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1.
Respir Res ; 22(1): 22, 2021 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-33468121

RESUMO

BACKGROUND: Increasing evidence shows that endothelial apoptosis contributes to cigarette smoke (CS)-induced disease progression, such as chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in CS-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from CS-induced apoptosis via regulating Notch1 signaling. METHODS: Human umbilical vein endothelial cells (HUVECs) were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24 h to explore the role of RESV in CSE induced endothelial apoptosis. 3-methyladenine (3-MA) or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD), γ-secretase inhibitor (DAPT) and Notch1 siRNA were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. RESULTS: Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT or Notch1 siRNA, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT or Notch1 siRNA induced apoptosis by activating Notch1 signaling. CONCLUSION: In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.


Assuntos
Apoptose/fisiologia , Autofagia/fisiologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Receptor Notch1/metabolismo , Resveratrol/farmacologia , Fumaça/efeitos adversos , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Fumar Cigarros/efeitos adversos , Fumar Cigarros/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
2.
Life Sci ; 266: 118911, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-33333049

RESUMO

AIM: Activation of transmembrane Notch-1 receptors through inflammatory cytokines is highly regulated by STAT-3 and NF-κB phosphorylation. Nimbolide (NMB) exhibits potent anti-inflammatory, anti-fibrotic, anticancer activities by targeting various pathways. Here, we have investigated the effect of NMB in regulation of STAT-3/NF-κB/Notch-1 axis in complete Freund's adjuvant (CFA) induced inflammatory arthritis (IA) model. MAIN METHODS: The anti-inflammatory and anti-arthritic activity of NMB was evaluated both in vitro (IL-1ß stimulated HIG-82 synovial fibroblasts) and in vivo (CFA induced rat model of IA) models. In vitro anti-arthritic activity was assessed by anti-migratory effect, while in vivo effects were evaluated through radiological and histological analysis. The effect of NMB on STAT-3, NF-κB, Notch-1 signaling pathways and proinflammatory cytokines were studied using western blot, immunohistochemistry and ELISA methods. Key findings NMB attenuated the migration of synovial fibroblasts in vitro. It reduced the progression of arthritis as evidenced from the improved radiological and histological abnormalities in arthritic rats. NMB significantly suppressed the nitrosooxidative stress and levels of pro-inflammatory cytokines. NMB also exhibited remarkable protective activity against upregulation of MAPK, STAT-3 and NF-κB phosphorylation mediated Notch-1 signaling pathway in synovial tissue of arthritic rats. SIGNIFICANCE: NMB may have clinical therapeutic value in rheumatoid arthritis by inhibiting STAT-3/NF-κB/Notch-1 axis and also by reducing the levels of proinflammatory cytokines.


Assuntos
Artrite Experimental/tratamento farmacológico , Adjuvante de Freund/toxicidade , Inflamação/tratamento farmacológico , Interleucina-1beta/toxicidade , Limoninas/farmacologia , Substâncias Protetoras/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Artrite Experimental/etiologia , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , NF-kappa B/genética , NF-kappa B/metabolismo , Fosforilação , Ratos , Ratos Wistar , Receptor Notch1/genética , Receptor Notch1/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
3.
Methods Mol Biol ; 2201: 27-34, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32975786

RESUMO

The interaction between neurons and glia is pivotal for the development of chronic opioid tolerance. One of the most important mechanisms of cell-to-cell interaction is the Notch signaling pathway. In this chapter we propose a double-immunofluorescence method to observe and quantify the colocalization of Notch-1 and mu-opioid receptor (MOR-1), using both neuronal and astrocyte markers.


