed knockout of Notch1 enhanced the radiosensitivity of NPC cells. The CCK-8 assay showed that cell proliferation and cell viability were significantly reduced in the IR+Notch1(-) group compared with the IR+Notch1(+) group(P<0.05). Annexin V-FITC/PI double staining assay showed that the IR+Notch1(-) group had the highest apoptosis rate compared with the other groups (P<0.05). Western blotting demonstrated that the expression of γH2AX was significantly increased after irradiation of Notch1 nasopharyngeal carcinoma cells, the expression of Cyclin-D1 was increased, and the ratio of Bax:Bcl-2 was higher. Conclusion:Knockout of Notch1 signaling molecule can effectively improve the radiosensitivity of NPC cells cultured in vitro, which may be a potential target for radiosensitization of NPC.


Assuntos
Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genética , Tolerância a Radiação , Receptor Notch1/genética , Apoptose , Linhagem Celular Tumoral/efeitos da radiação , Proliferação de Células , Técnicas de Inativação de Genes , Humanos , Carcinoma Nasofaríngeo/radioterapia , Neoplasias Nasofaríngeas/radioterapia
13.
Eur J Cell Biol ; 99(2-3): 151070, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32005345

RESUMO

Metastasis is the main cause of cancer related deaths, and unfolding the molecular mechanisms underlying metastatic progression is critical for the development of novel therapeutic approaches. Notch is one of the key signaling pathways involved in breast tumorigenesis and metastasis. Notch activation induces pro-metastatic processes such as migration, invasion and epithelial to mesenchymal transition (EMT). However, molecular mediators working downstream of Notch in these processes are not fully elucidated. CYR61 is a secreted protein implicated in metastasis, and its inhibition by a monoclonal antibody suppresses metastasis in xenograft breast tumors, indicating the clinical importance of CYR61 targeting. Here, we aimed to investigate whether CYR61 works downstream of Notch in inducing pro-metastatic phenotypes in breast cells. We showed that CYR61 expression is positively regulated by Notch activity in breast cells. Notch1-induced migration, invasion and anchorage independent growth of a normal breast cell line, MCF10A, were abrogated by CYR61 silencing. Furthermore, upregulation of core EMT markers upon Notch1-activation was impaired in the absence of CYR61. However, reduced migration and invasion of highly metastatic cell line, MDA MB 231, cells upon Notch inhibition was not dependent on CYR61 downregulation. In conclusion, we showed that in normal breast cell line MCF10A, CYR61 is a mediator of Notch1-induced pro-metastatic phenotypes partly via induction of EMT. Our results imply CYR61 as a prominent therapeutic candidate for a subpopulation of breast tumors with high Notch activity.


Assuntos
Neoplasias da Mama/genética , Proteína Rica em Cisteína 61/genética , Receptor Notch1/genética , Neoplasias da Mama/patologia , Feminino , Humanos , Transdução de Sinais
14.
Nucleic Acids Res ; 48(7): 3496-3512, 2020 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-32107550

RESUMO

Aberrant Notch signaling plays a pivotal role in T-cell acute lymphoblastic leukemia (T-ALL) and chronic lymphocytic leukemia (CLL). Amplitude and duration of the Notch response is controlled by ubiquitin-dependent proteasomal degradation of the Notch1 intracellular domain (NICD1), a hallmark of the leukemogenic process. Here, we show that HDAC3 controls NICD1 acetylation levels directly affecting NICD1 protein stability. Either genetic loss-of-function of HDAC3 or nanomolar concentrations of HDAC inhibitor apicidin lead to downregulation of Notch target genes accompanied by a local reduction of histone acetylation. Importantly, an HDAC3-insensitive NICD1 mutant is more stable but biologically less active. Collectively, these data show a new HDAC3- and acetylation-dependent mechanism that may be exploited to treat Notch1-dependent leukemias.


Assuntos
Histona Desacetilases/metabolismo , Leucemia/metabolismo , Receptor Notch1/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Linhagem Celular Tumoral , Inibidores de Histona Desacetilases/farmacologia , Humanos , Leucemia/enzimologia , Lisina/metabolismo , Camundongos , Mutação , Peptídeos Cíclicos/farmacologia , Estabilidade Proteica , Receptor Notch1/química , Receptor Notch1/genética
15.
Nat Commun ; 11(1): 460, 2020 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-31974363

RESUMO

Recent interest in the control of bone metabolism has focused on a specialized subset of CD31hiendomucinhi vessels, which are reported to couple angiogenesis with osteogenesis. However, the underlying mechanisms that link these processes together remain largely undefined. Here we show that the zinc-finger transcription factor ZEB1 is predominantly expressed in CD31hiendomucinhi endothelium in human and mouse bone. Endothelial cell-specific deletion of ZEB1 in mice impairs CD31hiendomucinhi vessel formation in the bone, resulting in reduced osteogenesis. Mechanistically, ZEB1 deletion reduces histone acetylation on Dll4 and Notch1 promoters, thereby epigenetically suppressing Notch signaling, a critical pathway that controls bone angiogenesis and osteogenesis. ZEB1 expression in skeletal endothelium declines in osteoporotic mice and humans. Administration of Zeb1-packaged liposomes in osteoporotic mice restores impaired Notch activity in skeletal endothelium, thereby promoting angiogenesis-dependent osteogenesis and ameliorating bone loss. Pharmacological reversal of the low ZEB1/Notch signaling may exert therapeutic benefit in osteoporotic patients by promoting angiogenesis-dependent bone formation.


Assuntos
Neovascularização Fisiológica/fisiologia , Osteogênese/fisiologia , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/farmacologia , Idoso , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/farmacologia , Células Endoteliais/metabolismo , Epigênese Genética , Feminino , Humanos , Camundongos Knockout , Camundongos Transgênicos , Pessoa de Meia-Idade , Neovascularização Fisiológica/genética , Osteogênese/efeitos dos fármacos , Osteogênese/genética , Osteoporose/terapia , Ovariectomia , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Receptor Notch1/genética , Receptor Notch1/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
16.
Yonsei Med J ; 61(2): 186-191, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31997628

RESUMO

An outbreak of fatal humidifier disinfectant lung injuries (HDLI) occurred in Korea. Human studies on mechanisms underlying HDLI have yet to be conducted. This study aimed to investigate methylation changes and their potential role in HDLI after exposure to HDs containing polyhexamethylene guanidine-phosphate. DNA methylation analysis was performed in blood samples from 10 children with HDLI and 10 healthy children using Infinium Human MethylationEPIC BeadChip. Transcriptome analysis was performed using lung tissues from 5 children with HDLI and 5 controls. Compared to healthy controls, 92 hypo-methylated and 79 hyper-methylated CpG sites were identified in children with HDLI at the statistical significance level of |Δß|>0.2 and p<0.05. NOTCH1 was identified as a candidate network hub gene in cases. NOTCH1 transcripts significantly increased in lung tissues from HDLI cases compared to unexposed controls (p=0.05). NOTCH1 may play an important role in pulmonary fibrosis of HDLI.


Assuntos
Desinfetantes/efeitos adversos , Umidificadores , Lesão Pulmonar/induzido quimicamente , Lesão Pulmonar/genética , Receptor Notch1/metabolismo , Transdução de Sinais , Criança , Metilação de DNA/genética , Feminino , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Guanidinas , Humanos , Lesão Pulmonar/patologia , Masculino , Receptor Notch1/genética , República da Coreia/epidemiologia , Transdução de Sinais/genética
17.
Chem Biol Interact ; 316: 108938, 2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31926151