Assuntos
Técnica Direta de Fluorescência para Anticorpo/métodos , Receptor Notch1/metabolismo , Receptores Opioides mu/metabolismo , Analgésicos Opioides/metabolismo , Animais , Tolerância a Medicamentos , Fluorescência , Humanos , Neurônios/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos
4.
Artigo em Japonês | MEDLINE | ID: mdl-33342936

RESUMO

Renal tubular cell death is caused by various extracellular stresses including toxic amounts of cadmium, an occupational and environmental pollutant metal, and is responsible for renal dysfunction. While cadmium exposure disrupts many intracellular signaling pathways, the molecular mechanism underlying cadmium-induced renal tubular cell death has not yet been fully elucidated. We have recently identified two important intracellular signaling pathways that promote cadmium-induced renal tubular cell death: the Notch1 signaling and activin receptor-like kinase (ALK) 4/5 signaling (also known as the activin-transforming growth factor ß receptor pathways). In this review paper, we introduce our previous experimental findings, focusing on Notch1 and ALK4/5 signaling pathways, which may uncover the molecular mechanisms involved in cadmium-induced renal tubular cell death.


Assuntos
Receptores de Ativinas Tipo I/metabolismo , Cádmio/toxicidade , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Poluentes Ambientais/toxicidade , Túbulos Renais/citologia , Túbulos Renais/patologia , Exposição Ocupacional/efeitos adversos , Receptor Notch1/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Animais , Humanos , Camundongos , Ratos
5.
PLoS One ; 15(12): e0244553, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33378407

RESUMO

Leydig cells represent the steroidogenic lineage of mammalian testis, which produces testosterone. Genetic evidence indicates the requirement of Notch signaling in maintaining a balance between differentiated Leydig cells and their progenitors during fetal development. In primary Leydig cells, Notch1 expression decreases with testicular development, while the expression of its ligand, Jagged1, remains relatively unchanged, suggesting that the roles of Jagged1 extend beyond Notch signaling. In addition, Jagged1 is known to be processed into its intracellular domain, which then translocate to the nucleus. In this study, we investigated the effect of Jagged1 intracellular domain (JICD) on steroidogenesis in Leydig cells. The independent overexpression of JICD in MA-10 Leydig cells was found to inhibit the activity of cAMP-induced Nur77 promoter. In addition, JICD suppressed Nur77 transactivation of the promoter of steroidogenic genes such as P450scc, P450c17, StAR, and 3ß-HSD. Further, adenovirus-mediated overexpression of JICD in primary Leydig cells repressed the expression of steroidogenic genes, consequently lowering testosterone production. These results collectively suggest that steroidogenesis in testicular Leydig cells, which is regulated by LH/cAMP signaling, is fine-tuned by Jagged1 during testis development.


Assuntos
Proteína Jagged-1/química , Proteína Jagged-1/genética , Células Intersticiais do Testículo/citologia , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Regiões Promotoras Genéticas , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Redes Reguladoras de Genes , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Domínios Proteicos , Transporte Proteico , Receptor Notch1/metabolismo , Transdução de Sinais , Esteroides/metabolismo
6.
Nat Commun ; 11(1): 5126, 2020 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-33046701

RESUMO

Cancer associated fibroblasts (CAFs) are a key component of the tumor microenvironment. Genomic alterations in these cells remain a point of contention. We report that CAFs from skin squamous cell carcinomas (SCCs) display chromosomal alterations, with heterogeneous NOTCH1 gene amplification and overexpression that also occur, to a lesser extent, in dermal fibroblasts of apparently unaffected skin. The fraction of the latter cells harboring NOTCH1 amplification is expanded by chronic UVA exposure, to which CAFs are resistant. The advantage conferred by NOTCH1 amplification and overexpression can be explained by NOTCH1 ability to block the DNA damage response (DDR) and ensuing growth arrest through suppression of ATM-FOXO3a association and downstream signaling cascade. In an orthotopic model of skin SCC, genetic or pharmacological inhibition of NOTCH1 activity suppresses cancer/stromal cells expansion. Here we show that NOTCH1 gene amplification and increased expression in CAFs are an attractive target for stroma-focused anti-cancer intervention.


Assuntos
Fibroblastos Associados a Câncer/metabolismo , Amplificação de Genes , Receptor Notch1/metabolismo , Neoplasias Cutâneas/metabolismo , Animais , Dano ao DNA , Feminino , Proteína Forkhead Box O3/genética , Proteína Forkhead Box O3/metabolismo , Humanos , Camundongos , Camundongos SCID , Receptor Notch1/genética , Pele/metabolismo , Neoplasias Cutâneas/genética
7.
Nat Commun ; 11(1): 5129, 2020 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-33046710

RESUMO

Zinc finger E-box binding homeobox 1 (Zeb1) has been demonstrated to participate in the acquisition of the properties of cancer stem cells (CSCs). However, it is largely unknown how signals from the tumor microenvironment (TME) contribute to aberrant Zeb1 expression. Here, we show that Zeb1 depletion suppresses stemness, colonization and the phenotypic plasticity of breast cancer. Moreover, we demonstrate that, with direct cell-cell contact, TME-derived endothelial cells provide the Notch ligand Jagged1 (Jag1) to neighboring breast CSCs, leading to Notch1-dependent upregulation of Zeb1. In turn, ectopic Zeb1 in tumor cells increases VEGFA production and reciprocally induces endothelial Jag1 in a paracrine manner. Depletion of Zeb1 disrupts this positive feedback loop in the tumor perivascular niche, which eventually lessens tumor initiation and progression in vivo and in vitro. In this work, we highlight that targeting the angiocrine Jag1-Notch1-Zeb1-VEGFA loop decreases breast cancer aggressiveness and thus enhances the efficacy of antiangiogenic therapy.


Assuntos
Neoplasias da Mama/metabolismo , Proteína Jagged-1/metabolismo , Células-Tronco Neoplásicas/metabolismo , Receptor Notch1/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Animais , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Feminino , Humanos , Proteína Jagged-1/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Fenótipo , Receptor Notch1/genética , Microambiente Tumoral , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
8.
Mol Pharmacol ; 98(5): 559-576, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32913140

RESUMO

The Notch family consists of four highly conserved transmembrane receptors. The release of the active intracellular domain requires the enzymatic activity of γ-secretase. Notch is involved in embryonic development and in many physiologic processes of normal cells, in which it regulates growth, apoptosis, and differentiation. Notch1, a member of the Notch family, is implicated in many types of cancer, including breast cancer (especially triple-negative breast cancer), leukemias, brain tumors, and many others. Notch1 is tightly connected to many signaling pathways that are therapeutically involved in tumorigenesis. Together, they impact apoptosis, proliferation, chemosensitivity, immune response, and the population of cancer stem cells. Notch1 inhibition can be achieved through various and diverse methods, the most common of which are the γ-secretase inhibitors, which produce a pan-Notch inhibition, or the use of Notch1 short interference RNA or Notch1 monoclonal antibodies, which produce a more specific blockade. Downregulation of Notch1 can be used alone or in combination with chemotherapy, which can achieve a synergistic effect and a decrease in chemoresistance. Targeting Notch1 in cancers that harbor high expression levels of Notch1 offers an addition to therapeutic strategies recruited for managing cancer. Considering available evidence, Notch1 offers a legitimate target that might be incorporated in future strategies for combating cancer. In this review, the possible clinical applications of Notch1 inhibition and the obstacles that hinder its clinical application are discussed. SIGNIFICANCE STATEMENT: Notch1 plays an important role in different types of cancer. Numerous approaches of Notch1 inhibition possess potential benefits in the management of various clinical aspects of cancer. The application of different Notch1 inhibition modalities faces many challenges.