RESUMO

Eugenol a phenylpropanoid, predominantly found in clove is a very common spice in daily cuisine. It already reported to have anti-breast cancer activity. In this study, the effect of eugenol on CSC (Cancer Stem Cell) markers and its main regulator ß-catenin both in vivo Ehrlich Ascites Carcinoma (EAC) cell line and in vitro MCF-7 cell line was investigated with that of the untreated group. The therapeutic doses were found to significantly induce apoptosis leaving normal mice and cells unaffected. The in-depth analysis revealed the downregulation of ß-catenin thereby facilitating its degradation by N-terminal phosphorylation of Ser37 residue. Significant downregulation of various CSC markers was also observed in vivo after eugenol treatment those are regulated by the intracellular status of ß-catenin. These findings were validated by the effect of eugenol on the formation of the secondary sphere in vitro. Notable downregulation of the enriched stemness of secondary mammosphere was detected by the significantly decreased percentage of CD44+/CD24-/low population after eugenol treatment along with their distorted morphology and smaller the number of spheres. The underlying mechanism revealed significant downregulation of ß-catenin and the set of CSC markers along with their reduced mRNA expression in secondary sphere culture. Therefore, it can be concluded from the study that eugenol exerts its chemotherapeutic potential by impeding ß-catenin nuclear translocation thereby promoting its cytoplasmic degradation as a result stemness is being suppressed potentially even if in the enriched state. Therefore the study contributes to reduce the cancer-induced complications associated with the CSC population. This will ultimately confer the longer and improved patient's life.


Assuntos
Apoptose/efeitos dos fármacos , Eugenol/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , beta Catenina/metabolismo , Animais , Sítios de Ligação , Linhagem Celular Tumoral , Regulação para Baixo , Eugenol/química , Eugenol/uso terapêutico , Feminino , Humanos , Camundongos , Simulação de Acoplamento Molecular , Neoplasias/tratamento farmacológico , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/metabolismo , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Fosforilação/efeitos dos fármacos , Receptor Notch1/genética , Receptor Notch1/metabolismo , Transplante Heterólogo , beta Catenina/química
18.
Mol Cell Biochem ; 464(1-2): 181-191, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31758376

RESUMO

Notch is an evolutionarily conserved signaling pathway with an important role in development and cell fate determination. Deregulation of Notch signaling has been associated with several pathological conditions, including cancer. Acting as an oncogene in some types of cancers and as a tumor suppressor in other, Notch effects seem to be highly context-dependent in solid tumors. In the present study, we aimed to investigate gene expression levels of Notch pathway constituents, including ligands, receptors, and target genes, during the early stages of inflammation-associated intestinal carcinogenesis. To achieve so, we used our recently developed mouse model, in which colon cancer arises in the absence of urokinase-type plasminogen activator (uPA) due to colitis induced by dextran sodium sulfate (DSS) treatment. Among the cell surface components, ligands Jag1/Jag2 and receptors Notch1/Notch2 were found to be significantly upregulated in the uPA-deficient protumorigenic inflammatory microenvironment. Moreover, several intracellular Notch modulators, i.e. Hes1, Hey1, and Klf4, were also shown to be deregulated with inflammation, yet irrespective of uPA status. Sox9 transcription factor, however, was significantly downregulated in the uPA-deficient/DSS-treated mice that developed colon adenomas as compared to the wild-type/DSS-treated group with no neoplasia identified. The latter finding supports a tumor suppressive role of Sox9 in intestinal carcinogenesis. Our results point towards an early activation of Notch signaling pathway at the receptor-ligand level in inflammation-associated colon neoplasmatogenesis developed in the absence of uPA. Interestingly, such activation may not be accompanied by deregulation of downstream Notch-target genes, possibly due to the effects of other inter-related signaling pathways.