Assuntos
Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Receptor Notch1/metabolismo , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Humanos , Transdução de Sinais/fisiologia
9.
Life Sci ; 258: 118154, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32735882

RESUMO

AIMS: Epithelial-to-mesenchymal transition (EMT) facilitates cell migration and invasion, and contributes to metastasis in bladder cancer. Within the perioperative period, anesthetic such as isoflurane have been found to affect cancer prognosis. In the study, we reported the tumor-promoting effect of isoflurane in bladder cancer. MATERIALS AND METHODS: Human bladder cancer cell lines T24 and BIU-87 were exposed to isoflurane at different concentrations. The immunofluorescent staining of Ki67, Annexin V-FITC/PI staining, Transwell invasion assays and wound-healing assays were performed to assess cell proliferation, apoptosis, invasion and migration. Expressions of EMT markers (E-cadherin, N-cadherin and Vimentin) and metastatic markers (Snail-1, Slug-1 and MMP-2/9) were determined by immunoblotting. Orthotopic tumor models and mice given tail vein injection of T24 cells were developed with or without 4-h exposure to 2% isoflurane. KEY FINDINGS: We found isoflurane promoted bladder cancer cell proliferation, invasion and migration but reduce apoptosis in a concentration-dependent manner. In addition, isoflurane was shown to increase HIF-1α and its nuclear accumulation in bladder cancer cells. HIF-1α knockdown inhibited bladder cancer cell proliferation and delayed EMT, which was reversed in the presence of 4-h exposure to 2% isoflurane. Likewise, we found isoflurane modulated ß-catenin/Notch1 pathways via HIF-1α. In vivo studies showed that isoflurane exposure accelerated formation of orthotopic bladder tumor and promoted hepatic metastases from carcinoma of the bladder. SIGNIFICANCE: Taken together, our study demonstrates that a frequently used anesthetic can exert a protumorigenic effect on bladder cancer. Isoflurane may serve as an important contributory factor to high recurrence following surgery.


Assuntos
Anestésicos Inalatórios/efeitos adversos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Isoflurano/efeitos adversos , Transdução de Sinais/efeitos dos fármacos , Neoplasias da Bexiga Urinária/induzido quimicamente , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Camundongos Endogâmicos C57BL , Invasividade Neoplásica/patologia , Metástase Neoplásica/patologia , Receptor Notch1/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia , beta Catenina/metabolismo
10.
Life Sci ; 261: 118306, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32828943

RESUMO

AIMS: Diabetic cardiomyopathy (DCM) is a common diabetes complication that can cause arrhythmia, heart failure, and even sudden death. Ranolazine is an antianginal agent used to treat chronic stable angina and has been demonstrated as an effective treatment for many cardiovascular diseases. However, the mechanism by which ranolazine alleviates DCM is unclear, motivating this study investigating the effects of ranolazine in DCM. MATERIALS AND METHODS: DCM rats were treated with one of three doses of ranolazine (10, 30, and 90 mg/kg/day) for 12 weeks. B-cell lymphoma 2 (Bcl-2), Bcl-2 associated X protein (Bax), cysteinyl aspartate specific proteinase-3 (Caspase-3), Notch homolog 1 (NOTCH1), and Neuregulin 1 (NRG1) expression was assayed using western blot and qRT-PCR. Cardiac changes were assayed using echocardiography, CT, HE staining, and Masson's trichrome staining. TUNEL staining and flow cytometry were used to detect cell apoptosis. NOTCH1 inhibitor (DAPT) was used to explore the mechanism of ranolazine. KEY FINDINGS: Compared with the DCM group, the ranolazine groups had no obvious weight loss and significantly decreased blood glucose levels. Ranolazine prevented diabetes-caused cardiac injury. Ranolazine also decreased the number of apoptotic cells and altered the expression of apoptosis-related mRNAs and proteins. Ranolazine-induced NOTCH1 activated NRG1 and inhibited the downstream apoptosis-related pathway, while DAPT partially inhibited ranolazine-induced NOTCH1 and NRG1 expression. SIGNIFICANCE: To our knowledge, this study is the first to demonstrate that ranolazine protects against DCM-induced apoptosis by activating the NOTCH1/NRG1 signaling pathway. Moreover, our study identified new mechanisms involved in DCM.