Assuntos
Neoplasias do Colo/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/metabolismo , Receptor Notch1/biossíntese , Receptor Notch2/biossíntese , Transdução de Sinais , Ativador de Plasminogênio Tipo Uroquinase/deficiência , Animais , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Camundongos , Camundongos Endogâmicos BALB C , Receptor Notch1/genética , Receptor Notch2/genética
19.
Cancer Lett ; 472: 70-80, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31874246

RESUMO

Liver cancer stem cells (LCSCs) initiate hepatocellular carcinoma (HCC) and contribute to its recurrence and treatment resistance. Studies have suggested ZBP-89 as a candidate tumor suppressor in HCC. We explored the role of ZBP-89 in the regulation of LCSCs. This study was performed in liver tissue samples from 104 HCC patients, 2 cell lines and mouse tumor models. We demonstrated that ZBP-89 was weakly expressed in LCSCs. Patients with high expression of LCSC markers displayed reduced survivals and higher recurrence rates after curative surgical operation. The expression of ZBP-89 was predictive for decreased recurrence. LCSC markers were negatively correlated with ZBP-89 in HCC tissues and in enriched liver tumor spheres. The exogenous expression of ZBP-89 attenuated the tumor-sphere formation and secondary colony formation capabilities of LCSCs in vitro and tumorigenicity in vivo. Furthermore, the negative effect of ZBP-89 on cancer stemness was Notch1-dependent. Localized with Notch1 intracellular domain (NICD1) in the nucleus, ZBP-89 repressed the Notch1 signaling pathway by competitive binding to NICD1 with MAML1. Collectively, ZBP-89 negatively regulates HCC stemness via inhibiting the Notch1 signaling.


Assuntos
Proteínas de Ligação a DNA/genética , Neoplasias Hepáticas/genética , Receptor Notch1/genética , Fatores de Transcrição/genética , Animais , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Autorrenovação Celular , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Transdução de Sinais/genética
20.
World J Gastroenterol ; 25(44): 6508-6526, 2019 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-31802831

RESUMO

BACKGROUND: Long noncoding RNAs (lncRNAs) are aberrant and play critical roles in gastric cancer (GC) progression and metastasis. Searching for coexpressed lncRNA clusters or representative biomarkers related to malignant phenotypes of GC may help to elucidate the mechanism of tumor development and predict the prognosis of GC. AIM: To investigate the prognostic value of NOTCH1 associated with lncRNA in T cell acute lymphoblastic leukemia 1 (NALT1) in GC and the mechanism of its involvement in GC invasion and metastasis. METHODS: RNA sequencing and corresponding clinical data were downloaded from The Cancer Genome Atlas database. The significance module was studied by weighted gene coexpression network analysis. A total of 336 clinical samples were included in the study. Gene silencing, reverse transcription polymerase chain reaction, western blotting, scrape motility assay, and Transwell migration assay were used to assess the function of hub-lncRNAs. RESULTS: At the transcriptome level, 3339 differentially expressed lncRNAs were obtained. weighted gene coexpression network analysis was used to obtain 15 lncRNA clusters and observe their coexpression. Pearson's correlation showed that blue module was correlated with tumor grade and survival. NALT1 was the hub-lncRNA of blue module and was an independent risk factor for GC prognosis. NALT1 was overexpressed in GC and its expression was closely related to invasion and metastasis. The mechanism may involve NALT1 regulation of NOTCH1, which is associated with lncRNA in T cell acute lymphoblastic leukemia, through cis regulation, thereby affecting the expression of the NOTCH signaling pathway. CONCLUSION: NALT1 is overexpressed and promotes invasion and metastasis of GC. The mechanism may be related to regulation of NOTCH1 by NALT1 and its effect on NOTCH signaling pathway expression.


Assuntos
Regulação Neoplásica da Expressão Gênica , RNA Longo não Codificante/metabolismo , Receptor Notch1/genética , Transdução de Sinais/genética , Neoplasias Gástricas/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Conjuntos de Dados como Assunto , Intervalo Livre de Doença , Feminino , Seguimentos , Gastrectomia , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Prognóstico , RNA-Seq , Receptor Notch1/metabolismo , Estômago/patologia , Estômago/cirurgia , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia , Análise de Sobrevida
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