Assuntos
Fármacos Cardiovasculares/farmacologia , Diabetes Mellitus Experimental/tratamento farmacológico , Cardiomiopatias Diabéticas/prevenção & controle , Ranolazina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Glicemia/efeitos dos fármacos , Fármacos Cardiovasculares/administração & dosagem , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/fisiopatologia , Cardiomiopatias Diabéticas/fisiopatologia , Relação Dose-Resposta a Droga , Masculino , Neuregulina-1/metabolismo , Ranolazina/administração & dosagem , Ratos , Ratos Sprague-Dawley , Receptor Notch1/metabolismo , Transdução de Sinais/efeitos dos fármacos
11.
Mol Cell ; 79(2): 342-358.e12, 2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32645368

RESUMO

Short linear motifs (SLiMs) drive dynamic protein-protein interactions essential for signaling, but sequence degeneracy and low binding affinities make them difficult to identify. We harnessed unbiased systematic approaches for SLiM discovery to elucidate the regulatory network of calcineurin (CN)/PP2B, the Ca2+-activated phosphatase that recognizes LxVP and PxIxIT motifs. In vitro proteome-wide detection of CN-binding peptides, in vivo SLiM-dependent proximity labeling, and in silico modeling of motif determinants uncovered unanticipated CN interactors, including NOTCH1, which we establish as a CN substrate. Unexpectedly, CN shows SLiM-dependent proximity to centrosomal and nuclear pore complex (NPC) proteins-structures where Ca2+ signaling is largely uncharacterized. CN dephosphorylates human and yeast NPC proteins and promotes accumulation of a nuclear transport reporter, suggesting conserved NPC regulation by CN. The CN network assembled here provides a resource to investigate Ca2+ and CN signaling and demonstrates synergy between experimental and computational methods, establishing a blueprint for examining SLiM-based networks.


Assuntos
Calcineurina/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Transporte Ativo do Núcleo Celular , Motivos de Aminoácidos , Biotinilação , Centrossomo/metabolismo , Simulação por Computador , Células HEK293 , Células HeLa , Humanos , Espectrometria de Massas , Monoéster Fosfórico Hidrolases/química , Fosforilação , Mapas de Interação de Proteínas , Proteoma/metabolismo , Receptor Notch1/metabolismo , Saccharomyces cerevisiae , Proteínas de Saccharomyces cerevisiae/metabolismo , Transdução de Sinais
12.
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
13.
Nat Commun ; 11(1): 3084, 2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32555153

RESUMO

Tumor-initiating stem-like cells (TICs) are defective in maintaining asymmetric cell division and responsible for tumor recurrence. Cell-fate-determinant molecule NUMB-interacting protein (TBC1D15) is overexpressed and contributes to p53 degradation in TICs. Here we identify TBC1D15-mediated oncogenic mechanisms and tested the tumorigenic roles of TBC1D15 in vivo. We examined hepatocellular carcinoma (HCC) development in alcohol Western diet-fed hepatitis C virus NS5A Tg mice with hepatocyte-specific TBC1D15 deficiency or expression of non-phosphorylatable NUMB mutations. Liver-specific TBC1D15 deficiency or non-p-NUMB expression reduced TIC numbers and HCC development. TBC1D15-NuMA1 association impaired asymmetric division machinery by hijacking NuMA from LGN binding, thereby favoring TIC self-renewal. TBC1D15-NOTCH1 interaction activated and stabilized NOTCH1 which upregulated transcription of NANOG essential for TIC expansion. TBC1D15 activated three novel oncogenic pathways to promote self-renewal, p53 loss, and Nanog transcription in TICs. Thus, this central regulator could serve as a potential therapeutic target for treatment of HCC.


Assuntos
Proteínas Ativadoras de GTPase/metabolismo , Células-Tronco Neoplásicas/citologia , Receptor Notch1/metabolismo , Adulto , Idoso , Animais , Carcinogênese/patologia , Carcinoma Hepatocelular/metabolismo , Divisão Celular , Linhagem Celular Tumoral , Transferência Ressonante de Energia de Fluorescência , Hepacivirus , Hepatócitos/citologia , Humanos , Fígado/metabolismo , Neoplasias Hepáticas/metabolismo , Camundongos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Fosforilação , Receptores Notch/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo
14.
Vascul Pharmacol ; 131: 106763, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32593718

RESUMO

Atherosclerosis (AS) is one of the most common cardiovascular events in patients with chronic renal insufficiency (CRI). During the development of CRI, uremic toxins, including indoxyl sulfate (IS), are pivotal risk factors for AS. However, the underlying mechanism between AS and IS has not been fully elucidated. The present study was designed to test our hypothesis that IS promotes the AS by regulating viability, proliferation, migration and apoptosis of endothelial cells and vascular smooth muscle cells. In this present study, our date showed that IS inhibited the cell viability of human umbilical vein endothelial cells (HUVECs) and human aortic vascular smooth muscle cells (HA-VSMCs) in a dose-dependent manner (P < .05). Moreover, IS inhibited the proliferation, migration and induced apoptosis of HUVECs and HA-VSMCs significantly (P < .05). However, inhibition of the miR-34a abolished these effects of IS in vitro, indicating that miR-34a is involved in the development of AS induced by IS. In addition, the luciferase reporter gene assay showed that up-regulating of miR-34a inhibited the Notch1 transcriptional activity remarkably (P < .05). The expression of Notch1 decreased after IS treatment, while miR-34a inhibitor attenuated this effect. Moreover, the expression of miR-34a-related proteins Wnt-1, Jag1, E2F1 and SIRT1 decreased, while the expression of p53 increased in HUVECs and HA-VSMCs after IS treatment. Consistently, blockage of miR-34a abolished the remarkable effects on protein expressions induced by IS. Taken together, this study showed that IS can inhibit the proliferation, migration and promote apoptosis of HUVECs and HA-VSMCs through the Notch1 signal and miR-34a-related proteins by up-regulating miR-34a. These findings may provide new insights into the underlying mechanism of AS in CRI.


Assuntos
Apoptose/efeitos dos fármacos , Aterosclerose/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Indicã/toxicidade , MicroRNAs/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Aterosclerose/genética , Aterosclerose/patologia , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , MicroRNAs/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Receptor Notch1/genética , Receptor Notch1/metabolismo , Transdução de Sinais , Regulação para Cima
15.
Nat Commun ; 11(1): 3256, 2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32591500

RESUMO

BRCA1 mutation carriers have a higher risk of developing triple-negative breast cancer (TNBC), which is a refractory disease due to its non-responsiveness to current clinical targeted therapies. Using the Sleeping Beauty transposon system in Brca1-deficient mice, we identified 169 putative cancer drivers, among which Notch1 is a top candidate for accelerating TNBC by promoting the epithelial-mesenchymal transition (EMT) and regulating the cell cycle. Activation of NOTCH1 suppresses mitotic catastrophe caused by BRCA1 deficiency by restoring S/G2 and G2/M cell cycle checkpoints, which may through activation of ATR-CHK1 signalling pathway. Consistently, analysis of human breast cancer tissue demonstrates NOTCH1 is highly expressed in TNBCs, and the activated form of NOTCH1 correlates positively with increased phosphorylation of ATR. Additionally, we demonstrate that inhibition of the NOTCH1-ATR-CHK1 cascade together with cisplatin synergistically kills TNBC by targeting the cell cycle checkpoint, DNA damage and EMT, providing a potent clinical option for this fatal disease.


Assuntos
Proteína BRCA1/deficiência , Carcinogênese/patologia , Receptor Notch1/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Alelos , Animais , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteína BRCA1/metabolismo , Morte Celular , Linhagem Celular Tumoral , Quinase 1 do Ponto de Checagem/metabolismo , Elementos de DNA Transponíveis/genética , Progressão da Doença , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Knockout , Mitose , Mutação/genética , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/genética
16.
Life Sci ; 257: 118010, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32598932

RESUMO

Podocyte injury is an early event and core in the development of focal segmental glomerular sclerosis (FSGS) that induces poor prognosis. Epithelial-mesenchymal transition (EMT) as a response of podocyte to injury leads to podocyte depletion and proteinuria. The abnormally reactivated NOTCH pathway may be involved in podocyte EMT. Baicalin, as a natural flavonoid compound, had significant inhibitory activity on tissue fibrosis and tumor cell invasion. However, its potential role and molecular mechanisms to injured podocyte in FSGS are little known. Here we found that baicalin could inhibit podocyte EMT markers expression and cell migration induced by TGF-ß1, accompanied by the up-regulated expression of slit diaphragm (SD) proteins and cell-cell adhesion molecule. Further investigation revealed that EMT inhibition of baicalin on injured podocyte is mainly mediated by the reduction of notch1 activation and its downstream Snail expression. Using the adriamycin-induced FSGS model, we determined that baicalin suppresses the Notch1-Snail axis activation in podocytes, relieves glomerulus structural disruption and dysfunction, and reduces proteinuria. Altogether, these findings suggest that baicalin is a novel renoprotective agent against podocyte EMT in FSGS and indicate its underlying mechanism that involves in negative regulation of the Notch1-Snail axis.


Assuntos
Flavonoides/farmacologia , Glomerulosclerose Segmentar e Focal/tratamento farmacológico , Proteinúria/tratamento farmacológico , Animais , Biomarcadores/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Doxorrubicina/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Flavonoides/metabolismo , Glomerulosclerose Segmentar e Focal/metabolismo , Glomérulos Renais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Podócitos/metabolismo , Proteinúria/metabolismo , Receptor Notch1/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
17.
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
18.
Toxicol Appl Pharmacol ; 396: 115001, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32277947

RESUMO

Mebendazole (MBZ) is a tubulin-suppressive antihelmintic agent with low toxicity, which has been repurposed to treat different types of tumors. Chemoresistance is quite common in refractory or relapsed T cell acute lymphoblastic leukemia (T-ALL), which leads to dismal chances of recovery. In this study, MBZ was found to suppress the proliferation and reduce the viability of T-ALL cell line, CCRF-CEM, and its chemoresistant derivative, CEM/C1, at nanomolar concentrations. The inhibitive effects were found to be dose-dependent and not to be affected by the chemoresistance of CEM/C1 cells. Cell cycle arrest, caspase 3/7 activation and tubulin disruption were found in the MBZ-treated T-ALL cells. Notch1 signaling, which is often aberrantly activated in T-ALL cells, was showed to be suppressed by MBZ treatments. MBZ administration in murine T-ALL models also suppressed the growth of CEM/C1 cells, indicating that MBZ may be developed as a therapeutic agent for chemoresistant T-ALLs. The mRNA levels of the Notch1 and Hes1 were also confirmed to be suppressed by MBZ in vivo, which was consistent with the in vitro observations. This study demonstrated, for the first time, that MBZ could inhibit chemoresistant T-ALL cells both in vitro and in vivo, and the Notch1 signaling pathway was suppressed by MBZ treatment.


Assuntos
Antineoplásicos/uso terapêutico , Mebendazol/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Moduladores de Tubulina/uso terapêutico , Animais , Western Blotting , Caspase 3/metabolismo , Caspase 7/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Neoplasias , Receptor Notch1/metabolismo , Tubulina (Proteína)/metabolismo
19.
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
20.
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
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